Susan J Baserga | Maria Carmo-Fonseca | Ingrid Grummt | Danièle Hernandez-Verdun |
Julian A Hiscox | Denis L J Lafontaine | Angus I Lamond | David A Matthews | Jo Milner |
Mark O J Olson| Craig S Pikaard | John J Rossi |Carlos Rubbi | Peter Shaw | Michael Taliansky |
Robert Y Tsai | Elisa Varela | Robert J White | Adrian Whitehouse |
Susan J Baserga
Ribosome biogenesis in eukaryotic cells
Departments of Molecular Biophysics & Biochemistry
Genetics and Therapeutic Radiology
Yale University School of Medicine
New Haven, CT 06520-8024
USA
Email: susan.baserga@yale.edu
Biography
Susan Baserga received a B.S. in Biology from Yale College and an M.D. and Ph.D. (Human Genetics) from Yale in 1988. She conducted post-doctoral training in the Department of Molecular Biophysics and Biochemistry at Yale with Joan Steitz. Susan is currently an Associate Professor at Yale University with a primary appointment in Molecular Biophysics & Biochemistry and joint appointments in the Departments of Genetics and Therapeutic Radiology. The focus of Susan’s research is on the function of ribonucleoproteins in pre-rRNA processing and pre-ribosome assembly. Her research is principally funded by the National Institutes of Health and the National Science Foundation.
Abstract
Ribosome biogenesis is a complex process requiring the coordinated expression of rRNA and protein moieties and their assembly in the eukaryotic nucleolus. In order to better understand each aspect of this process, we are using an array of genetic, biochemical, and cell biological techniques in the yeast Saccharomyces cerevisiae. My laboratory focuses on the role of the ribonucleoprotein and protein complexes involved in generating the mature rRNAs. Using innovative proteomics techniques, my laboratory has recently identified the protein components of a large nucleolar ribonucleoprotein that is required for processing of the 18S small subunit rRNA. This RNP, which we termed the SSU processome, is composed of the U3 snoRNA and 40 proteins. Currently, projects in the lab are aimed at determining the architecture of this RNP and the functions of individual proteins in 18S processing. We approach this question from several perspectives, using genetic and biochemical methods to identify direct interactions between components, and cryo electron microscopy to visualize the complex in three dimensions. Through these studies we have discovered and characterized several unique protein motifs and their specific roles in rRNA processing.
Key primary references
Granneman, S., Bernstein, K.A., Bleichert, F. & Baserga, S.J. A comprehensive mutational analysis of yeast DExD/H box RNA helicases required for 18S rRNA synthesis. In press for Molecular and Cellular Biology.
Gallagher, J.E., Dunbar, D.A., Granneman, S., Mitchell, B.M., Osheim, Y., Beyer, A.L. & Baserga, S.J. (2004) RNA polymerase I transcription and pre-rRNA processing are linked by specific SSU processome components. Genes & Dev. 18; 2506-2517.
Osheim, Y.N., French, S.L., Keck, K.M., Champion, E.A., Spasov, K., Dragon, F., Baserga, S.J. & Beyer, A.L. (2004) Pre-18S ribosomal RNA is structurally compacted into the SSU processome prior to being cleaved from nascent transcripts in Saccharomyces cerevisiae. Mol. Cell 16; 943-954.
Dragon, F., Gallagher, J.E.G., Compagnone-Post, P.A., Mitchell, B.M., Porwancher, K.A., Wehner, K.A., Wormsley, S., Settlage, R.E., Shabanowitz, J., Osheim, Y., Beyer, A.L., Hunt, D.F. & Baserga, S.J. (2002) A large nucleolar U3 ribonucleoprotein required for 18S rRNA biogenesis. Nature 417; 967-970.
Key reviews
Granneman, S. & Baserga, S.J. (2005) Crosstalk in gene expression: coupling and co-regulation of rDNA transcription, pre-ribosome assembly and pre-rRNA processing. Curr. Op. Cell Biol. 17; 281-286.
