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Overview

The Department of Cellular and Molecular Biology


is scaffolded by a multidisciplinary scientific
project focused on the role that the cell, and
the way it functions at the molecular level, plays
as core element of Biology. The Department
understands as Cell Biology not only the study of
the cell as isolated entity but also its integration
into tissues and into the development of organs
and organisms. To that aim it involves research
groups interested in a variety of model systems,
including microbes, be they prokaryotic or
eukaryotic, invertebrates and vertebrates. These
groups exploit a range of methodologies, such
as genetics, physiology, molecular biology and
biochemistry, biophysics, optical microscopy and
omics. In addition, it includes groups decidedly
exploring bottom-up synthetic approaches to
reconstruct minimal cytomimetic systems by
means of the controlled assembly of proteins or
nucleic acids in defined structures, thus allowing
their integration into lipid vesicles and other
cytomimetic compartments. These objectives
have clear implications for Nanotechnology and
Biotechnology.
Miguel ngel Pealva
Department Head

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Biologa Celular
y Molecular
Cellular & Molecular Biology

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Miguel ngel Pealva Soto Eduardo Antonio Espeso Fernndez


Biologa Molecular y Celular de Aspergillus | Aspergillus Molecular and Cellular Biology

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Germn Rivas Caballero Carlos Alfonso Botello, Mercedes Jimnez Sarmiento y Silvia Zorrilla Lpez
Bioqumica de Sistemas de la Divisin Bacteriana | Systems Biochemistry of Bacterial Division

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Rafael Giraldo Surez M. Elena Fernndez-Tresguerres Rodrguez-Vigil y Juan Francisco Gimnez Abin
Ensamblajes Macromoleculares Microbianos Sintticos | Synthetic Microbial Macromolecular Assemblies

108

Jorge Bernardo Schvartzman Blinder Dora Beatriz Krimer Smunis y Pablo Hernndez Valenzuela
Biologa Molecular de los Cromosomas | Molecular Biology of the Chromosomes

110

Miguel Angel Vidal Caballero


El Sistema Polycomb de Regulacin Epigentica | Epigenetic Control by the Polycomb Group of Genes

112

Patricia Boya
Funciones de la Autofagia en la Fisiopatologa de los Organismos | Roles of Autophagy in Health and Disease

114

Jess del Mazo Martnez


Biologa Molecular de la Gametognesis | Molecular Biology of Gametogenesis

116

Rosa Mara Lozano Puerto Blanca Prez-Maceda


Reconocimiento Clula-Biomaterial | Cell-Biomaterial Recognition

118

Susana Moreno Daz de la Espina


Matriz Nuclear y Regulacin de la Organizacin y Funcionalidad Nuclear
Nuclear Matrix and Regulation of the Nuclear Organization and Function

120

Jos Luis Barbero Esteban Lucas Snchez Rodrguez


Dinmica Cromosmica en Meiosis | Chromosomal Dynamics in Meiosis

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Biologa Celular y Molecular | Cellular & Molecular Biology

Miguel ngel Pealva Soto

Eduardo Antonio Espeso Fdez.

Profesor de Investigacin
penalva@cib.csic.es

Cientfico Titular
eespeso@cib.csic.es
PhD, 1982
Universidad Autnoma de Madrid
Postdoctoral
Antibiticos SA (Madrid)
Institut de Genetique et Microbiologie,
Universidad de Paris (Orsay, Paris)
Cientfico Titular, 1987
Jefe de Grupo, 1987
Profesor de Investigacin, 2001
CIB, CSIC
Visiting Scientist, 2005-2006
MRC Laboratory of Molecular Biology
(Cambridge, UK)
Elegido miembro, 2000
EMBO

PhD, 1989
Universidad Complutense de Madrid
Postdoctoral, 1997-1999
Imperial College London

EMBO-Postdoctoral Fellow
Contratado, 2001-2004
Ramn y Cajal
Cientfico Titular, 2004
Jefe de Grupo, 2004
CIB, CSIC

Secretario, 2004-2008
Grupo Especializado de Hongos Filamentosos
y Levaduras (SEM)

Otros miembros | Other lab members:


Herbert N. Arst (Ad honorem)
Elena Reoyo Hernndez
Areti Pantazopoulou
Mario Pinar Sala
Manuel Snchez Lpez-Berges

Maria Villarino Prez


Victor Garca Tagua
Laura Mellado Maroas
Daniel Lucena Agell
Patricia Hernndez Ortiz

Maria-Tsampika Manoli
Miguel Hernndez Gonzlez
http://www.cib.csic.es/es/grupo.php?idgrupo=8

Biologa Molecular y Celular


de Aspergillus
Aspergillus nidulans es un modelo gentico apropiado para estudiar exocytosis polarizada y transporte a larga
distancia por microtbulos y actina. Su trfico intracellular se asemeja al de metazoos, pero el hongo es haploide,
genticamente manipulable y conveniente para microscopa.

ediante la combinacin de abordajes genticos y bioqumicos


con microscopa multidimensional in vivo, estudiamos la organizacin y la dinmica del Golgi y del sistema endovacuolar,
centrndonos en GTPasas RAB y ARF, sus reguladores y sus efectores.
El Golgi de Aspergillus est formado por cisternas dispersas que pueden resolverse por microscopa ptica. Pretendemos comprender los
mecanismos de maduracin de cisternas del Golgi y especcamente
la biognesis de carriers post-Golgi en el TGN, as como las diferentes
rutas por las que membrana y cargo salen del ER. Nuestro trabajo tiene
importantes implicaciones tanto en medicina como en agricultura (la
patogenicidad de los hongos hacia humanos y plantas dependen estrictamente de la exocitosis y los hongos son sensibles a ciertas drogas
antitumorales) y tambin en el campo de la biotecnologa, dado que una
parte substancial del portafolio de enzimas industriales se fabrica con
especies de Aspergillus como factoras celulares.
Muchas rutas biosintticas y catablicas estan sujetas a regulacin
transcripcional. Estudiamos las seales, los receptores, la transduccin
de la seal y los mecanismos que modican tanto las actividades como
la localizacin celular de factores de transcripcin. En los eucariotas
el transporte de los factores transcripcionales al interior nuclear es un
punto clave en la regulacin de su actividad. Usando como modelo
diferentes factores nucleares queremos entender los mecanismos de
sealizacin y transporte entre citoplasma y ncleo en un organismo
con organizacin celular cenoctica (multinucleado). En especial nos
centramos en aquellos que median en la respuesta al estrs por cationes
y la alcalinidad como son los factores con dedos de zinc SltA y CrzA.
El estudio de estos reguladores pemite abordar la sealizacin mediada
por calcio/calcineurina, analizar la protelisis como mecanismo de activacin postraduccional y el papel de la tolerancia al estrs en procesos
de virulencia fngica.

Figura 1 | Figure 1
Portada del nmero de Julio de la revista Autophagy, por el artculo de Pinar et al.
que demuestra que las membranas de la ruta de autofagia en hongos derivan de
estructuras asociadas con el ER que se asemejan a los omegasomas de metazoos.
Cover of the July 2013 issue of the journal Autophagy, for the article by Pinar et al. showing
that fungal autophagic membranes derive from ER-associated srtuctures resembling
metazoan omegasomes.

Financiacin | Funding
BIO2012-30695 (MINECO)

S2010/BMD-2414 (Comunidad de Madrid)


IPT-2011-0752-900000 (MINECO)
BFU2012-33142 (MINECO)

RD12/0018/0007 (ISCIII-FEDER)

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Biologa Celular y Molecular | Cellular & Molecular Biology

Aspergillus Molecular
and Cellular Biology
Aspergillus nidulans is a genetic model well suited
for studying polarised exocytosis and long-distance
transport mediated by actin and microtubules.
Intracellular traffic resembles that of metazoan cells,
yet the organism is haploid, genetically amenable
and microscopy-friendly.

y combining genetic and biochemical approaches with in vivo


multidimensional microscopy, we are studying the organization and
dynamics of the Golgi and endovacuolar systems, focusing on RAB
and ARF GTPases, their regulators and their eectors. The Aspergillus
Golgi is formed by non-stacked early and late Golgi cisternae that can
be resolved by optical microscopy. We are studying the mechanisms of
cisternal maturation in the Golgi, and specically the mechanisms that
determine the biogenesis of post-Golgi carriers in the TGN, as well as the
dierent pathways for the exit of membrane and cargo from the endoplamic reticulum. Our work has important implications for both medicine and
agriculture (fungal pathogenicity to plants and humans is strictly dependent
on exocytosis and fungal cells are sensitive to certain anti-tumour drugs)
and major ones for biotechnology, as a substantial share of the industrial
enzyme catalogue is produced with Aspergillus species as cell factories.
Most biosynthetic and catalytic pathways are transcriptionally regulated.
We study signals, receptors, signaling transduction and the mechanisms
behind the activation and cellular localisation of transcription factors. In
eukaryotes, nuclear transport is a key regulatory step in the regulation of a
transcription factor activity. Using as models diverse nuclear factors we try
to understand the mechanisms involved in signaling and track between
cytoplasm and nucleus in a coenocytic (multi nuclear) organism. Specically
we focus on the zinc-nger transcription factors SltA and CrzA that mediate
in the responses to cation and alkaline pH stresses. Studying these regulators allow us to investigate the calcium-calcineurin mediated signaling,
proteolysis as a mechanism of posttranslational activation and the role of
stress tolerance in fungal virulence.

Publicaciones Seleccionadas
Selected Publications
Pinar, M, A Pantazopoulou & MA Pealva [2013] Live-cell imaging of Aspergillus
nidulans autophagy: RAB1 dependence, Golgi independence and ER involvement.
Autophagy 9: 1-20. (Journal Cover).
Pinar, M, Pantazopoulou, A, Arst, HN, Jr, and Pealva, MA [2013] Acute inactivation
of the Aspergillus nidulans Golgi membrane fusion machinery: correlation
of apical extension arrest and tip swelling with cisternal disorganization. Mol.
Microbiol. 89: 228-248. (Editorial Minireview).

Etxebeste O, Villarino M, Markina-Iarrairaegui A, Arajo-Bazn L, Espeso EA. [2013]


Cytoplasmic dynamics of the general nuclear import machinery in apically growing
syncytial cells. PLoS One 8(12):e85076.
Shantappa S, Dhingra S, Hernndez-Ortiz P, Espeso EA, Calvo AM. [2013] Role of
the zinc finger transcription factor SltA in morphogenesis and sterigmatocystin
biosynthesis in the fungus Aspergillus nidulans. PLoS One. 8(7):e68492.

Hernndez-Ortiz P, Espeso EA. [2013] Phospho-regulation and nucleocytoplasmic


trafficking of CrzA in response to calcium and alkaline-pH stress in Aspergillus
nidulans. Mol Microbiol. 89(3):532-51.

Figura 2 | Figure 2
Sealizacin del factor CrzA y anlisis fenotpico de cepas nulas CrzA y SltA. A) CrzA
muestra diferentes estados de fosforilacin y la adicin de calcio o la alcalinizacin del
medio altera el patrn de fosforilacin. RC=resting cells. B) La ausencia de CrzA causa
sensibilidad al calcio mientras que una cepa nula sltA es sensible a una gran variedad
de cationes, y ambas a pH alcalino.
Signalling of CrzA factor and phenotypic analyses of null crzA and sltA strains. A) CrzA
displays different phosphorylation states. Addition of calcium or medium alkalinisation
alter the phospho-pattern. RC=resting cells. B) Absence of CrzA activity results in calcium
sensitivity. A null sltA strain is sensitive to a large variety of cations, both null strains are
sensitive to alkalinity.

Zhang, J, R Qiu, HN Arst, Jr, MA Penalva, and X Xiang [2014] HookA is a novel
dynein-early endosome linker critical for cargo movement in vivo. Journal of Cell
Biology 204:1009-1026. (Editorial comment as Journal Focus).

Pantazopoulou, A, M Pinar, X Xiang & MA Pealva [2014] Maturation of late Golgi


cisternae into RabERAB11 exocytic post-Golgi carriers visualized in vivo. Mol Biol
Cell 25: 2428-2443 (edited by Benjamin Glick)

Arst, HN, Jr, Hernndez-Gonzlez, M, Pealva, MA, and Pantazopoulou, A. [2014]


GBF/Gea mutant with a single substitution sustains fungal growth in the absence of
BIG/Sec7. FEBS Lett 588, 4799-4786.

