Our lab is dedicated to gaining mechanistic insight into crucial molecular processes in the life of the eukaryotic cell. Our two main research topics are the role of cytoskeletal dynamics in cell division, and the molecular mechanisms governing the flow of genetic information. The unifying principle in our work is the emphasis on studying macromolecular assemblies as whole units of molecular function via visualization of their functional states and regulatory interactions. We use electron microscopy (EM) and image analysis, biochemical and biophysical assays, towards a molecular understanding of our systems of interest.
Cytoskeleton and Cell Division
In our microtubule cytoskeleton studies we are interested in defining the conformational landscape of tubulin as defined by its nucleotide and assembly states, in order to obtain detail mechanistic understanding of the process of microtubule dynamic instability. Furthermore, to gain insight into how MT dynamics are regulated and utilized in the cellular context, we are studying the interactions of microtubule ends with cellular factors involved in chromosome capture, alignment and segregation. Our studies aim to provide fine details of the highly regulated interactions between microtubules and kinetochore complexes and other microtubule-interacting proteins essential for mitosis progression.
We have expanded our cytoskeletal studies to include the molecular understanding of septin assembly and function in cytokinesis. We have characterized the molecular architecture of septin assembly units, their polymerization into different assembly forms, and the relevance of their interaction with phospho-inositides. More recently we have moved into the realm of in vivo studies of the cytokinetic process in yeast. We are deepening our understanding of all of these aspects of septin structure and function and extending our studies to the characterization of septin interaction with cellular partners.
Find out more about our cytoskeleton studies.
Gene Regulation and the Central DogmaOur studies of nucleic acid transactions have been most powerful at defining the overall architecture of a large number of macromolecular assemblies involved in the critical initiation steps in DNA replication, RNA transcription and translation, for which no crystallographic information yet exist (e.g. ORC, TFIID, eIF3), as well as putting the crystal structures of essential elements in RNA processing and protein degradation in the context of fully functional complexes (e.g. RLC, proteosome). Our aim is to gain mechanistic insight that goes beyond overall architecture, by pushing resolution, describing conformational landscapes, and relating structural states to function via the analysis of interactions with ligands and regulatory factors.
Regulated gene transcription in eukaryotes requires the assembly of a complex molecular machinery that includes general factors, activators, cofactor complexes and chromatin modifying and remodeling factors. We have recently described the structural dynamics of human TFIID and its relevance for recognition of core promoter DNA. We have also used an in vitro reconstituted system to study the stepwise assembly of human TBP, TFIIA, TFIIB, Pol II, TFIIF, TFIIE and TFIIH onto promoter DNA using cryo-electron microscopy. Our structural analyses provide pseudo-atomic models at various stages of transcription initiation that illuminate critical molecular interactions.
He, Y., Fang, J., Taatjes, D.J., and Nogales, E. (2013) Structural visualization of key steps in human transcription initiation. Nature 495, 481-486.
Cianfrocco, M.A., Kassevitis, G.A., Grob, P, Fang, J., Juven-Gershon, T., Kadonaga, J.T. and Nogales, E. (2013) Human TFIID binds core promoter DNA in a reorganized structural state. Cell 152, 120-131.
Ciferri, C., Lander, G.C., Maiolica, A., Herzog, F., Aebersold, R. and Nogales, E. (2012) Structure of the polycomb represive complex 2 and implications for gene silencing. eLIFE, e00005.
Lander, G.C., Estrin, E., Matyskiela, M.E., Bashore, C., Nogales, E. and Martin, A. (2012) Complete subunit architecture of the proteosome regulatory particle. Nature 482,186-191.
Garcia, G.III, Bertin, A., Li, Z., McMurray, M., Thorner, J. and Nogales, E. (2011) Subunit-dependent modulation of septin assembly: budding yeast septin Shs1 promotes ring and gauze formation. JCB 195, 993-1004.
Wiedenheft, B., Lander, G.C., Zhou, K., Jore, M.M., Brouns, S.J.J., van der Oost, J., Doudna, J.A., and Nogales, E. (2011) Structrues of the RNA-guided surveillance complex from a bacterial immune system. Nature 477, 486-489.
Alushin, G., Ramey, V.H., Pasqualato, S., Ball, D., Grigorieff, N., Musacchio, A. and Nogales, E. (2010) The Ndc80 kinetochore complex forms oligomeric arrays along microtubules. Nature 467, 805-810.
Bertin, A., McMurray, M.A., Grob, P., Park, S-S., Garcia, G. III, Patanwala, I., Ng, H-L., Alber, T.C., Thorner, J. and Nogales, E. (2008) Saccharomyces cerevisiae septins: Supramolecular organization of hetero-oligomers and the mechanism of filament assembly. PNAS 105, 8274-8279.
Westermann, S., Wang, H-W., Avila-Sakar, A., Drubin, D.G., Nogales, E. and Barnes, G. (2006) The Dam1 kinetochore ring complex moves processively on depolymerizing microtubule ends. Nature, 440, 565-569.
Siridechadilok, B., Fraser, C.S., Hall, R.J., Doudna, J.A. and Nogales, E. (2005) Structural roles for human translation factor eIF3 in the initiation of protein synthesis. Science, 310, 1513-1515.
Wang, H-W. and Nogales, E. (2005) The nucleotide-dependent bending flexibility of tubulin regulates microtubule assembly. Nature, 435, 911-915.
For a full publication listing please visit http://cryoem.berkeley.edu/pubs.shtml.