Granneman, S. & Baserga, S.J. (2004) Ribosome biogenesis: of knobs and RNA processing. Exp. Cell Res. 296; 43-50.
The nucleolus and RNA editing
Instituto de Medicina Molecular
Faculdade de Medicina
Universidade de Lisboa
Av. Prof. Egas Moniz 1649-028 Lisboa
Portugal
Email: carmo.fonseca@fm.ul.pt
Biography
Maria Carmo-Fonseca obtained her MD in 1983 and the PhD in Cell Biology in 1988. Since 1998, she is Professor at the Faculty of Medicine, University of Lisbon. Carmo-Fonseca did a post-doc at EMBL (Heidelberg, Germany) from 1989 to 1992. Since 2002 she serves as Executive Director of the Institute of Molecular Medicine in Lisbon, a national center of excellence for biomedical research. Carmo-Fonseca is also head of the Cell Biology Department at this Institute. The focus of Carmo’s research is the regulation of gene expression within the sophisticated environment that is the nucleus of a living cell. The group is particularly interested in understanding how mRNA biogenesis is regulated and how errors in this process lead to human disease.
Abstract
A defining feature of eukaryotic cells is the generation of protein diversity either post-transcriptionally by alternative splicing and RNA editing or post-translationally by modification of amino acids in proteins. One of the most recently discovered post-translational modification mechanism in eukaryotes involves the covalent attachment of the small ubiquitin-like modifier, SUMO, to target proteins. We have recently described that proteins modified by SUMO-1 are present in the nucleolus, that SUMO-1 in the nucleolus co-localizes with the RNA editing enzyme ADAR1 and that this enzyme represents a novel substrate for sumoylation. ADAR1 (adenosine deaminase that acts on RNA) is a member of the family of enzymes that catalyse the conversion of adenosine to inosine in double-stranded RNA. In mammals there are three ADAR enzymes, termed ADAR1, ADAR2, and ADAR3. Inactivation of editing enzymes in mice and in the fruit fly has resulted in profound neurological phenotypes. Several lines of evidence suggest that ADAR activity is tightly controlled in the cell. In our work we demonstrate that ADAR1 is modified by SUMO at lysine residue 418. Substitution of this amino acid residue by arginine, which cannot be modified by SUMO, affects the editing activity of the enzyme. Our finding that ADAR1 co-localizes with SUMO-1 in the nucleolus and that sumoylation of ADAR1 reduces editing activity suggests that the nucleolus represents a “sink” for inactive ADAR1 in the cell.
Key primary references
Desterro, J., Keegan, L., Lafarga, M., Berciano, M.T., O’Connell, M., & Carmo-Fonseca, M. (2003) Dynamic association of RNA editing enzymes with the nucleolus. J. Cell Sci. 116; 1805-1818.
Desterro, J.M., Keegan L.P., Jaffray, E., Hay R.T., O’Connell, M.A. & Carmo-Fonseca, M. (2005) SUMO-1 Modification Alters ADAR1 Editing Activity. Mol. Biol. Cell. 16; 5115-5126.
Key review
Carmo-Fonseca, M., Mendes-Soares, L., and Campos, I. (2000) To be or not to be in the nucleolus. Nature Cell Biol. 2; E107-E112.
Regulation of ribosomal RNA gene transcription
Molecular Biology of the Cell II
German Cancer Research Center
Im Neuenheimer Feld 280
D-69120 Heidelberg
Germany
Email: I.Grummt@DKFZ-Heidelberg.de
Biography
Ingrid Grummt received her PhD in 1970 at the Humboldt-University in Berlin, worked as a Postdoc at the German Academy of Sciences in Berlin-Buch and the Max-Planck-Institute of Biochemistry in Munich, led a research group since 1977, and is Head of a Research Division at the German Cancer Research Center in Heidelberg. Her research focuses on the molecular mechanisms that regulate RNA polymerase I transcription.