Pealva MA, Lucena-Agell D, Arst HN Jr. [2014] Liaison alcaline: Pals entice
non-endosomal ESCRTs to the plasma membrane for pH signaling. Curr Opin
Microbiol. 22C:49-59.

Bertuzzi M, Schrettl M, Alcazar-Fuoli L, Cairns TC, Muoz A, Walker LA, Herbst S,


Safari M, Cheverton AM, Chen D, Liu H, Saijo S, Fedorova ND, Armstrong-James D,
Munro CA, Read ND, Filler SG, Espeso EA, Nierman WC, Haas H, Bignell EM. [2014]
The pH-responsive PacC transcription factor of Aspergillus fumigatus governs
epithelial entry and tissue invasion during pulmonary aspergillosis. PLoS Pathog.
10(10):e1004413.
Liu W, Mellado L, Espeso EA, Sealy-Lewis HM. [2014] In Aspergillus nidulans the
suppressors suaA and suaC code for release factors eRF1 and eRF3 and suaD
codes for a glutamine tRNA. G3 (Bethesda). 4(6):1047-57.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Germn Rivas Caballero


Investigador Cientfico
grivas@cib.csic.es
PhD, 1989
Universidad Autnoma de Madrid
Postdoctoral, 1990-1993
NIH, Bethesda, USA
Biozentrum, Univ. Basilea, CH

Investigador, 1994
Cientfico Titular, 1995
Jefe de Grupo,1996
Investigador Cientfico, 2006
CIB, CSIC

Investigadores del equipo | Staff scientists:


Carlos Alfonso Botello
Mercedes Jimnez Sarmiento

Silvia Zorrilla Lpez

Otros miembros | Other lab members:


Victor Hernndez Rocamora
Elisa Jimnez Cabr
Begoa Monterroso Marco
Concepcin Garca Montas

Ana Raso Alonso


Marta Sobrinos Sanguino
Noelia Ropero
Alicia Rodrguez

http://www.cib.csic.es/es/grupo.grivas

Bioqumica de Sistemas de
la Divisin Bacteriana
Nuestro objetivo es entender cmo los elementos de la maquinaria de la
divisin bacteriana (el divisoma) funcionan como un sistema integrado de
interacciones moleculares para ejercer su funcin esencial. Desarrollamos
y aplicamos novedosos abordajes de reconstitucin bioqumica para
construir, con un conjunto mnimo de protenas, ensamblajes funcionales
de divisin en ausencia de clulas.

Organizacin y reconstruccin bioqumica de componentes del anillo


FtsZ en sistemas de membrana: La
divisin bacteriana est mediada por un conjunto de protenas que interaccionan en el sitio
de divisin, ensamblando un anillo dinmico
que dispara la citoquinesis y forma parte del
divisoma. En Escherichia coli, la protena FtsZ,
principal elemento del anillo septal, se ancla a la
membrana interna por la accin de las protenas
ZipA y FtsA, formando el primer ensamblaje
molecular del divisoma, el proto-anillo. El posicionamiento del anillo en el punto medio est
regulado por dos sistemas de control (el complejo MinCDE y la oclusin del nucleoide - SlmA)
que inhiben su formacin en lugares equivocados. Estudiamos las actividades e interacciones
de FtsZ en reconstrucciones mnimas del protoanillo estructuradas en sistemas de membrana, como nanodiscos, microesferas, bicapas,
vesculas y microgotas (Fig. 1). Investigamos la
accin de MinCDE y SlmA sobre las propiedades de divisomas mnimos.

2 Reactividad macromolecular y organizacin en entornos aglomerados y connados citomimticos: El ensamblaje del


divisoma in vivo tiene lugar en entornos
caracterizados por la presencia de altas
concentraciones de macromolculas, que
pueden estructurarse como redes dinmicas solubles o asociadas a la membrana.
Aplicamos y diseamos reconstrucciones
sintticas de estos microentornos para investigar el impacto de la exclusin de volumen y
la unin a membranas sobre las propiedades
y el comportamiento de divisomas mnimos.
3 Bioqumica fsica de interacciones macromoleculares: Investigamos las propiedades
de asociacin de FtsZ con otros elementos
del divisoma mediante ultracentrifugacin
analtica, dispersin de luz y espectroscopas
de uorescencia (Fig. 2). En paralelo, estudiamos las interacciones de FtsZ con elementos del divisoma asociados a membrana
mediante ensayos bioqumicos y biofsicos.

Financiacin | Funding
HEALTH-F3-2009-223432. (Comunidad Europea)
RGP0050-2010. (Human Frontier Science

Program)

BIO2011-28941-C03. (MINECO)

Publicaciones
Seleccionadas
Selected Publications
Ahijado-Guzmn R, Alfonso C, Reija B, Salvarelli E,
Mingorance J, Zorrilla S, Monterroso B, Rivas G [2013]
Control by potassium of the size-distribution of
Escherichia coli FtsZ polymers is independent of
GTPase activity. J. Biol. Chem. 288:27358-27365.
Cabr EJ, Snchez-Gorostiaga A, Carrara P, Ropero
N, Casanova M, Palacios P, Stano P, Jimnez M,
Rivas G, Vicente M [2013] Bacterial division proteins
FtsZ and ZipA induce vesicle shrinkage and cell
membrane invagination. J. Biol. Chem. 288:2662526634.

Hernndez-Rocamora VM, Garca-Montas C, Reija


B, Monterroso B, Margolin W, Alfonso C, Zorrilla S,
Rivas G [2013] MinC shortens FtsZ protofilaments by
preferentially interacting with GDP-bound subunits.
J. Biol. Chem. 288:24625-24635.

Jimnez M, Cabr EJ, Raso A, Martos A, Rivas


G [2013] Giant vesicles: a powerful tool to
reconstruct bacterial division assemblies in cell-like
compartments. Environ. Microbiol. 15:3158-3168.

Mellouli S, Monterroso B, Vutukuri HR, te Brinke E,


Chokkalingam V, Rivas G, Huck WTS [2013] Selforganization of the bacterial cell-division protein FtsZ
in confined environments. Soft Matter 9:1049310500.

Monterroso B, Alfonso C, Zorrilla S, Rivas G [2013]


Combined light scattering, ultracentrifugation and
fluorescence correlation spectroscopy studies on the
associations and assembly of the Escherichia coli
cell division FtsZ protein. Methods 59:349-362.

Rivas G, Alfonso C, Jimnez M, Monterroso B, Zorrilla


S [2013] Macromolecular interactions of bacterial cell
division FtsZ protein: From quantitative biochemistry
and crowding to reconstructing minimal divisomes in
the test tube. Biophys. Rev. 5:63-77.
Ahijado-Guzmn R, Prasad J, Rosman C, Henkel
A, Tome L, Schneider D, Rivas G, Snnichsen C
[2014] Plasmonic nanosensors for simultaneous
quantification of multiple protein-protein binding
affinities. Nano Lett. 14:5528-5532.

Alvira S, Cullar J, Rhl A, Yamamoto S, Itoh H,


Alfonso C, Rivas G, Buchner J, Valpuesta JM [2014]
Structural characterization of the substrate-transfer
mechanism in Hsp70/Hsp90 folding machinery
mediated by Hop. Nat Commun. 5:5484. doi:
10.1038/ncomms6484.
Rivas G, Vogel SK, Schwille P [2014] Reconstitution
of cytoskeletal protein assemblies for large-scale
membrane transformation. Curr. Opin. Chem.
Biol. 22C:18-26.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Figura 1 | Figure 1
Reconstitucin de elementos del protoanillo en vesculas. Mtodo de emulsin (A)
para encapsular y polimerizar FtsZ dentro de GUVs permeables (B). (C) Contraccin
de GUVs debido a la interaccin de polmeros de FtsZ y ZipA asociada a la membrana
(0, 5, 10 min). (D) Bloqueo de la contraccin al aadir un pptido de FtsZ que inhibe la
interaccin FtsZ-ZipA (Cabr et al. 2013; Rivas et al. 2014).
Reconstitution of proto-ring elements in vesicles. Water-in-oil droplet transfer method (A)
used to encapsulate and polymerize FtsZ inside permeable GUVs (B). (C) Shrinking of GUVs
through the interaction of FtsZ with membrane-associated ZipA (0, 5 and 10). (D) Blocking
shrinkage by the addition of an FtsZ-derived peptide that inhibits FtsZ-ZipA interaction.
(Cabr et al. 2013; Rivas et al. 2014).

Figura 2 | Figure 2
Anlisis biofsico de los diferentes comportamientos de los oligmeros de GDPFtsZ (gris) y los polmeros de GTP-FtsZ (negro) a partir de medidas de velocidad de
sedimentacin (A), dependencia con la concentracin de la dispersin de luz esttica
(B), dispersin de luz dinmica (C) y espectroscopa de correlacin de fluorescencia
(D). (Monterroso et al. 2013).
Biophysical analysis of the different behavior of GDP-FtsZ oligomers (grey) and GTPFtsZ polymers (black) from measurements of sedimentation velocity (A), concentration
dependence static light scattering (B), dynamic light scattering (C) and fluorescence
correlation spectroscopy (D). (Monterroso et al. 2013).

Systems Biochemistry
of Bacterial Division
Our research aims at understanding how the elements of the bacterial division machinery (the divisome) work
together as an integrated system of molecular interactions to fulfill its essential function. To address these
questions we develop and apply novel biochemical reconstitution approaches to build, with a minimum set of
elements, functional division assemblies in the absence of cells.

Biochemical organization and reconstruction of FtsZ ring components in


minimal membrane systems: Bacterial
division is mediated by a set of proteins
that interact at the division site to assemble a dynamic ring, a structure that drives
cytokinesis, forming part of the divisome. In
Escherichia coli, the FtsZ protein, main element of the septal ring, is anchored to the inner
membrane by the action of the proteins ZipA
and FtsA, forming the rst molecular assembly
of the divisome, the proto-ring. The positioning
of the ring at midcell is regulated by two control systems (MinCDE complex and nucleoid
occlusion - SlmA) that inhibit its formation at
wrong places. We study the activities and

interactions of FtsZ in minimal reconstructions


of the proto-ring structured in membrane systems as nanodics, microbeads, bilayers, vesicles and microdroplets (Fig. 1). We investigate
the concerted action of MinCDE and SlmA on
the properties of minimal divisomes.
2 Macromolecular reactivity and organization in crowded and conned cell-like
environments: The assembly of the divisome in vivo takes place in environments
characterized by the presence of high concentrations of macromolecules, which may
be structured as soluble and membraneassociated dynamic networks. We apply
and design synthetic reconstructions of

these microenvironments to investigate the


impact of excluded volume and surface
binding eects on the properties and behavior of minimal divisomes.
3 Physical biochemistry of macromolecular interactions: We investigate the association properties of FtsZ with itself and with
other divisome elements using analytical
ultracentrifugation, light scattering and uorescence spectroscopies (Fig. 2). In parallel, we study the interactions of FtsZ with
membrane-associated division elements by
biochemical and biophysical assays.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Rafael Giraldo Surez


Profesor de Investigacin
rgiraldo@cib.csic.es
PhD, 1991
Universidad Complutense de Madrid

Postdoctoral, 1992-1994
Divisin de Estudios Estructurales,
Laboratorio de Biologa Molecular del MRC
(Cambridge, UK)

Investigador Contratado MEC, 1995-1999


Cientfico Titular, 2000-2008
Investigador Cientfico, 2008-2009
Profesor de Investigacin, 2010
CIB, CSIC
Miembro, 2010
Academia Europea

Investigadores del equipo | Staff scientists:


M. Elena Fernndez-Tresguerres Rodrguez-Vigil
Juan Francisco Gimnez Abin

Otros miembros | Other lab members:


Mara Moreno del lamo
Cristina Fernndez Fernndez
Ftima Gasset Rosa
Laura Molina Garca