Abstract
Regulation of rRNA synthesis is a key factor that controls cell growth and balances the effects of growth factors to prevent uncontrolled cell proliferation. The key factor that directs growth-dependent regulation of rDNA transcription is TIF-IA, a transcription factor whose activity fluctuates in response to cell growth and serves a key function in adapting cellular biosynthetic activities to cell growth. We have inactivated the murine TIF-IA gene by homologous recombination in mice and embryonic fibroblasts (MEFs). TIF-IA-/- embryos die before/at embryonic day 9.5, displaying retardation of growth and development. In MEFs, Cre-mediated depletion of TIF-IA leads to disruption of nucleoli, cell cycle arrest, upregulation of p53, and induction of apoptosis. The striking correlation between perturbation of nucleolar function, elevated levels of p53 and induction of cell suicide supports the view that the nucleolus is a stress sensor that regulates p53 activity.
In addition, we study rDNA regulation at the epigenetic level. Ribosomal genes exist in two distinct types of chromatin, an “open” one that is permissive to transcription and a “closed” one that is transcriptionally refractive. In exploring the mechanisms that establish and propagate the active and silent state of rRNA genes, we have identified and functionally characterized a chromatin remodeling complex, termed NoRC, that mediates rRNA gene silencing. NoRC recruits DNA methyltransferase and histone deacetylase to the rDNA promoter, thereby triggering heterochromatin formation and silencing of rDNA transcription. Heterochromatin formation appears to be mediated through interaction of TIP5, the large subunit of NoRC, with 100-250 nucleotide RNAs that match the rDNA promoter sequence. Mutations that abrogate RNA binding of TIP5 impair the association of NoRC with rDNA and fail to induce heterochromatin formation. Knockdown of IGS transcripts abolishes the nucleolar localization of NoRC, decreases DNA methylation and enhances rDNA transcription. The results reveal an important contribution of small IGS transcripts in alterations of chromatin structure and epigenetic control of the rDNA locus.
Key primary references
Li, Y., Santoro, R. & Grummt, I. (2005) The chromatin remodeling complex NoRC controls replication timing of ribosomal RNA genes. EMBO J. 24; 120-127.
Mayer, C., Bierhoff, H. & Grummt, I. (2005) The nucleolus as a stress sensor: JNK inactivates the transcription factor TIF-IA and down-regulates rRNA synthesis. Genes & Dev. 19; 933-941.
Yuan, X., Zhou, Y., Casanova, E., Chai, M., Kiss, E., Gröne, H.-J., Schütz, G. & Grummt, I. (2005) Genetic inactivation of the transcription factor TIF-IA leads to nucleolar disruption, cell cycle arrest and p53-mediated apoptosis. Mol. Cell. 19; 77-89.
Zhou, Y. & Grummt, I. (2005) The PHD finger/bromodomain of NoRC interacts with acetylated histone H4K16 and is sufficient for rDNA silencing. Current Biol. 15; 1434-1438.
Key reviews
Grummt, I. & Pikaard, C.S. (2003) Epigenetic control of ribosomal RNA gene transcription. Nature Review Mol. Cell. Biol. 4; 641-649.
Grummt, I. (2003) Life on a planet of its own: regulation of RNA polymerase I transcription in the nucleolus. Genes & Dev. 17; 1691-1702.
Mayer, C. & Grummt, I. (2005) Cellular stress and nucleolar function. Cell Cycle 4; 1036-1038.
Nucleolar assembly and the cell cycle
Nuclei and cell cycle, Institut Jacques Monod,
CNRS, Université Paris VI and Paris VII,
2 place Jussieu, 75251 Paris Cedex 05,
France
Email: dhernand@ccr.jussieu.fr
Biography
Danièle Hernandez-Verdun received a BSc in Biology and a PhD in Development and Reproduction from Faculty of Science, Paris (1974). She spent a post-doctoral training period in the Karolinska Institutet Stockholm, Sweden. She obtained a permanent research position in the CNRS on a mouse developmental project in Paris, and then moved to a new group and has been interested in nuclear, mostly nucleolar organization. Joining the Cell Biology Department of the IJM in 1988, she developed a research team focussed on the nucleolus during the cell cycle. She used successively many cell biology approaches with presently a particular interest for living cell investigations. She was president of the French Society of Electron Microscopy. The research funding is mainly from the CNRS and from a Cancer Research grant (ARC).
Abstract
One of the fundamental features of nuclear organization is that many components of the RNA synthesis and processing machineries are organized in compartments. This implies that the recruitment of dedicated machineries and formation of discrete nuclear domains are crucial events at the beginning of interphase. The nucleolus is the first active domain to be assembled after mitosis. Its functions depend on recruitment of the nucleolar machineries involved in processing of ribosomal RNAs (rRNA) as well as rDNA transcription activation. During mitosis, the RNA polymerase I transcription machinery is assembled on ribosomal genes and repressed by the CDK1-cyclin B pathway. The proteins involved in rRNA processing are distributed around the chromosomes. New results concerning the translocation of processing machinery on transcription sites will be presented. We examined the pathway of the rRNA processing machinery in living cells, and the role of the prenucleolar bodies (PNBs), on the migration of the machinery from the chromosome periphery to sites of rDNA transcription. Protein interactions along the recruitment pathway were investigated by time-lapse analysis of fluorescence resonance energy transfer (FRET). Interestingly interactions between processing proteins occurred first in PNBs before being recruited in nucleoli. The dynamics of these interactions suggest that PNBs are pre-assembly platforms for rRNA processing complexes.
Key primary references
Angelier, N., Tramier, M., Louvet, E., Coppey-Moisan, M., Savino, T.M., De Mey, J.R. & Hernandez-Verdun, D. (2005) Tracking the interactions of rRNA processing proteins during nucleolar assembly in living cells. Mol. Biol. Cell, 16; 2862-71.
Sirri, V., Hernandez-Verdun, D. & Roussel, P. (2002) Cyclin-dependent kinases govern formation and maintenance of the nucleolus. J. Cell Biol.. 156; 969-981.
Savino, T.M., Gébrane-Younès J., De Mey, J., Sibarita, J-B. & Hernandez-Verdun, D. (2001) Nucleolar assembly of the rRNA processing machinery in living cells. J. Cell Biol. 153; 1097-1110.
Verheggen, C., Almouzni, G. & Hernandez-Verdun, D. (2000) The ribosomal RNA processing machinery is recruited to the nucleolar domain before RNA polymerase I during Xenopus laevis. development. J. Cell Biol. 149; 293-305.
Key reviews
Hernandez-Verdun, D., Roussel, P. & Gébranne-Younès J. (2002) Emerging concepts of nucleolar assembly. J. Cell Sci. 115; 2265-2270.
Hernandez-Verdun, D. & Roussel, P. (2003) Regulators of nucleolar functions. Progress in Cell Cycle Research, 5; 301-308. Eds: Meijer L., Jézéquel A. and Roberge M.
Trafficking of viral proteins to the nucleolus
Institute of Cellular and Molecular Biology
Astbury Centre for Structural Molecular Biology
University of Leeds, Leeds, LS2 9JT.
UK
Email: j.a.hiscox@leeds.ac.uk
Biography
Julian A. Hiscox received a BSc in Genetics from University College London (1991) and a PhD in Microbiology from the University of Reading (1994). He conducted post-doctoral training in the Department of Microbiology University of Alabama at Birmingham (USA) and the Division of Molecular Biology at the Institute for Animal Health (Compton, UK). Julian is a Senior Lecturer in Virology at the University of Leeds; prior to joining the faculty at Leeds in 2003 he was a Lecturer in Virology at the University of Reading. The focus of Julian’s research is on the coronavirus nucleocapsid protein, both in terms of trafficking to the nucleolus, affect on the host cell cycle and biophysical characterisation. His research is principally funded by the Biotechnology and Biological Sciences Research Council (BBSRC).