Ada Revilla Garca


Ana Mara Serrano Lpez
Maria Cruz Snchez Martnez

http://www.cib.csic.es/es/grupo.php?idgrupo=61

Ensamblajes Macromoleculares
Microbianos Sintticos
Mediante aproximaciones de Biologa Sinttica, desarrollamos mdulos derivados de RepA, una protena de
replicacin propia de plsmidos bacterianos, que permiten controlar el ensamblaje amiloide en condiciones
cuasifisiolgicas y construir en microorganismos una proteinopata amiloide modelo genrica. Esperamos
as poder deconstruir e intervenir las rutas y mecanismos de citotoxicidad compartidos entre las amiloidosis
bacterianas y humanas.

uestro grupo desvel (1998-2008) el mecanismo que activa, en


bacterias Gram-negativas, la replicacin de plsmidos por protenas de la familia RepA. Descubrimos que una conformacin
funcionalmente activa se selecciona por la unin de RepA a secuencias
de DNA, que actan como efectores alostricos, y por la chaperona
DnaK (Hsp70). Recientemente (2007-2012), encontramos que la unin
al DNA tambin promueve in vitro el ensamblaje como bras amiloides
de un dominio winged-helix (WH1) en RepA, al igual que sucede con
las protenas causantes de las encefalopatas espongiformes y de la
enfermedad de Parkinson. Como prueba de concepto, la amiloidognesis es inhibida por una molcula que interere con la unin de RepAWH1 al DNA.
Durante los dos ltimos aos (2012-2014), hemos estudiado cmo se
comporta RepA-WH1 in vivo. Cuando se expresa en E. coli, fuera de su
contexto funcional natural y fusionada a una protena marcadora uorescente, RepA-WH1 causa una proteinopata amiloide sinttica. Aunque

Patentes | Patents
Rafael Giraldo y Mara Moreno del lamo. 25 septiembre 2013. Anticuerpo
monoclonal B3h7 anti-oligmeros amiloides RepA-WH1, hibridoma que lo
produce y aplicaciones. OEPM / P201331391

Financiacin | Funding
CSD2009-00088 (MINECO)
BIO2012-30852 (MINECO)

no es un agente infeccioso, por lo que se considera un prionoide,


RepA-WH1 es capaz de moldear su conformacin amiloide sobre molculas de la misma protena, tanto in vitro como in vivo. Hemos caracterizado, mediante microudica, su propagacin vertical de clula madre
a clulas hijas. Los linajes bacterianos transmiten epigenticamente dos
estirpes amiloides alternativas de RepA-WH1: mltiples partculas globulares de toxicidad aguda o un nico agregado elongado, que reduce
en menor medida la proliferacin celular. La chaperona DnaK modula la
interconversin entre ambas estirpes de RepA-WH1.
RepA-WH1 parece mimetizar dedignamente en bacterias la patogenicidad que amiloides con relevancia clnica ejercen sobre las mitocondrias,
orgnulos que descienden de -proteobacterias de vida libre adquiridas
como endosimbiontes. El prionoide bacteriano sinttico RepA-WH1 es
un modelo mnimo y bioseguro para desentraar los mecanismos y vas
comunes a las proteinopatas amiloides.

Publicaciones Seleccionadas
Selected Publications
Lane AB, Gimnez-Abin JF, Clarke DJ [2013] A novel chromatin tether domain
controls topoisomerase IIa dynamics and mitotic chromosome formation. J Cell
Biol 203:471-486.
Gasset-Rosa F, Coquel AS, Moreno-del lamo M, Chen P, Song X, Serrano AM,
Fernndez-Tresguerres ME, Moreno-Daz de la Espina S, Lindner AB, Giraldo R
[2014] Direct assessment in bacteria of prionoid propagation and phenotype
selection by Hsp70 chaperone. Mol Microbiol 91:1070-1087.

Molina-Garca L, Giraldo R [2014] Aggregation interplay between variants of the


RepA-WH1 prionoid in Escherichia coli. J Bacteriol 196:2536-2542.

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By means of Synthetic Biology approaches, we are developing modules derived from RepA, a DNA replication
protein in bacterial plasmids, which allow control on amyloid assembly under quasi-physiological conditions and the
construction in microorganisms of a generic amyloid proteinopathy. We thus aim later to deconstruct, and enable
intervention on, the pathways and mechanisms of cytotoxicity shared by human and bacterial amyloidosis.

ur group had pioneered (1998-2008)


knowledge on the mechanisms enabling initiation of plasmid DNA replication, in Gram-negative bacteria, by proteins
of the RepA family. We had discovered that
a functionally active RepA conformation is
selected by specic DNA sequences, acting
as allosteric eectors, and by DnaK, a chaperone of the Hsp70 family. More recently (20072012), we found that binding to DNA also promotes in vitro the assembly into amyloid bres
of a winged-helix domain (WH1) in RepA, as
described for proteins involved in spongiform
encephalopathies and Parkinsons disease. As
a proof of concept, RepA-WH1 amyloidogenesis can be inhibited by a molecule interfering
with binding to DNA.

In the last two years (2012-2014), we have


focussed our research on how RepA-WH1
behaves in vivo. When it is expressed in
E. coli, uncoupled from its natural functional
context as a fusion to a uorescent protein
reporter, RepA-WH1 causes a synthetic amyloid proteinopathy. Although RepA-WH1 is not
infectious, therefore behaving as a prionoid,
it templates the amyloid conformation by
cross-seeding, both in vitro and in vivo. We
have surveyed through microuidics the vertical transmission of RepA-WH1 amyloidosis
from mother to daughter bacterial cells. We
have discovered that bacterial lineages epigenetically propagate two alternative amyloid
strains of RepA-WH1: either multiple globular
particles with acute cytotoxicity, or a single

elongated aggregate, mildly detrimental to cell


proliferation. DnaK chaperone modulates the
conformational switch between both strains
of RepA-WH1.
RepA-WH1 seems to parallel in bacteria
the pathogenicity of clinically relevant protein amyloids on mitochondria, organelles
descending from free-living -proteobacteria
that subsequently underwent symbiosis. The
bacterial synthetic prionoid RepA-WH1 might
be a suitable, minimal and bio-safe model
system for untangling key pathways common
to amyloid proteinopathies.

Biologa Celular y Molecular | Cellular & Molecular Biology

Synthetic Microbial Macromolecular


Assemblies

Figura 1 | Figure 1
La amiloidognesis de RepA-WH1 in vitro (dcha.) implica la disociacin, mediada por efector (DNA), de dmeros en monmeros metaestables que se ensamblan jerrquicamente en
filamentos y fibras amiloides entrelazados. Estructuras resueltas en colaboracin con A. Romero (2003) y O. Llorca (2015). En E. coli los agregados amiloides (izda., sectores rojos) son
citotxicos y verticalmente transmisibles.
RepA-WH1 amyloidogenesis in vitro (right) implies effector (DNA)-mediated dissociation of dimers into metastable monomers that assemble hierarchically into intertwined amyloid filaments and variably
twisted fibres. Structures solved in collaboration with the groups of A. Romero (2003) and O. Llorca (2015). In E. coli, amyloid aggregates (left, red sectors) are cytotoxic and vertically inheritable.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Jorge Bernardo Schvartzman Blinder


Profesor de Investigacin
schvartzman@cib.csic.es
PhD, 1979
Universidad Politcnica de Madrid, Espaa

Postdoctoral, 1980-1982
Brookhaven National Laboratory (New York,
USA)
Fullbright Fellow, 1987-1989
Albert Einstein College of Medicine
(New York, USA)
Cientfico Titular, 1985
Jefe de Grupo, 1980
Investigador Cientfico, 2002
Profesor de Investigacin, 2007
CIB, CSIC

Investigadores del equipo | Staff scientists:


Dora Beatriz Krimer Smunis

Pablo Hernndez Valenzuela

Otros miembros | Other lab members:


Jorge Cebrin Castillo
Vanessa Fernndez Calleja
Alicia Castn Garca
Jos Manuel Belmonte Rodrguez

Celia Bolomburu
Idoia Garca Hernando
Leticia Garca Martnez
Mara-Luisa Martnez-Robles

http://www.cib.csic.es/es/grupo.php?idgrupo=2

Biologa Molecular de los Cromosomas


Nos interesa la interrelacin y coordinacin de los procesos biolgicos en los que est involucrado el DNA:
replicacin, transcripcin, reparacin y recombinacin, cmo estn regulados y cmo modifican o son afectados
por factores genticos, epigenticos y ambientales como la topologa del DNA, la organizacin de la cromatina y el
estrs nutricional.

) Utilizamos la electroforesis bidimensional en geles de agarosa


para demostrar que la movilidad electrofortica de molculas
de igual masa vara dependiendo de si estn superenrolladas,
encadenadas o anudadas. Esto nos ha permitido identicar las condiciones ptimas para distinguir cada familia de topoismeros. Tambin
analizamos el comportamiento de minicromosomas circulares y lineares
de Saccharomyces cerevisiae en clulas sincronizadas en presencia y
ausencia de la topoisomerasa 2 (topo 2). Los resultados indican que la
topo 2 no es necesaria para la replicacin y segregacin de cromosomas
articiales lineales de levaduras de pequeo tamao (YACs). b) Hemos
estudiado la replicacin del DNA durante la diferenciacin terminal en
clulas eritroleucmicas murinas (MEL). Utilizamos la incorporacin de
BrdU, la citometra de ujo, el peinado del DNA y la tincin por inmunouorescencia indirecta para demostrar que la velocidad de progreso
de las horquillas replicativas se ralentiza y la distancia entre orgenes
disminuye a medida que las clulas dejan de proliferar y se acumulan
en G1. Proponemos que este comportamiento es general causado por
la heterocromatinizacin, conrmada por la acumulacin progresiva de
la HP1 que caracteriza la diferenciacin celular terminal. c) Una de las
causas ms importantes de inestabilidad genmica es la parada de las
horquillas replicativas del DNA. Estudiamos los mecanismos celulares
que previenen la parada de horquillas y aquellos que operan sobre las
horquillas detenidas para prevenir la inestabilidad que su reactivacin
puede ocasionar. Nos interesan especialmente el bloqueo de las horquillas ocasionado por la colisin entre las maquinarias replicativa y transcripcional y por el estrs topolgico del DNA, en cuya liberacin las DNA
topoisomerasas juegan un papel central. Deciencias en estos mecanismos son la base molecular de varias enfermedades, caracterizadas por
una alta inestabilidad gentica y predisposicin al cncer.

Publicaciones Seleccionadas
Selected Publications
Schvartzman JB, Martnez-Robles ML, Hernndez P, Krimer DB [2013] The benefit
of DNA supercoiling during replication. Biochemical Society Transactions 41:
646-651.
Schvartzman JB, Martnez-Robles ML, Hernndez P, Krimer DB [2013] Plasmid DNA
topology assayed by two-dimensional agarose gel electrophoresis. In Methods
Mol Biol 1054: 121-132, DNA Electrophoresis: Methods and Protocols (Svetlana
Makovets, ed.) Springer Science Business Media, New York.
Fernndez-Nestosa MJ, Monturus ME, Snchez Z, Torres F, Fernndez A, Fraga
M, Hernndez P, Schvartzman JB, Krimer DB [2013] DNA methylation-mediated
silencing of PU.1 in leukemia cells resistant to cell differentiation. SpringerPlus 2,
392 DOI:10.1186/2193-1801-2-392.

Figura 2 | Figure 2
Molculas de DNA aisladas de clulas MEL DS19 durante la diferenciacin extendidas
por peinado molecular. La deteccin inmunocitoqumica de tramos marcados
secuencialmente con IdU (en rojo) seguido de CldU (en verde) permite identificar los
sitios de iniciacin de la replicacin y la distancia entre orgenes as como calcular la
velocidad de progreso de las horquillas.
Selected DNA molecules isolated from MEL DS19 cells along differentiation stretched by
DNA combing. The immunocytological detection of sequentially labelled tracks with IdU
(red) followed by CldU (green) allows the identification of replication origins and inter-origin
distances as well as the calculation of the rate of replication fork progression.