Abstract
The nucleolus is a dynamic sub-nuclear structure involved in ribosome biogenesis, RNA processing, control of cell growth and mediating responses to cell stress and is therefore crucial to the normal operation of the eukaryotic cell. In order to traffic to the nucleolus proteins must usually contain a targeting motif. However, unlike cellular nuclear localisation signals there is no obvious consensus sequence for the nucleolar targeting/retention of proteins to the nucleolus. Studying these signals can be problematic as many nucleolar retention motifs are part of nuclear localisation signals and in some cases nuclear export signals. Utilising the coronavirus nucleocapsid (N) protein we have been investigating nucleolar localisation with the avian and severe acute respiratory (SARS) coronavirus N proteins and the subsequent consequences for the host cell function and virus replication. We have delineated nucleolar and nuclear targeting and also nuclear export signals in the N protein and have mapped an eight amino acid arginine rich motif, which can direct an exogenous protein to the nucleolus. Molecular modelling of the N protein nucleolar retention signal revealed that this motif is available for interaction with cellular factors, which may mediate nucleolar localisation. Our data suggests that not only is the primary sequence of a nucleolar localisation signal important for its function but also the context with regard to the three dimensional structure of the protein.
Key primary references
Chen, H., Gill, A., Dove, B.K., Emmett, S.R., Kemp, F.C., Ritchie, M.A., Dee, M. & Hiscox, J.A. (2005) Mass spectroscopic characterisation of the coronavirus infectious bronchitis virus nucleoprotein and elucidation of the role of phosphorylation in RNA binding using surface plasmon resonance. J. Virol. 79; 1164-1179.
Chen, H., Wurm, T., Britton, P., Brooks, G. & Hiscox, J.A. (2002) Interaction of the coronavirus nucleoprotein with nucleolar antigens and the host cell. J. Virol. 76; 5233-5250.
Wurm, T., Chen, H., Britton, P., Brooks, G. & Hiscox, J.A. (2001) Localisation to the nucleolus is a common feature of coronavirus nucleoproteins and the protein may disrupt host cell division. J. Virol. 75; 9345-9356.
You, J.-H., Dove, B.K., Enjuanes, L., DeDiego, M.L., Alvarez, E., Howell, G., Heinen, P., Zambon, M. & Hiscox, J.A. (2005) Sub-cellular localisation of the severe acute respiratory syndrome coronavirus nucleocapsid protein. J. Gen. Virol. 86; 1-8.
Key reviews
Hiscox, J.A. (2002) Brief review: The nucleolus - a gateway to viral infection? Archives of Virol. 147; 1077-1089.
Hiscox, J.A. (2003) The interaction of animal cytoplasmic RNA viruses with the nucleus to facilitate replication. Virus Research 95; 13-22.
Nucleolus & evolution
Université Libre de Bruxelles
Belgium
Email: denis.lafontaine@ulb.ac.be
Web: www.ulb.ac.be/sciences/rna
Biography
Denis LJ Lafontaine received a BSc and a PhD in Molecular Biology & Genetics from the University of Namur (Belgium). He conducted post-doctoral training in the Gene Expression programme (EMBL, Heidelberg) and The Welcome Trust Center for Cell Biology (University of Edinburgh). Denis is currently a group leader in the Institute of Molecular Biology & Medecine at the University of Brussels. Denis is working on several aspects of RNA metabolism (including, pre-rRNA synthesis, processing & modification, pre-ribosome assembly & trafficking) and organelle morphogenesis. His research is principally funded by the Fonds National de la Recherche Scientifique (FNRS).