Cebrin J, Monturus ME, Martnez-Robles ML, Hernndez P, Krimer DB, Schvartzman


JB [2014] Topoisomerase 2 Is Dispensable for the Replication and Segregation
of Small Yeast Artificial Chromosomes (YACs). PLoS ONE 9(8): e104995.
DOI:10.1371/journal.pone.0104995.
Cebrin J, Kadomatsu-Hermosa MJ, Castn A, Martnez V, Parra C, FernndezNestosa MJ, Schaerer C, Martnez-Robles ML, Hernndez P, Krimer DB, Stasiak A,
Schvartzman JB [2014] Electrophoretic Mobility of Supercoiled, Catenated and
Knotted DNA Molecules. Nucleic Acids Res DOI: 10.1093/nar/gku1255.

Cebrin J, Castn A, Martnez V, Parra C, Kadomatsu-Hermosa MJ, FernndezNestosa MJ, Schaerer C, Hernndez P, Krimer DB, Schvartzman JB [2015] Direct
evidence for the formation of precatenanes during DNA replication. Journal of
Biol Chem DOI: 10.1074/jbc.M115.642272.

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We are interested in the relationships and coordination between biological processes where DNA is involved:
replication, transcription, repair and recombination, how are they regulated and how they alter or are affected
by genetic, epigenetic and environmental factors such as DNA topology, chromatin organization and nutritional
stress.

) We used two-dimensional (2D) agarose gel electrophoresis to


show that for molecules of the same mass the electrophoretic
mobility of supercoiled, catenated and knotted DNAs dier as
the electrophoretic conditions change. This allowed us to identify
the optimal conditions to distinguish each family of topoisomers. We
analyzed also the behavior of circular and linear minichromosomes of
Saccharomyces cerevisiae in synchronized cells in the presence and
absence of topoisomerase 2 (topo 2). The results obtained indicated
that topo 2 is dispensable for the replication and segregation of small
linear yeast articial chromosomes (YACs). b) We investigated DNA
replication during terminal cell dierentiation in murine erythroleukemia
(MEL) cells. We used BrdU labeling, cell ow cytometry, genome-wide
DNA combing and indirect immunouorescent staining to show that the
rate of replication fork movement slowdown and the inter-origin distance
becomes shorter as cells stop proliferating and accumulate in G1. We
propose this is a general feature caused by the heterochromatinization,

conrmed by the progressive accumulation of HP1 that characterizes


terminal cell dierentiation. c) DNA replication fork arrest is one of the
most important causes of genomic instability. We study the cellular
mechanisms that prevent fork arrest and those operating at the arrested
forks to prevent the instability that their reactivation may cause. We are
especially interested in the induction of fork arrest produced upon collision between DNA replication and RNA transcription machineries and
by the accumulation of topological DNA stress, in whose release DNA
topoisomerases play a central role. Deciencies in these mechanisms
are the molecular basis of several diseases characterized by a high
genetic instability and cancer predisposition.

Biologa Celular y Molecular | Cellular & Molecular Biology

Molecular Biology of the Chromosomes

Financiacin | Funding
BFU2011-22489 (MINECO)

Figura 1 | Figure 1
Anlisis de la topologa del DNA. A) La electroforesis bidimensional en geles de agarosa con distintas concentraciones de cloroquina permite distinguir todos los topoismeros de
molculas circulares covalentemente cerradas (CCCs). As se puede identificar el topoismero ms abundante y calcular la densidad de superenrollamiento. B) El recubrimiento del
DNA con la protena RecA permite visualizar molculas encadenadas por microscopa electrnica.
Analysis of DNA topology. A) Two-dimensional (2D) agars gel electrophoresis in the presence of different concentrations of chloroquine allows the identification of all the topoisomers of covalentlyclosed circles (CCCs). This technique can be used to recognize the most abundant topoisomer to calculate supercoiling density. B) Covering DNA molecules with the bacterial protein RecA allows
the identification of catenanes by electron microscopy.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Miguel Angel Vidal Caballero


Investigador Cientfico
mvidal@cib.csic.es
PhD, 1985
Universidad Complutense de Madrid

Postdoctoral, 1985-1989
National Institute for Medical Research
(MRC, UK)
Cientfico Titular, 1991
Jefe de Grupo, 1991
Investigador Cientfico, 2008
CIB, CSIC

Otros miembros | Other lab members:


Mnica Bravo Madrigal
Katarzyna Starowicz
Fabio Nicolini

https://www.cib.csic.es/es/grupo.php?idgrupo=14

El Sistema Polycomb
de Regulacin
Epigentica
El grupo de genes Polycomb (PcG) codifica
reguladores epigenticos que forman complejos con
actividad modificadora de cromatina. Bien conocidos
como reguladores transcripcionales durante el
desarrollo embrionario, participan crticamente en
diferenciacin y homeostasis celulares, actuando
sobre progenitores. Nuestro trabajo se centra en los
complejos que monoubiquitinan la histona H2A.

a monoubiquitinacin de la histona H2A (lisina 119) correlaciona


con estados de transcripcionalmente reprimidos y, fundamentalmente, depende de RING1A y RING1B, E3 protein ligasas
del sistema Polycomb. Estas proten ligasas son parte esencial de los
complejos PRC1, uno de los dos tipos de ensamblajes moleculares del
sistema Polycomb. RING1A y RING1B, as como RYBP, una subunidad
comn a todos los complejos no cannicos PRC1, fueron identicados
en el laboratorio. En la actualidad, usamos modelos murinos de prdida
de funcin y otros que expresan formas modicadas de subunidades
PRC1 para el estudio funcional y bioqumico de subunidades PRC1.
El compartimento hematopoytico y clulas pluripotentes (neural,
ES) son nuestros modelos de trabajo. Uns observacin general es la
apreciacin de que las subunidades PRC1 promueven proliferacin/
supervivencia celulares. As, la inactivacin combinada de los homlogos Ring1A y Ring1B resultan en paradas proliferativas casi inmediatas,
debida al aumento de niveles de reguladores negativos de proliferacin
que detienen el ciclo celular antes de la fase S. Adems, estas clulas
muestran alteraciones en replicacin y evidencia de inestabilidad genmica. RING1A y RING1B pueden actuar sobre el proceso replicativo en
s, o sobre las vas de reparacin disparadas por el estress replicativo. Dado
el efecto dominante que el bloqueo del ciclo celular tiene sobre cualquier
otro tipo de anlisis de las clulas mutantes sera importante desacoplar
esta actividades de las asociadas con regulacin transcripcional.
Para esclarecer los mecanismos de accin de RING1A y RING1B utilizamos aproximaciones protemicas dirigidas a la identicacin de protenas asociadas. Con este n usamos una lnea de ratones que expresa
una forma modicada de RING1B que permite su aislamiento ecaz,
previo al anlisis por cromatografa lquida y espectrometra de masas.
El trabajo est enfocado a lneas hematopoyticas.

Figura 1 | Figure 1
Asociacin de RING1B con la maquinaria de replicacin. Sitios de interaccin de
RING1B con la abrazadera de replicacin PCNA, detectada en un ensayo de ligacin
en proximidad (PLA) como focos coloreados en rojo. DNA nuclear (teido con DAPI,
A) y replicando (marcado mediante incorporacin de EdU, un anlogo de timidina, en
verde, B). Barra, 10 m.
Asociation of RING1B with the replication machinery. Proximity ligation assay (PLA) detects
RING1B association to the replicative slide clamp PCNA, visualized as red foci. Total DNA
(DAPI, A) and replicating DNA (B, EdU). Scale bar, 10 m.

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The Polycomb group (PcG) of genes encode epigenetic regulators assembled in complexes displaying
chromatin modifying activities. Well known developmental regulators, they participate in cell differentiation
and tissue homeostasis with an emphasis in progenitor cells. We focus on core subunits of complexes that
monoubiquitylate histone H2A.

ING1A and RING1B, the evolutionary conserved Polycomb E3


ligases, are responsible for most of histone H2A (lysine 119)
monoubiquitylation, a modication that correlates with transcriptional repression. RING1A and RING1B are part of the core of PRC1
complexes, one of the two major categories of Polycomb assembles.
We identied RING1A and RING1B and also RYBP, a subunit unique to
non-canonical PRC1 complexes. We use loss-of-function mouse models
as well as mouse strains that express tagged variants for functional and
biochemical analysis regarding transcriptional and non-transcriptional
activities.
Currently, we investigate roles of RING1 and RYBP proteins in hematopoyetic and neural homeostasis and in ES cells pluripotency. A general observation arising from these studie is the positive role that PRC1
subunits have on cell proliferation/survival. Thus, compound inactivation
of Ring1A and Ring1B paralogs leads to acute proliferation arrest that
involves a variety of proliferation inhibitors that prevent entry in S-phase.

A more detailed analysis of these cells, however, allowed the identication


of RING1A and RING1B activities during replication and genome stability.
Work to ascertain whether it is a role in assisting replication or in xing replicative stress is underway. Understanding these activities would also help
to overcome the dominant, obscuring eects that impaired cell cycle progression has on the analysis of transcriptional programs in mutant cells.
We have set up a proteomic approach aimed at identifying RING1A/
RING1B partners that could illuminate mechanisms in transcriptional and
non-transcriptional functions. It uses a mouse model carrying a knockedin modication of the Ring1B locus that expresses a tagged-Ring1B
protein. The model, one as physiological as it can get, also permits the
access to PRC1 complexes in primary cells or in ex-vivo expanded primary cells that are not often available as established tissue culture cells
lines. Ongoing works focuses on hematopoietic cell lineages.

Biologa Celular y Molecular | Cellular & Molecular Biology

Epigenetic Control by the Polycomb


Group of Genes

Financiacin | Funding
BFU2010-18146 (MINECO)

Oncocycle S2010/BMD2470 (CAM)


FP7-People-2011-ITN

SAF2013-47997-P (MINECO)

Publicaciones Seleccionadas
Selected Publications
Morimoto-Suzki N, Hirabayashi Y, Tyssowski K, Shinga J,Vidal M, Koseki H, Gotoh
Y [2014] The polycomb component Ring1B regulates the timed termination
of subcerebral projection neuron production during neocortical development.
Development 141:4343-53.

Vidal M [2014] Polycomb complexes: chromatin regulators required for cell diversity
and tissue homeostasis. pp95-139. C. Bonifer and P.N. Cockerill (eds.).
Transcriptional and Epigenetic Mechanisms Regulating Normal and Aberrant Blood Cell
Development, Epigenetics and Human Health, Springer-Verlag Berlin Heidelberg.
Kondo T, Isono K, Kondo K, Endo TA, Itohara S, Vidal M, Koseki H [2014] Polycomb
potentiates Meis2 activation in midbrain by mediating interaction of the promoter
with a tissue-specific enhancer. Dev Cell 28:94-101.

Figura 2 | Figure 2
Mitosis aberrante en clulas mutantes que carecen de RING1A y RING1B. La tincin
de DNA con DAPI muestra un puente cromosomal probablemente consecuencia de
replicacin incompleta. Barra, 10 m.
Aberrant mitosis of RING1A and RING1B-deficient cells. DAPI-stained mitosis showing
chromosomal bridges indicating incomplete DNA replication. Scale bar, 10 m.

Martnez-Gmez AI, Villegas S, Aguado-Llera C, Bacarizo J Cmara-Artigas A, Vidal


M Neira JL [2014] The isolated N terminus of Ring1B is a well-folded, monomeric
fragment with native-like structure. Protein Eng Des Sel 27:1-11.