Abstract
In eukaryotes, ribosome synthesis largely takes place in a specialized nuclear domain – the nucleolus. It has recently become apparent that this organelle is involved in the biogenesis of most cellular ribonucleoprotein particles (RNPs), as well as in cell-cycle regulation, making it central to gene expression. The field has traditionally acknowledged that each nucleolus is organized in three morphologically distinct compartments. Here, however, we discuss our view that in fact many eukaryotes have bipartite nucleoli. We propose that, during evolution, a third nucleolar compartment emerged at the transition between the anamniotes and the amniotes, following a substantial increase in size of the rDNA intergenic region. We believe that these conclusions have important implications for understanding the structure–function relationships within this key cellular organelle.
Key primary references
Thiry, M. & Lafontaine, D.L.J. (2005) Birth of a nucleolus: the evolution of nucleolar compartments. Trends in Cell Biol. 15; 194-199.
Hoang, T., Peng, W.-T., Vanrobays, E., Krogan, N., Hiley, S., Beyer, A., Osheim, Y., Greenblatt, J., Hughes, T.R., & Lafontaine, D.L.J. (2005) Esf2p, a U3-associated factor required for the initial steps of small ribosomal subunit synthesis. Mol. Cell. Biol. 25; 5523-5534.
Colau, G., Thiry, M., Leduc, V., Bordonné, R., & Lafontaine, D.L.J. (2004) The small nucle(ol)ar RNA cap trimethyltransferase is required for ribosome synthesis and intact nucleolar morphology. Mol. Cell. Biol. 24; 7976-7986.
Key reviews
Lafontaine, D.L.J. (2004). Eukaryotic ribosome synthesis. In Protein synthesis and ribosome structure (ed. K. Nierhaus), 107-143. Wiley-InterScience.
Lafontaine, D.L.J. & Tollervey, D. (2001) The function and synthesis of ribosomes. Nature Reviews Mol. Cell. Biol. 2; 514-20.
Lafontaine, D.L.J. & Tollervey, D. (1998) Birth of the snoRNPs: the evolution of the modification-guide snoRNAs. Trends Biochem. Sci. 23; 383-8.
Angus I Lamond
Nucleolar Dynamics: the ins and outs of the nucleolar proteome
Division of Gene Regulation and Expression,
Wellcome Trust Biocentre
MSI/WTB Complex
University of Dundee
Dundee, DD1 5EH.
UK
Email: angus@lifesci.dundee.ac.uk
Biography
Angus Lamond received a B.Sc. in Molecular Biology from the University of Glasgow (1981) and a PhD in Molecular Biology from the University of Cambridge (1984). He conducted post-doctoral training with Professor Phillip Sharp in the Cancer Center of M.I.T. (USA) and in 1987 moved to the European Molecular Biology Laboratory (EMBL) in Heidelberg as a group leader. Angus moved to his current position at the University of Dundee in 1995, as a Professor of Biochemistry and Wellcome Trust Principal Research Fellow, where he currently heads the Division of Gene Regulation and Expression. Angus’ research is concentrated on studying structure/function relationships and dynamic processes in the cell nucleus, using a combination of quantitative proteomics and time-lapse imaging techniques. The Lamond group is principally funded by the Wellcome Trust.
Abstract
We are studying the functional organization of mammalian cell nuclei with the aim of understanding how specific nuclear compartments are assembled and how individual protein and RNA-protein complexes are targeted to these compartments and are able to move between them (see www.LamondLab.com). We are using a combination of quantitative proteomic methods, involving metabolic labeling of proteins with amino acids containing heavy isotopes 13C and 15N, and quantitative in vivo imaging techniques. Experiments using multiplexed heavy isotope labeling (SILAC) and quantitative mass spectrometry have identified changes in the protein composition of the nucleolus under different metabolic conditions and at specific stages of cell cycle progression. Parallel studies using quantitative digital fluorescence microscopy confirm the dynamic behaviour of nucleolar proteins. A detailed description of the nucleolar proteome and the SILAC data is available in a searchable online database (see http://lamondlab.com/nopdb/). Recently we have used pulse-labeling in combination with SILAC and time-lapse fluorescence imaging to characterize the interactions of ribosomal proteins with nucleoli and their flux in the pathway of ribosomal subunit assembly. This work offers a general approach for characterizing dynamic changes in the composition of either organelles or multi-protein complexes.