Frangini, A. Sjberg, M., Romn-Trufero, M., Dharmalingam, G., Haberle,V., Bartke,


T.,Lenhard, B., Malumbres, M., Vidal M, Dillon N [2013] The Aurora B Kinase
and the Polycomb Protein Ring1B Combine to Regulate Active Promoters in
Quiescent Lymphocytes. Mol Cell 51:647661.
Yokobayashi S, Liang CY, Kohler H, Nestorov P, Liu Z, Vidal M, van Lohuizen M,
Roloff TC, Peters AH [2013] PRC1 coordinates timing of sexual differentiation of
female primordial germ cells. Nature 495: 236-240.

van Arensbergen J, Garca-Hurtado J, Maestro MA, CorreaTapia M, Rutter G, Vidal


M, Ferrer J [2013] Ring1B bookmarks genes in pancreatic embryonic progenitors
for repression in adult beta cells. Genes Dev. 27:52-63.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Patricia Boya
Cientifica titular
pboya@cib.csic.es
PhD, 2000
Universidad de Navarra

Postdoctoral, 2001-2005
CNRS (Pars, Francia)
Universisty of Cambridge (UK)
Contrato, 2005-2009
Ramn y Cajal

Cientfica Titular, 2009


CIB, CSIC

Otros miembros | Other lab members:


Lorena Esteban Martnez
Raquel Gmez Sintes
Luca Garca Ledo
Esther Seco Martn

Ana Serrano Puebla


Sergio Rivas Muoz
Elena Sierra Filardi

http://www.cib.csic.es/es/grupo.php?idgrupo=73

Funciones de la Autofagia en la
Fisiopatologa de los Organismos
En nuestro laboratorio utilizamos modelos celulares y animales para comprender el papel de la autofagia en
la fisiologa y la patologa de los organismos. Este es un proceso de degradacin intracelular que permite la
eliminacin y el reciclaje de componentes celulares. Es una importante respuesta frente al ayuno nutricional,
participa en la degradacin de orgnulos celulares y permite la supervivencia en situaciones de estrs.

l inters de nuestro laboratorio se centra en entender por que el


proceso de la autofagia es esencial para mantener la homeostasis
de las clulas y qu patologas subyacen a alteraciones de este
mecanismo de degradacin intracelular.
La importancia del proceso de autofagia queda patente por la letalidad
embrionaria de animales decientes en algunos de los genes Atg. En nuestro grupo estudiamos la relacin de la autofagia con procesos esenciales
para las clulas como la proliferacin, diferenciacin y la muerte celular.
Hemos demostrado que este proceso es importante para la diferenciacin
neuronal ya que animales decientes de autofagia no generan neuronas
maduras y poseen defectos en neuritogenesis. Por otro lado hemos
demostrado que la induccin temprana de la autofagia durante procesos
neurodegenerativos supone una respuesta citoprotectora. Dao axonal
producido in vivo en animales decientes de autofagia aumenta los niveles
de muerte celular y por el contrario la induccin farmacolgica de este
proceso retrasa el proceso de neurodegeneracin. Estamos as mismo
interesados en la relacin de la autofagia con procesos de envejecimiento
del sistema nervioso y hemos demostrado una disminucin de la actividad
de autofagia que podra en parte estar compensado por otros mecanismos
de degradacin lisosomal como la autofagia mediada por chaperonas.
Adems y estrecha colaboracin con empresas espaolas estamos buscando nuevos productos que sean capaces de modular estos procesos.
Hemos puesto a punto varios mtodos de cribado para la determinacin
de nuevos compuestos que induzcan o bloqueen el proceso de autofagia y
que puedan luego ser aplicados a la terapia para enfermedades humanas.

Figura 1 | Figure 1
Distribucin de la poblacin total del clulas ganglionares de la retina de ratn
montada en plano y teidas para el factor de transcripcin Brn3a (A y B), y
representacin del mapa de isodensidades de nmero de clulas ganglionares (C y D).
Retinal ganglion cell distribution in mouse retinal flatmounts stained for the transcription factor
Brna (A and B). Isodensity map of retinal ganglion cell numbers (C and D).

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In our group we use cellular and animal models to


understand the role of autophagy in the physiology
and pathology of organisms. Autophagy is an
intracellular degradative process that allows the
elimination and recycling of cellular constituents.
This process is induced in many stress situations
acting as a cytoprotective response.

e want to understand why the process of autophagy is essential


to maintain cellular homeostasis and how deregulations in this
mechanism can inuence several pathological situations.

Animals decient for several autophagy regulators, the Atg genes,


die during embryonic development revealing the importance of this
process to maintain cellular homeostasis. In our group we study the
relationship of autophagy with essential processes of proliferation,
dierentiation and cell death. We have recently demonstrated that
autophagy is essential for neuronal dierentiation since autophagydecient animals generate reduced numbers of neurons in vitro and
have defects in neuritogenesis. We have also shown that autophagy
is an early cytoprotective response during several neurodegenerative
conditions. Axonal damage in autophagy-decient animals increases
cell death and conversely, pharmacological upregulation of this process
increases neuronal survival. In addtion we are interested in the role of
autophagy during the aging process in the nervous system and have
recently found a decrease in the activity of macroautophagy that seems
to be partially compensated by un upregulation of other lysosomal
pathways as chaperone mediated autophagy.
We also collaborate with several companies in the search of new
autophagy regulators. We have developed several screening methods
to nd new autophagy inducers and inhibitors that could we used as
new therapies for the treatment of human diseases.

Publicaciones Seleccionadas
Selected Publications
Esteban-Martnez L, Boya P. [2015] Autophagic flux determination in vivo
and ex vivo. Methods. Jan 30. pii: S1046-2023(15)00014-6. doi: 10.1016/j.
ymeth.2015.01.008.
Rodrguez-Muela N, Hernndez-Pinto AM, Serrano-Puebla A, Garca-Ledo L, Latorre
SH, de la Rosa EJ, Boya P [2014] Lysosomal membrane permeabilization and
autophagy blockade contribute to photoreceptor cell death in a mouse model of
retinitis pigmentosa. Cell Death Differ. 2014 Dec 12. doi: 10.1038/cdd.2014.203.

Gaband-Rodrguez E, Boya P, Labrador V, Dotti CG, Ledesma MD [2014] High


sphingomyelin levels induce lysosomal damage and autophagy dysfunction in
Niemann Pick disease type A. Cell Death Differ 2014, Jan 31 (doi: 10.1038/
cdd.2014.4).
Boya P, Diaz-Meco MT, Rubinsztein D, Sass M. [2014] Autophagy researchers.
Autophagy. Mar;10(3):393-6. doi: 10.4161/auto.27581.

Wang F, Bexiga MG, Anguissola S, Boya P, Simpson JC, Salvati A, Dawson KA.
[2013] Time resolved study of cell death mechanisms induced by amine-modified
polystyrene nanoparticles. Nanoscale. 2013 Nov 21;5(22):10868-76. doi: 10.1039/
c3nr03249c.
Boya P, Codogno P. [2013] Cell biology: Recycling in sight. Nature. 2013 Sep
5;501(7465):40-2. doi: 10.1038/501040.
Boya P, Reggiori F, Codogno P. [2013] Emerging regulation and functions of
autophagy. Nat Cell Biol. 2013 Jul;15(7):713-20. doi: 10.1038/ncb2788.

Oeste CL, Seco E, Patton WF, Boya P, Prez-Sala D [2012] Histochem Cell Biol.
2013 May;139(5):659-70. doi: 10.1007/s00418-012-1057-6.
Rodrguez-Muela N, Koga H, Garca-Ledo L, de la Villa P, de la Rosa EJ, Cuervo
AM, Boya P. [2013] Balance between autophagic pathways preserves retinal
homeostasis. Aging Cell. 2013 Jun;12(3):478-88. doi: 10.1111/acel.12072.

Biologa Celular y Molecular | Cellular & Molecular Biology

Roles of
Autophagy
in Health
and Disease

Marta Mauro-Lizcano, Lorena Esteban-Martnez, Esther Seco, Ana Serrano-Puebla,


Lucia Garcia-Ledo, Claudia Figueiredo-Pereira, Helena L A Vieira and Patricia Boya
[2014] New method to assess mitophagy flux by flow cytometry. Autophagy, in
press. http://dx.doi.org/10.1080/15548627.2015.1034403.

Financiacin | Funding
i-link0701 (CSIC 2014-2015)

SAF2012-36079 (MINECO 2013-2016)

INNPACTO IPT-010000-2010-48 (MICINN 2010-2013)


CONSOLIDER CDS2010-00045 (MICINN 2011-2016)
DSM 2013-2014

Provital 2013-2016

Figura 2 | Figure 2
Corte de una retina de ratn que
expresa constitutivamente el
marcador de autofagosomas LC3
unido a la protena fluorescente GFP
(tincin en verde). En rojo se han
marcado las mitocondrias que se han
teido utilizando el anticupero para la
protena mitocondrial TOMM20, y en
azul se observan los ncleos teidos
con el marcador para DNA DAPI.
Section of a retina from the
GFP-LC3 mouse, an animal model
that constitutively expresses the
autophagosomal marker LC3 coupled
to the fluorescent protein GFP in green.
Mitochondria stained with TOMM20 are
labelled in red and nuclei are revealed
with the DNA marker DAPI in blue.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Jess del Mazo Martnez


Investigador Cientfico
jdelmazo@cib.csic.es
PhD, 1978
Universidad Complutense de Madrid

Research Associated, 1987-1989


California Institute of Technology , CALTECH,
Pasadena L.A. (California, USA)
Profesor Honorfico, 2006
Universidad de Valparaso (Chile)
Cientfico Titular, 1981
Jefe de Grupo, 1984
Investigador Cientfico, 2006
CIB, CSIC

Otros miembros | Other lab members:


Jess Garca Lpez
Miguel Angel Brieo Enrquez
Cristina Templado Meseguer
Daro Fernndez Zoppino

Eduardo Larriba Tornel


Julio Buay Noboa
Eleni Papadopoulou

http://www.cib.csic.es/es/grupo.php?idgrupo=18

Biologa
Molecular de la
Gametognesis
Nuestro inters se ha centrado en los ltimos aos
en la biognesis y funcin de diferentes RNAs
reguladores, no-codificantes, de pequeo tamao
(tales como miRNAs, piRNAs , endo-siRNAs,
snoRNAs) en el desarrollo y la diferenciacin de la
lnea germinal y la reproduccin en mamferos y su
papel en la desregulacin gentica y epigentica
mediada por algunos reprotxicos ambientales.

os RNAs pequeos no-codicantes (sncRNAs) son considerados


como importantes reguladores postranscripcionales en el desarrollo de las clulas germinales. Adems de microRNAs (miRNAs)
endo-siRNAs y PIWIRNAs (piRNAs), otros sncRNAs: pequeos RNA
nucleolares (snoRNAs), o derivados de tRNAs o rRNAs parecen desempear importantes funciones reguladoras en la gametognesis y la
fertilizacin. Combinando secuenciacin masima (NGS), bioinformtica
y biologa celular y molecular estamos caracterizando el panorama de
expresin de sncRNAs en la diferenciacin desde clulas germinales
primordiales (PGC) hasta gametos y sus implicaciones en la fertilizacin
y el desarrollo de preimplantacin temprano. Por ejemplo, mientras
algunos snoRNAs y miRNAs se expresan abundantemente en PGCs
son reemplazados por piRNAs en espermatozoides y por endo-siRNAs
en ovocitos y cigotos. Es interesante comprobar como las secuencia de
variantes de miRNA son mas abundantes en la espermatogneis que
sus correspondientes formas cannicas.
Otras alternativas de funcionales de miRNAs como los mecanismos de
edicin de sncRNAs tambin se estn analizando en nuestro sistema.
Asi, la sustitucin mediante ADAR de Adenosina por Inosina (reconocida como guanosina por la maquinaria celular [A-to-I]) en precursores
de miRNAs, afecta el procesamiento de miRNAs. Descubrimos que la
edicin activa y la degradacin de molculas de precursor de RNAs
editados ocurre durante el perodo de perifertilization.
Tambin estamos aplicando estos estudios de regulacin gnica a la
valoracin del efecto de reprotxicos. Mltiples estudios han demostrado la asociacin entre exposicin a sustancias txicas ambientales, tales
como los llamados disruptores endocrinos, y disfunciones del desarrollo
en las clulas germinales. Estamos estudiando cmo la exposicin prenatal, incluso a bajos niveles de exposicin, a estos compuestos puede
inducir cambios epigenticos en la expresin de miRNAs en PGCs en
las siguientes generaciones no expuestas.

Figura 1 | Figure 1
Apoptosis en clulas germinales primordiales (PGCs) de ratones machos expuestos
durante su vida fetal a vinclozolina (un fungicida utilizado en la agricultura, con efectos
antiandrognicos). Co-deteccin en microscopa confocal de apoptosis por TUNEL y
marcaje especfico con SSEA-1 para clulas PGC (13,5 das postcoitum). En la parte
superior de la figura: Testis junto con el MesoNefros.
Apoptosis in primordial germ cells (PGCs) of male mice exposed during fetal life to vinclozolin
(a widely used fungicide in agriculture, with antiandrogenic effects). Examples of co-detection
by confocal microscopy analysis of apoptosis by TUNEL and SSEA-1 positive PGCs cells
(from 13.5 days post coitum embryos). Testis showed at the top of the figure along with the
MesoNephros.

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In recent years, our interest has been focused in


the biogenesis and function of various small noncoding regulatory RNAs (such as miRNAs, piRNAs,
endo-siRNAs, snoRNAs) in the development and
differentiation of the germline and reproduction in
mammals and their role in genetic and epigenetic
deregulation mediated by some environmental
reprotoxicants.

he small non-coding RNAs (sncRNAs) are considered as postranscriptional key regulators of germ cell development. In addition to microRNAs (miRNAs) endo-siRNAs and PIWI-interacting
RNAs (piRNAs), other sncRNAs generated from small nucleolar RNAs
(snoRNAs), -tRNAs or rRNAs-derevatives may also play important regulatory roles in gametogenesis and fertilization. Combining next generation sequencing (NGS), bioinformatics and cell and molecular biology
approaches we are characterizing the regulatory landscape of small
non-coding RNAs during germ cell dierentiation from primordial germ
cells (PGCs) to gameta and the consequences of the expression of the
dierent classes of these small RNAS in fertilization and early preimplantation development. Both, microRNAs and snoRNA-derived small RNAs
are abundantly expressed in PGCs but transiently replaced by piRNAs
in spermatozoa and endo-siRNAs in oocytes and zygotes. Interestingly,
miRNA sequence variants also shows an increment of non-canonical
microRNA forms along male germ cell dierentiation.

Publicaciones Seleccionadas
Selected Publications
Garca-Lpez J., Alonso L., Crdenas D.B., Artaza-Alvarez H, Hourcade J.de D.,
Martinez S., Brieo-Enrquez M. A., and del Mazo J. [2015] Diversity and functional
convergence of small non-coding RNAs in male germ cell differentiation and
fertilization. RNA, 21: 946-962.

Brieo-Enrquez M. A., Garca-Lpez J., Crdenas D. B., Guibert S., Cleroux


E., Dd L., Hourcade J.de D, Pknicov J., Weber M. and J. del Mazo [2015]
Transgenerational paternal effects of prenatal germ cell exposure to vinclozolin
are mediated by microRNAs. PLoS ONE 10(4): e0124296. doi:10.1371/journal.
pone.0124296.
J. del Mazo, J. Garca-Lpez and M. Weber [2014] Epigenetic traits of testicular
cancer: from primordial germ cells to germ cell tumors. Epigenomics, June
2014, Vol. 6, (3): 253-25.

Garca-Lpez J., Hourcade JdD, Alonso L., Crdenas D.B. and del Mazo J. [2014]
Global characterization and target identification of piRNAs and endo-siRNAs in
mouse gametes and zygotes. BBA-Gene Regulatory Mechanisms. 1839:463475.
G. M. Oresti,. J. Garca-Lpez, M. I. Aveldao and J. del Mazo [2013] Cell-typespecific regulation of genes involved in testicular lipid metabolism: fatty acidbinding proteins, diacylglycerol acyltransferases and perilipin. Reproduction
146, 471-480.
J. Garca-Lpez, M.A. Brieo-Enrquez and J. del Mazo [2013] MicroRNA
biogenesis and variability. BioMolecular Concepts. 4(4): 367380.

M. A. Brieo-Enrquez, J. Gaca-Lpez, J. del Mazo [2013] Especificidad de los


alteradores endocrinos en la expresin gnica durante el desarrollo. Revista de
Salud Ambiental, 13: 67-69.

Biologa Celular y Molecular | Cellular & Molecular Biology

Molecular Biology
of Gametogenesis

Garca-Lpez, J., Hourcade, JdD, and del Mazo, J [2013] Reprogramming of


microRNAs by adenosine-to-inosine editing and the selective elimination of edited
microRNA precursors in mouse oocytes and preimplantation embryos. Nucleic
Acids Research 41: 54835493.

J. del Mazo M.A. Brieo-Enrquez, J. Garca-Lpez, L.A. Lpez-Ferndez and M. De


Felici. [2013] Endocrine disruptors, gene deregulation and male germ cell tumors.
International Journal of Developmental Biology 57: 225 - 239.

Other functional alternatives in the miRNAs such as RNA editing mechanisms are also being analysed in our system. Adenosine-to Inosine
(A-to-I) editing represents a post-transcriptional modication of doublestranded RNA, including miRNA precursors. Inosine is recognized as
guanosine (G) by the cell machinery, which aects the subsequent
processing of edited molecules. We discovered that both active editing
and the degradation of edited precursor RNA molecules occur during the
perifertilization period.
We are also applying these basic gene regulatory aspects to the eect of
reprotoxicants. Multiple studies demonstrated the association between
exposure to environmental toxicants -such as the so-called endocrine
disruptors- and developmental dysfunctions in germ cells. We are studying how prenatal exposure to this compounds could induce induces epigenetic changes in the expression of miRNAs in PGCs in the following
non-exposed generations, even at low level of exposure.

Figura 2 | Figure 2

Financiacin | Funding
11-MRES-PNRPE-9-CVS-072-N210064934 (Ministre de lEcologie,

du Developpment Durable, des Transports et du Logement.


Repblica Francesa)
201020E016 (PIE)
BFU2013-42164-R (MINECO)

Se ha demostrado que RNAs derivados de pequeos RNAs nucleolars (snoRNAs)


pueden actuar como silenciadores de genes. SNORD21 mostr actividad del tipo
miRNA. Se muestra la prediccin de estructura de SNORD21 como ejemplo de snoRNA
detectados en las clulas germinales y cigotos y la potencial region de interaccion entre
caja C/D (posible gua para metilacin del rRNA) del snoRNAs y el rRNA.
It has been demonstrated that several small RNAs derived from snoRNAs can act as gene
silencers. Predicted secondary structures of the SNORD21 as an example of small nucleolar
RNAs (snoRNA) detected in germ cells and zygotes. SNORD21 showed miRNA activity.
Potential base pairing interactions between C/D Box snoRNAs and rRNA are showed. The
D Box motif serves as a guide for rRNA methylation.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Rosa Mara Lozano Puerto


Cientfica Titular
rlozano@cib.csic.es
PhD, 1990
Universidad Autnoma de Madrid

Postdoctoral, 1991-1993
University of California Berkeley (USA)

Investigador Contratado, 1993-2001


MEC, CIB
Cientfica Titular, 2001
Jefe de Grupo, 2008
CIB, CSIC

Investigadora del equipo | Staff scientist:


Blanca Teresa Prez-Maceda

Otros miembros | Other lab members:


Mara Encarnacin Lpez Fernndez
Natalia Soledad Fagali

http://www.cib.csic.es/grupo.php?idgrupo=67

Reconocimiento Clula-Biomaterial
Se investiga la respuesta celular y molecular de la clula al interaccionar con materiales metlicos y cermicos de
aplicacin en reparacin sea. Entre los metlicos se analizan aleaciones de Cobalto-Cromo y de biodegradables
de base Magnesio y, entre los cermicos, las hidroxiapatitas. Se estudian los efectos de las partculas metlicas del
desgaste-corrosin del material implantado. En cermicos, se disean superficies con distintas protenas.

as aleaciones Cobalto-Cromo con alto contenido en carbono


(CoCrHC) se proponen como material alternativo para prtesis
de cadera. Las bajas tasas de desgastecorrosin que presentan
stas aleaciones contrastan con las del par polietileno/metal, ampliamente utilizado en clnica y que se caracteriza por generar gran cantidad de
partculas en el lugar del implante, que se han relacionado con procesos
de osteolisis y prdida nal de la prtesis lo que ha motivado la bsqueda
de otros materiales. Es por esta razn por la que se propone el estudio de
las aleaciones de CoCrHC y del efecto de las partculas que se generan
como consecuencia del proceso de tribocorrosin de este material. Los
efectos producidos en la clula por los productos derivados del desgastecorrosin de las aleaciones CoCrHC, partculas e iones, se evalan en las
lneas celulares representativas del entorno de la prtesis osteoarticular.
El Mg y sus aleaciones son materiales interesantes debido a las propiedades que presentan: son ligeros, su mdulo elstico y densidad

son semejantes a los del hueso, son reabsorbibles, sus productos de


corrosin no son txicos y son fcilmente excretados en la orina. Sin
embargo, la principal limitacin reside en que la velocidad de degradacin de stos es mayor que la velocidad necesaria para la regeneracin
del tejido. Se pretende, mediante la aplicacin de tratamientos, controlar
la velocidad de degradacin y disear materiales de base Mg cuya
reabsorcin est sincronizada con la regeneracin del tejido seo a
reparar. Se estudia adems, el efecto de las partculas de Mg sobre la
respuesta celular.
Se disean nuevas supercies biomimticas sobre materiales cermicos
como son las hidroxiapatitas sustituidas con silicio mediante la funcionalizacin con distintas protenas, como son ciertos factores de crecimiento que favorecen la vascularizacin del tejido estimulando la actividad
biolgica de otros pptidos implicados en el crecimiento del tejido seo.

Figura 1 | Figure 1
Microscopa multidimensional en tiempo real in vivo de macrfagos J774 en presencia de partculas de magnesio. Imgenes del cultivo de macrfagos expuesto a las partculas de
Mg (1 mg/ml; negro) durante distintos tiempos (0 y 24 horas). A las 24 horas, los macrfagos se dividen y algunos mueren al interaccionar con las partculas que se van degradando
durante el cultivo. (Referencia Alvarez F. y colab. en Microscopy: advances in scientific research and education).
In vivo time-lapse multidimensional microscopy of macrophages J774 in presence of magnesium particles. Images of macrophages culture exposed to Mg particles (1 mg/ml; black images) at
different times (0 and 24 hours). At 24 h., some macrophages duplicate and those located close to particles displayed morphological changes that developed in cell death. Mg particles degrade and
become clear. (Reference Alvarez F. et al. in Microscopy: advances in scientifc research and education).

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Research is focus in the analysis of the molecular and cell response upon interaction with metallic and ceramic
materials for application in bone tissue repair. Metallic materials as cobalt-chromium alloys with high carbon
content and some biodegradable magnesium-base, and ceramics as hydroxyapatites are selected. The effect on the
cell of metallic debris, particles and ions, are under study. In ceramics, new surfaces are designed with proteins.

otal hip replacement by metallic biomaterials with loss of function


has become an important concern in human health. The most
widespread clinical substitution for hip substitution is given by
the polyethylene/metal joint replacement. However, excessive wear of
polyethylene causes the production of particles that is believed to be the
major cause of the progressive osteolysis and subsequent loosening of
prosthesis. This problem has strongly driven the use of Metal on Metal
(MoM) combinations as a replacement in joint prosthesis, specically,
made of CoCr alloys, due to their substantially low corrosion and wear
rates. But unfortunately, there are still wear particles and ions that are
released in the body with these implants. The eect of the wear debris
and ions derived from the tribocorrosion process of the MoM in the cell
response are under analysis in studies with those cell lines that better
represent the osteoarticular prostheses cell microenvironment.

osteoconductivity. The density, elastic modulus and compressive strength


properties of Mg are more similar to bone. Mg is a necessary element for
the incorporation of calcium to bone and to stimulate the growth of new
tissue, is non-toxic and degrades in body uids, making suitable for orthopedic applications. In spite of the desirable properties, Mg-based materials have very high corrosion rate that need to be controlled. Research
pretends with dierent approaches synchronize the degradation kinetic of
Mg-based materials and bone tissue repair and analyze the eect of Mg
particles on cell response.

Biologa Celular y Molecular | Cellular & Molecular Biology

Cell-Biomaterial Recognition

New ceramic materials hydroxyapatite-based are under study upon functionalization with proteins, as are several growth factors that promotes
tissue vascularization and increase the biological activity of other peptides
involved in bone tissue growth.