Key primary references
Andersen, J.S., Lyon, C.E., Fox, A.H., Leung, A.K.L., Lam, Y.W., Steen, H., Mann, M. & Lamond, A.I. (2002) Directed Proteomic Analysis of the Human Nucleolus. Current Biol. 12; 1-11.
Fox, A.H., Lam, Y.W., Leung, A.K.L., Lyon, C.E., Andersen, J., Mann, M. & Lamond, A.I. (2002) Paraspeckles: A Novel Nuclear Domain. Current Biol. 12;13-25.
Leung, A.K.L. & Lamond, A.I. (2002) In vivo Analysis of NHPX reveals a Novel Nuclear Localisation Pathway involving a Transient Accumulation in Splicing Speckles. J. Cell Biol. 157; 615-629.
Leung, A.K.L., Gerlich, D., Miller, G., Lyon, C., Lam, Y.W., lleres, D., Daigle, N., Zomerdijk, J., Ellenberg, J. & Lamond, A.I. (2004) Quantitative Kinetic Analysis of Nucleolar Breakdown and Reassembly during Mitosis in live Human Cells. J. Cell Biol. 166; 787-800.
Andersen, J.S., Lam, Y.W., Leung, A.K.L., Ong, S-E., Lyon, C.E., Lamond, A.I. & Mann, M. (2005) Nucleolar Proteome Dynamics. Nature, 433; 77-82.
Key reviews
Leung, A.K.L. & Lamond, A.I. (2003) The Dynamics of the Nucleolus. Critical Reviews in Eukaryotic Gene Expression. 13; 49-64.
Lamond, A.I. & Spector, D.L. (2003) Nuclear Speckles: A Model For Nuclear Organelles. Nature Reviews Mol. Cell Biol. 4; 605-612.
Lam, Y.W., Trinkle-Mulcahy, L. & Lamond, A.I. (2005) The nucleolus. J. Cell Sci. 118; 1335-1337.
The role of the nucleolus in viral infection
Department of Cellular and Molecular Medicine
University of Bristol
Bristol
BS8 1TD
UK
Email: d.a.matthews@bristol.ac.uk
Biography
Dr Matthews received a B.Sc. in Microbiology and Virology from the University of Warwick, a M.Sc. in Biological Sciences by Research from the University of Warwick and a PhD in Biochemistry from the University of St Andrews. He initially worked on the molecular epidemiology of respiratory syncytial virus at Warwick University before moving on to adenovirus research at St Andrews for his PhD. His post-doctoral training was originally on adenovirus gene therapy vector development at McMaster University (Canada) and later on adenovirus-host cell interactions at the University of Leeds where he secured a personal fellowship from the Medical Research Council. Dr Matthews is currently a Lecturer in Virology at the University of Bristol where the focus of his research is on adenovirus interactions with the host cell. In particular the work focuses on interactions with components of the nucleolus and with the nuclear import machinery. His research has been primarily funded by the Medical Research Council and the Wellcome Trust.
Abstract
During infection with adenovirus, rRNA processing is disrupted and eventually rRNA synthesis is down regulated. In addition, major nucleolar antigens are redistributed within the infected nucleus. We have been using this viral system to ask where the nucleolar antigens are localised relative to the viral replication proteins and what role they might play during a viral infection. In addition we are interested in which viral proteins are targeted to the nucleolus and for what purpose. So far we have identified three viral proteins that are targeted to the nucleolus and established the molecular basis of this phenomenon. We have also shown that at least one is capable of disrupting the nucleolus. In addition we have been examining the potential for nucleolar antigens to play a role in the replication of viral DNA.