Magnesium (Mg) and its alloys are biodegradable materials suitable


for bone repair application due to its biodegrability, reabsorbability and

Publicaciones Seleccionadas
Selected Publications
Lozano RM*, Prez-Maceda BT, Carboneras M, Onofre-Bustamante E, GarcaAlonso MC, Escudero ML [2013] Response of MC3T3-E1 osteoblasts, L929
fibroblasts and J774 macrophages to fluoride surface-modified AZ31 magnesium
alloy. Journal of Biomedical Materials Research: Part A. 101: 2753-2762.
*Corresponding author.
Alvarez F, Lozano Puerto R, Prez-Maceda B, Grillo C, Schilardi P, Fernndez
Lorenzo M [2013] Efecto de micropartculas de Mg con y sin tratamiento con
KF en clulas osteoblsticas y macrofagos. The Journal of the Argentine
Chemical Society. 100: 48-52.

Billi F, Iglesias C, Onofre E, Lozano RM, Prez-Maceda B, Rubio JC, Escudero ML,
Garca-Alonso MC [2013] Characterization of oxidized TiAlV after fretting-corrosion
tests using near-field microscopy. British Journal of Surgery. Abstract. 100
(Suppl. 1): 13.

Bodeln O, Iglesias-Urraca C, Daz I, Lozano RM, Prez-Maceda BT, Clemente C,


Alobera MA, Garca-Alonso MC, Rubio Surez JC, Escudero ML. [2014] Analysis
of metallic traces from biodegradation of AZ31 magnesium alloy in rat organs.
British Journal of Surgery. Abstract. 101 (Suppl. 1): 6.

Iglesias-Urraca C, Bodeln O, Daz I, Lozano RM, Prez-Maceda B-T, Clemente C,


Alobera MA, Garca-Alonso MC, Rubio Surez JC, Escudero ML. [2014] Clinicalradiological and histological correlation of AZ31 alloy used as a prosthetic implant.
British Journal of Surgery. Abstract. 101 (Suppl. 1): 6.
Alvarez F, Lozano Puerto RM, Prez-Maceda BT, Grillo CA, Fernndez Lorenzo MA
[2014] Effect of Mg particles on MC3T3-E1 and J774 cellular cycle. Influence of
fluoride treatment. European Cells and Materials. Vol 28 (Suppl 3): 65.

Alvarez F, Lozano Puerto RM, Prez-Maceda BT, Grillo CA, Fernndez Lorenzo MA
[2014] Multidimensional microscopy: A suitable technique to follow in vivo the
interaction between biodegradable biomaterials and cells. In Microscopy:
advances in scientific research and education Microscopy book n
6 Vol.1. Editor: A. Mndez-Vilas. Editorial: Formatex Research Center (Badajoz,
Espaa): 523-529.

Financiacin | Funding
MAT2011-29152-C02-02 (MINECO)

CAM S2009/MAT1472 (CAM) Grupo asociado

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Biologa Celular y Molecular | Cellular & Molecular Biology

Susana Moreno Daz de la Espina


Investigadora Cientfica
smoreno@cib.csic.es
PhD, 1975
Universidad Complutense Madrid
Postdoctoral, 1976-1978
DKFZ (Heidelberg, Alemania)

Visiting Scientist, 1987


NCI/NIH. Frederick (MD, USA)

Cientfica Titular, 1979


Jefa de Grupo, 1981
Jefa de Laboratorio, 1997
Investigadora Cientfica, 2002
CIB, CSIC

Otros miembros | Other lab members:


Malgorzata Ciska
Ana Ugidos Valladares
Mercedes Carnota Romero

http://www.cib.csic.es/es/grupo.php?idgrupo=31

Matriz Nuclear y Regulacin de la


Organizacin y Funcionalidad Nuclear
Nuestro objetivo es el anlisis de la lmina nuclear de plantas. Hemos determinado los anlogos funcionales
de las laminas de metazoos, las protenas NMCP y caracterizado sus dos homlogos NMCP1 y NMCP2 en la
monocotilednea Allium cepa y su interaccin con las protenas SUN que son las nicas protenas asociadas a
laminas conservadas en plantas y forman parte de los complejos que unen el ncleoesqueleto y citoesqueleto en
metazoos y plantas.

a lmina nuclear est muy conservada en eucariotas, aunque slo


bien caracterizada en metazoos; sus principales componentes,
las laminas, no estn conservados en otros eucariotas. En nuestro laboratorio investigamos las protenas que componen la lmina en
plantas que carecen de genes de laminas. Hemos analizado varias protenas candidatas a realizar funciones de laminas en plantas. El anlisis
bioinformtico de las protenas NMCP, unas de las principales candidatas, revel que se trata de una familia muy conservada que comparte
muchas caractersticas con las laminas de metazoos como son los dos
tipos bsicos, la estructura tripartita con un largo segmento central coiled coil con capacidad de dimerizacin y varias secuencias especcas
conservadas (Fig 1; Ciska et al., JXB 2013), que unido a su capacidad
de dimerizar y formar lamentos, localizacin en la lmina, asociacin a
protenas SUN, expresin regulada durante el desarrollo e implicacin en
algunas de las funciones de las laminas como la regulacin del tamao

Figura 1 | Figure 1
Estructura de NMCPs y laminas. Ambas presentan una distribucin similar de coiled
coils (cajas naranja), regiones conservadas (barras verdes), sitios para cdk1 (barras
rojas), NLS (cajas verdes) y un segmento de aa cidos (caja roja). Las NMCPs carecen
de plegamiento Ig (elipse negra) y la caja CAAX de laminas pero tienen el extremo
C-terminal conservado. Regiones que dirigen las NMCP a la EN (*).

y forma nuclear, organizacin de la heterocromatina y asociacin del


ncleoesqueleto al citoesqueleto, nos han permitido establecer que
constituyen los anlogos funcionales de las laminas en plantas (Ciska
and Moreno Daz de la Espina, PSB 2013). La produccin de anticuerpos contra dominios conservados de las protenas nos ha permitido
determinar las caractersticas de las protenas NMCP1 y NMCP2 endgenas en Allium cepa, su localizacin nuclear y presencia en el ncleoesqueleto. Estamos caracterizando tambin en este sistema las protenas
que interaccionan con ellas, principalmente las SUNs. Todos estos datos
junto con otros de la literatura nos han llevado a establecer por primera
vez un modelo de organizacin de la lmina en plantas (Fig 2; Ciska and
Moreno Daz de la Espina, FPS 2014). El grupo trabaja integrado en el
International Plant Nucleus Consortium (http://bms.brookes.ac.uk/ipnc).

Financiacin | Funding
BFU2010-15900 (MINECO)
PIE 201020E019 (CSIC)

Structural analogies of NMCPs and lamins. Both display a similar distribution of coiled coils
(orange boxes), conserved regions (green bars), cdk1 phosphorylation sites (red bars), a NLS
(green boxes) and a stretch of acidic aminoacids (red boxes). NMCPs lack the Ig fold (black
ellipse) and the CAAX box of lamins, but have a conserved C-terminus. Regions mediating NE
localization of NMCPs (*).

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The plant lamina and its main interacting partners. The NMCP-based lamina binds to NPCs through Nup136 and NUA, and associates to nucleocytoplasmic linkers by binding of NMCPs to SUNs
and plant specific KASH proteins (WIP). These complexes connect the lamina with the actin cytoskeleton and the -TuC complexes but also anchor proteins to the ONM (RanGAP). Not elucidated
interactions (?).

Nuclear Matrix
and Regulation
of the Nuclear
Organization and
Function
Our aim is the analysis of the plant nuclear lamina.
We have determined the functional analogs of
metazoan lamins in plants, the NMCP protein
family and analyzed the two homologs MMCP1 and
NMCP2 in the monocot Allium cepa, as well as their
interactions with SUN proteins that are the only
lamin-binding proteins conserved in plants and form
the protein complexes that link the nucleoskeleton
and cytoskeleton in metazoan and plants.

Publicaciones Seleccionadas
Selected Publications
Ciska M, Moreno Diaz de la Espina S [2014] The intriguing plant nuclear lamina.
Front Plant Sci. 5, 166. DOI: 10.3389/fpls.2014.00166.
Gasset-Rosa F, Coquel AS, Moreno-del lamo M, Chen P, Song X, Serrano AM,
Fernndez-Tresguerres ME, Moreno-Daz de la Espina S, Lindner AB, Giraldo R

he nuclear lamina is highly conserved in eukaryotes, but only


well characterized in metazoa. The main components of the
metazoan lamina are the lamins that are not conserved in other
eukaryotes. We investigate the proteins that form the lamina in plants
that lack genes for lamins. We have analyzed several protein candidates
to play lamin functions in plants. The bioinformatic analysis of NMCP
proteins, the main candidates to play lamin functions in plants, revealed
that this is a highly conserved family of proteins that share multiple
characteristics with metazoan lamins as are the two basic types, the
tripartite structure with a long central coiled coil domain with dimerization ability, and several conserved specic domains (Fig 1; Ciska et al.,
JXB 2013), which along with their ability to dimerize and form laments,
localization in the lamina, binding to SUN proteins, developmentally
regulated expression and involvement in some of the nuclear functions
regulated by lamins such as nuclear shape and size determination,
heterochromatin organization and association of the nucleoskeleton to
cytoskeleton allowed us to establish that they constitute the functional
analogs of metazoan lamins in plants (Ciska and Moreno Daz de la
Espina, PSB 2013). Antibodies produced against conserved regions of
NMCP1 and NMCP2 allowed us the characterization of the endogenous
proteins in this system and the determination of their nuclear localization and presence in the nucleoskeleton. We are also characterizing the
proteins interacting with NMCPs in this system, mainly the SUNs. The
above results along with others in the literature allowed us to establish
for the rst time a model for the organization of the plant nuclear lamina
(Fig 2; Ciska and Moreno Daz de la Espina, FPS 2014). The groups
belongs to the International Plant Nucleus Consortium (http://bms.
brookes.ac.uk/ipnc)

Biologa Celular y Molecular | Cellular & Molecular Biology

Figura 2 | Figure 2
La lmina vegetal y sus principales interacciones. La lmina de protenas NMCP se une a los PNC a traves de Nup136 y NUA, y se asocia a los complejos que conectan ncleo y
citoplasma por unin de las NMCPs a SUNs y protenas KASH especficas (WIP). Estos complejos conectan la lmina con el citoesqueleto de actina y los complejos -TuC pero
tambin anclan protenas a la MNE (RanGAP). Interacciones no aclaradas (?).

[2014] Direct assessment in bacteria of prionoid propagation and phenotype


selection by Hsp70 chaperone. Mol Microbiol 91: 1070-1087.

Ciska M, Moreno Daz de la Espina S [2013] NMCP/LINC proteins. Putative lamin


analogs in plants? Plant Signaling & Behavior 8: 12e26669.

Ciska M, Masuda K, Moreno Daz de la Espina S [2013] Lamin-like analogues in


plants: the characterization of NMCP1 in Allium cepa. J Exp Bot 64: 1553-1564.

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Biologa Celular y Molecular | Cellular & Molecular Biology

Jos Luis Barbero


Esteban

Investigador Cientfico
jlbarbero@cib.csic.es
PhD, 1981
Universidad Complutense de Madrid

Lucas Snchez
Rodriguez

Associate Research, 1984


NYU Medical Center. Pathology Department.
Dr. Angel Pellicer laboratory (New York, USA)

Profesor de Investigacin
lsanchez@cib.csic.es
PhD, 1976
Universidad Complutense de Madrid

Postdoctoral, 1977-1979
Investigador Asociado, 1979-1981
Zoological Institute University of Zurich

Researcher, 1983-1996
Pharmacia/Antibioticos Pharma

Jefe de Grupo, 1981-1984


European Molecular Biology Laboratory
(Heidelberg, Alemania)

Group Leader, 1996-2006


Pharmacia/Department of Immunology
and Oncology. CNB.

Cientfico Titular, 1985


Investigador Cientfico,1989
Profesor de Investigacin, 2004
CIB, CSIC

Investigador Cientfico, 2006


CIB, CSIC

Otros miembros | Other lab members:


http://www.cib.csic.es/es/grupo.php?idgrupo=64

Dinmica Cromosmica
en Meiosis
El complejo de cohesinas y el control de la dinmica
de dicho complejo en la cromatina son esenciales
para la correcta segregacin cromosmica. Errores
en estos mecanismos conducen a la muerte celular,
patologas como el sndrome de Down, la formacin
de tumores, la infertilidad y otras cohesinopatas.

n colaboracin con los grupos de AM Pends (Centro de


Investigacin del Cancer, Salamanca) y de JA Suja (Universidad
Autnoma de Madrid) se han estudiadado diferentes protenas
denominadas cohesin-regulators que controlan la dinmica del complejo
de cohesinas, no solo durante la segregacin cromosmica, sino tambin
en procesos de control de la expresin gnica en los que las cohesinas
actan modelando la estructura de ciertas regiones de los cromosomas.
Esencialmente hemos estudiado la topoisomerasa alfa-2, las acetiltransferasas ESCO1 y ESCO2 y la histona quinasa haspin en mamferos. Por otra
parte, nuestra investigacin se ha centrado en estudiar en colaboracin
con diferentes laboratorios, las patologas producidas como consecuencia
de la falta de funcin de la cohesina especca de meiosis STAG3, identicada y caracterizada por primera vez en 2001 en mi laboratorio. El estudio
en ratones decientes en STAG3 demostr la necesidad de su funcin para
la fertilidad (referencia 7). El descubrimiento de mutaciones en humanos
de la STAG3 que conducen a la patologa denominada como Premature
ovarian failure y su posible implicacin en cncer de ovario se ha publicado en la prestigiosa New England J. Med. con la participacin de nuestro
laboratorio (referencia 6). L. Snchez est trabajando sobre la evolucin de
los mecanismos de determinacin sexual.

Maria Fernanda Ruiz Lorenzo

Chromosomal Dynamics
in Meiosis
Are there link between human syndromes with
physical and mental problems, a tumor growing out
of control and the incapability to contribute to next
generation? This question can be answered if we
look at the biological functions of a protein complex,
named cohesin.

n collaboration with the groups of AM Pends ( Cancer Research


Center, Salamanca) and of JA Suja (Autonoma University of Madrid)
we have study the function of dierent proteins called cohesinregulators that control the dynamics of the cohesin complex, not only
during the chromosomal segregation, but also in processes of control
of the gene expression in which the cohesins act shaping the structure
of certain regions of the chromosomes, such as the topoisomerase
II, the acetyltransferases ESCO1 and ESCO2 and of the histone-kinase
haspin in mammals. On the other hand, our recent research has centered on studying, in collaboration with dierent laboratories, the pathologies produced as consequence of the lack of function of the meiosis
specic cohesin STAG3, which was identied and characterized by the
rst time in 2001 in my laboratory. The study in decient mice in STAG3
demonstrated the need of his function for the fertility (reference 7). The
discovery of mutations in human STAG3, which drive to the pathology
named as Premature ovarian failure and his possible implication
in cancer of ovary has been published in the prestigious journal New
England J. Med. with the participation of our laboratory (reference 6). L.
Snchez is working on the evolution of sex-determining mechanisms.

Publicaciones Seleccionadas
Selected Publications
Barbero JL [2013] Cohesin Complexes: Modulators of Chromatin Organization
Control Gene Expression in Immune System. In: Advances in Medicine and
Biology Vol. 58.pp. 167-176. Editor: Leon V. Berhardt. Nova Science Publisher,
Inc. New York. USA. ISBN: 978-1-62257-803-0.
Calvente A, Viera A, Parra MT, de la Fuente R, Suja JA, Page J, Santos JL, Garca de la
Vega C., Barbero JL, Rufas JS [2013] Dynamics of cohesin subunits in grasshopper
meiotic divisions. Chromosoma 122: 77-91.(DOI 10.1007/s00412-012-0393-6).

Barbero JL [2013] Genetic basis of cohesinopathies. The Application of


Clinical Genetics 6:15-23.

Roco Gmez, Alberto Viera, Ins Berenguer, Elena Llano, Alberto M. Pends, Jos
Luis Barbero, Akihiko Kikuchi and Jos A. Suja [2014] Cohesin removal precedes
topisomerase II -dependent decatenation at centromeres in male mammalian
meiosis II. Chromosoma. 123:129-146. DOI 10. 1007/s00412-013-0434-9.

Figura 1 | Figure 1
Implicacin del complejo de cohesinas y sus reguladores en las patologas humanas
denominadas cohesinopatas.
Cohesin and cohesin regulators in human cohesinopathies.

Financiacin | Funding
Bases Moleculares de la Aneuploida en Cncer: Control de la Segregacin
y Estabilidad Cromosmica. AP 98712012 (2012-2014) FUNDACIN DE

INVESTIGACIN MDICA MUTUA MADRILEA.

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Barbero, JL [2014] Molecular Genetics of Cohesinopathies. In: eLS. John


Wiley&Sons Ltd, Chichester. doi: 10.1002/9780470015902.a0025309.

Caburet S., Arboleda VA, Llano E., Overbeek PA, Barbero JL, Oka K., Harrison W.,
Vaiman D., Ben-Neriah Z., Garcia-Tuon I., Fellous M., Pends AM, Veitia RA and
Vilain E [2014] Mutant Cohesin in Premature Ovarian Failure. New England J.
Med. 370:943-949.

Llano E., Gmez-H L.,Garca-Tuon I., Snchez Martn M., Caburet S., Barbero JL,
Schimenti JC, Veitia RA and Pends AM [2014] STAG3 is a strong candidate gene
for male infertility. Human Mol. Genet. 23:3421-3431.

Ruiz, M.F., Sarno, F., Zorrilla, S., Rivas, G. and Snchez, L. (2013) Biochemical and
functional analysis of Drosophila-Sciara chimeric Sex-lethal proteins. PLoS ONE
8(6): e65171.
Snchez, L. (2014) Sex determining mechanisms in insects based on imprinting
and elimination of chromosomes. Sexual Development 8: 83-103.

24/6/15 12:54

PhD, 2005
Universidad Complutense de Madrid

Postdoctoral, 2005-2009
MRC-Laboratory of Molecular Biology
(Cambridge, UK)
Postdoctoral, 2009-2012
Investigadora Ramn y Cajal, 2012
CIB, CSIC

Publicaciones Seleccionadas
Selected Publications
Oliva MA, Martin-Galiano AJ, Sakaguchi Y, Andreu JM [2012] Tubulin homolog TubZ
in a phage partition system. Proc Natl Acad Sci USA 109 (20):7711-6.

http://www.cib.csic.es/en/grupo.php?idgrupo=43

Financiacin | Funding
RYC-2011-07900 (Ministerio de Ciencia e Innovacin)

BFU2013-47014-P (MINECO, cofinanciado con fondos FEDER)

Tubulinas y FtsZ:
Modulacin del
Ensamblaje de Protenas
Bases moleculares de la segregacin de factores de
virulencia por sistemas de particin tipo III.

l xito de los factores de virulencia radica en su habilidad para


mantenerse en las bacterias. Los sistemas de particin ayudan al
posicionamiento de estos elementos durante la divisin. Hemos
identicado un sistema tipo III en el fago c-st de C. Botulinum (codica
para el botox-C), donce el complejo nucleoproteico tubCR permite
anclar el fago a la protena motora TubZ para su movilizacin. Adems
descubrimos la existencia de un regulador conservado, TubY. Ahora nos
centramos en entender cmo funciona el sistema.

Francisco Javier Ruiz Dueas


Investigador Ramn y Cajal
fjruiz@cib.csic.es
PhD, 1999
Universidad Complutense de Madrid

Postdoctoral, 1999-2000
Centro di Ricerca di Risonanze Magnetiche,
CERM (Florencia, Italia)
Postdoctoral, 2001-2002
Investigador I3P, 2003-2005
Investigador contratado, 2006-2009
Investigador Ramn y Cajal, 2010
CIB, CSIC
http://www.cib.csic.es/lignina/lignina_en.html

Financiacin | Funding
RYC-2009-04798
KBBE-2010-4-265397 (EC FP7)
BIO2011-26694 (MICINN)

KBBE-2013-7-613549 (EC FP7)

CSP15-1609 (U.S. Department of

Energy Joint Genome Institute,


DOE-JGI)

Biotecnologa para la
Biomasa Lignocelulsica
Bsqueda e ingeniera de nuevas peroxidasas
fngicas de alto potencial redox.

uestra actividad cientca se centra en la obtencin de nuevas


peroxidasas con propiedades catalticas de inters que se
puedan emplear en procesos industriales de oxidacin, en sustitucin de reactivos qumicos agresivos. Para ello se ha diseado una
estrategia que consiste en la bsqueda de genes de nuevas enzimas de
tipo peroxidasa en secuencias de genomas fngicos, y en el posterior
diseo a medida de sus propiedades catalticas y estabilidad mediante
el empleo de tcnicas de ingeniera de protenas.

05 _ DPTO BIOLOGIA CELULAR Y MOLECULAR _ 15-06-24.indd 121

Oliva MA, Andreu JM [2014] Tubulin and FtsZ superfamily of protein assembly
machines. (review) In: eLS, Encyclopedia of Life Sciences (e-Book chapter;
John Wiley & Sons, Ltd. Chichester) doi: 10.1002/9780470015902.a0025586.

Andreu JM, Oliva MA [2013] Purification and assembly of bacterial tubulin BtubA/B
and constructs bearing eukaryotic tubulin sequences. Methods in Cell Biology
115, 269-281.

Tubulins & FtsZ:


Targeting Proteins
Self-Assembly
Molecular basis of virulence factors segregation by
type III partition systems.

he extraordinary success of virulence elements relies on their


stable maintenance in bacteria. Partition systems are positioning
systems that evenly distribute those DNAs during division. We
identied a type III partition system in Clostridium botulinum phage c-st
(encodes Botox-C), where the centromere-like tubC is recognized by
TubR and anchored to tubulin-like TubZ to mobilise the phage. We also
discovered a conserved regulator, TubY. Now we focus on the understanding of how this system works.

Programa Ramn y Cajal | Ramn y Cajal Program

Mara A. Oliva Blanco


Investigadora Ramn y Cajal
marian@cib.csic.es

Publicaciones Seleccionadas
Selected Publications
Barrasa JM, Blanco MN, Esteve-Ravents F, Alts A, Checa J, Martnez AT, RuizDueas FJ [2014] Wood and humus decay strategies by white-rot basidiomycetes
correlate with two different dye decolorization and enzyme secretion patterns on
agar plates. Fungal Genet Biol 72: 106-114.

Fernndez-Fueyo E, Ruiz-Dueas FJ, Martnez MJ, Romero A, Hammel KE, Medrano


FJ, Martnez AT [2014] Ligninolytic peroxidase genes in the oyster mushroom
genome: heterologous expression, molecular structure, catalytic and stability
properties, and lignin-degrading ability. Biotechnol. Biofuels 7:2.

Ruiz-Dueas FJ, Lundell T, Floudas D, Nagy LG, Barrasa JM, Hibbett DS, Martnez
AT [2013] Lignin-degrading peroxidases in Polyporales: an evolutionary survey
based on ten sequenced genomes. Mycologia 105: 14281444.

Hori C, Ishida T, Igarashi K, Samejima M, Suzuki H, Master E, Ferreira P, Ruiz-Duenas


FJ, Martnez AT, Covert S, Blanchette B, Cullen C [2014] Analysis of the Phlebiopsis
gigantea genome, transcriptome and secretome provides insight into its pioneer
colonization strategies of wood. PLOS Genetics 10: Pages: e1004759.

Linde D, Coscoln C, Liers C, Hofrichter M, Martnez AT, Ruiz-Dueas FJ [2014]


Heterologous expression and physicochemical characterization of a fungal dyedecolorizing peroxidase from Auricularia auricula-judae. Protein Expr Purif 103:
28-37.

Biotechnology for
Lignocellulosic Biomass
Screening and engineering of new high redoxpotential fungal peroxidases.

ur research activity aims to obtain novel peroxidases with catalytic properties of interest that can be used in industrial oxidation processes, substituting harsh chemical reagents. To attain
this objective, a strategy has been designed consisting of searching
for genes encoding new type-peroxidase enzymes in fungal genome
sequences, and subsequent tailored designing of their catalytic properties and stability by using protein engineering techniques.

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