References
Lee, T.W.R., Lawrence, F.J., Dauksaite, V., Blair, G.E., Akusjärvi, G. & Matthews, D.A. (2004) Precursor of human adenovirus core polypeptide Mu targets the nucleolus and modulates the expression of E2 proteins. J. Gen. Virol. 85;185-196.
Lee T.W.R., Blair, G.E. & Matthews, D.A. (2003) Adenovirus core protein VII contains distinct sequences that mediate targeting to the nucleus, nucleolus, and colocalisation with human chromosomes.J. Gen. Virol. 84; 3423-3428.
Matthews, D.A. (2001) Adenovirus protein V induces redistribution of nucleolin and B23 from nucleolus to cytoplasm. J. Virol. 75; 1031-8.
Matthews, D.A. & Russell, W.C. (1998) Adenovirus core protein V is delivered by the invading virus to the nucleus of the infected cell and later in infection is associated with nucleoli. J. Gen. Virol. 79; 1671-5.
Jo Milner
The nucleolus, stress and cancer
YCR P53 Research Group
Department of Biology
University of York
YORK
UK
Email: ajm24@york.ac.uk
Biography
Jo Milner was a State Scholar at the University of London where she obtained a BSc in Zoology. She gained her PhD from the University of Cambridge and went on to study the correlation between mitochondrial ultra-structure and the biochemical activity of steroid biosynthetic enzymes located within the inner mitochondrial membrane. She was a Research Fellow at Harvard Medical School, USA, and subsequently a Beit Memorial Research Fellow at the University of Cambridge. She continued independent research as a Senior Research Associate in Cambridge, studying commitment from quiescence into the cell cycle, and established functional links between the conformational structure and flexibility of the tumour suppressor protein p53. In 1991 she was appointed Yorkshire Cancer Research Professor of Cell Biology in the Department of Biology at the University of York. Her research interests centre upon (i) protein structure, modifications and functions of the tumour suppressor p53, and (ii) the regulation of apoptosis in cancer versus non-cancer cells.
Abstract
Sirtuins are a new class of histone deacetylases implicated in diverse pathogenetic roles, including cancer. They impact upon the architecture of the genome and affect gene expression by their capacity to deacetylate chromatin-interacting proteins. SIRT1 is functionally linked with the p53 tumour suppressor, which it inactivates via deacetylation. SIRT1 is thus believed to attenuate a p53 response following exposure of cells to genotoxic stress. Single cell analysis has demonstrated that activation of p53 in response to UV-mediated damage requires involvement of the nucleolar chromatin, identifying the nucleolus as a stress sensor in mammalian cells. SIRT1 also has the capacity to function as a cancer-specific survival factor under basal non-stress conditions. Current studies now aim to characterise the impact of diverse stress conditions upon the nucleolus and thus to explore the repertoire of stress responses that may be linked with the nucleolus. Comparison of cancer and non-cancerous cells may reveal cancer-related differences in the behaviour of the nucleolus under defined conditions of stress.
Key References
Ford, J., Jiang, M.J. and Milner, J. (2005) Cancer-specific functions of SIRT1 enables human epithelial cancer cell growth and survival. Cancer Research,65; 10457-10463.
Jiang, M. and Milner, J. (2003) Bcl-2 constitutively suppresses p53-dependent apoptosis in colorectal cancer cells. Genes Dev.17; 832-837.
Jiang, M. and Milner, J. (2002) Selective silencing of viral gene expression in HPV-positive human cervical carcinoma cells treated with siRNA, a primer of RNA interference. Oncogene21; 6041-6048.
Rubbi, C.P. and Milner, J. (2003) Disruption of the nucleolus mediates stabilization of p53 in response to DNA damage and other stresses. EMBO J.22; 6068-6077.
Key Review
Milner, J. (2004) Death without stress - the stiletto approach against disease. The Biochemist26; 16-18.