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David Stillman

Professor of Pathology

David Stillman

B.A. University of California, Berkeley

Ph.D. University of California, San Diego

Research

References

david.stillman@path.utah.edu

David Stillman's Lab Page

David Stillman's PubMed Literature Search

Molecular Biology Program

Gene Regulation and Chromatin Structure

Research

We want to understand eukaryotic transcriptional regulation at the molecular level. Problems in gene regulation underlie many human diseases. We study gene regulation in yeast because of the powerful genetic and molecular tools that are available. Importantly, the transcription regulatory machinery is conserved between yeast and vertebrates, and insights gained from studies in yeast are generally universal.

Changes in chromatin structure leading to gene activation. We have studied activation of the yeast HO gene, which is transiently expressed during the cell cycle in only one of the two cells following mitotic division. Chromatin immunoprecipitation (ChIP) experiments show that transcriptional coactivator complexes get recruited to one region of the HO promoter early in the cell cycle, and then migrate to a different promoter region. Chromatin disassembly occurs in waves both along the length of the promoter and during the cell cycle, and three different chromatin factors are required for disassembly of nucleosomes, each at different regions of the HO promoter. The SBF DNA-binding factor binds to the promoters of HO and the G1 cyclin genes, and recruits three distinct factors: the Rpd3(L) histone deacetylase that inhibits gene activation, the FACT chromatin reorganizing complex that stimulates transcription, and the cyclin dependent kinase (CDK) that activates these genes as cells pass the G1/S transition by expelling the Rpd3(L) inhibitor from the promoter. Current experiments are to understand how these chromatin changes are regulated and how they control gene activity.

Memory at the HO promoter. We have identified several types of “memory” at the HO promoter. When deprived of nutrients and cells arrest in G1 of the cell cycle, the HO promoter “remembers” earlier events at the promoter by the stable binding of the SBF and Mediator factors; this allows the promoter to be activated when nutrients are added and cells reenter the cell cycle. There is a distinct form of G1 cell cycle arrest that occurs in response to mating pheromone, which activates the Ste12 transcription factor. From far upstream at the HO promoter, Ste12 activates a long noncoding RNA, and the polymerase making this ncRNA appears to disrupt binding of SBF and Mediator. As a result, memory is eliminated and HO is not expressed in the first cell cycle following cell cycle arrest by pheromone. We have evidence for a second type of memory at HO, a form of transcriptional repression that persists longer than a cell cycle. Experiments are in progress to define this repression molecularly.

Promoter specificity of transcriptional factors. Specific gene expression is controlled by transcription factors binding to elements present in promoters and enhancers. Yeast has two transcription factors, Swi5 and Ace2, that show similar patterns of cell cycle regulation, that have nearly identical zinc finger DNA-binding domains, and recognize the same DNA sequences in vitro. Despite these similarities, Swi5 and Ace2 activate transcription of different genes in vivo, in addition to a common set of target genes. HO is a "Swi5-only" gene where Swi5 binds and activates in vivo, but Ace2 cannot bind in vivo. Since Ace2 binds to these sites in vitro, promoter specificity is determined by mechanisms that control factor binding. Experiments are in progress to determine how chromatin structure can prevent one factor from binding while allowing another protein, with the same DNA-binding domain, to bind. Both Swi5 and Ace2 bind to "Ace2-only" promoters in vivo, but only Ace2 is able to actually activate transcription of these genes; thus, Swi5 binds to these promoters but fails to activate. We have identified other proteins bound nearby at these promoters that function as selective “anti-activators,” blocking Swi5 from activating transcription while allowing Ace2 to activate. Current experiments are studying the mechanisms of these selective anti-activators. These studies are quite relevant to mammalian gene regulation, because in many cases it has been shown that multiple transcription factors recognize the same sequence, but that simple DNA-binding at a promoter is not sufficient for gene activation in vivo.

FACT in transcriptional elongation. We are studying the FACT chromatin reorganizing complex in collaboration with Tim Formosa’s laboratory. In addition to the functions at promoters (like HO) and DNA replication, FACT promotes transcriptional elongation. Current studies investigate how FACT functions to properly reassemble nucleosomes following passage of an elongating RNA polymerase.

Stillman Figure

References

  1. Chiaro, T., R. Soto, , W.Z. Stephens, J.L. Kubinak, C. Petersen, L.Gogokhia, R. Bell, J.C. Delgado, J. Cox, W. Voth, J. Brown, D. Stillman, Ryan O’Connell, Anne Tebo, and June L. Round. (2017) A member of the mycobiota modulates host metabolism to increase intestinal permeability and disease. Science Translational Medicine 9 (380). pii: eaaf9044. 
  2. Yu, Y., R.M. Yarrington, E.B. Chuong, N.C. Elde, and D.J. Stillman. (2016) Disruption of Promoter Memory by Synthesis of a Long Noncoding RNA. Proc. Natl. Acad. Sci. USA 113:9575-80.
  3. Yarrington, R.M., J.S. Rudd, and D.J. Stillman. (2015) Spatiotemporal cascade of transcription factor binding required for promoter activation. Mol. Cell Biol. 35:688-98.
  4. Parnell, E.J., Y. Yu, R. Lucena, Y. Yoon, L. Bai, D.R. Kellogg, and D.J. Stillman. (2014) The Rts1 Regulatory Subunit of PP2A Phosphatase Controls Expression of the HO Endonuclease via Localization of the Ace2 Transcription Factor Localization. J. Biol. Chem. 289:35431-25437.
  5. Zapata, J., N. Dephoure, T. Macdonough, Y. Yu, E.J. Parnell, M. Mooring, S.P. Gygi, D.J. Stillman, and D.R. Kellogg. (2014) PP2ARts1 is a master regulator of pathways that control cell size. J. Cell Biol. 204:359–376.
  6. Voth, W.P., S. Takahata, J.L. Nishikawa, B.M. Metcalfe, A.M. Näär, and D.J. Stillman. (2014) A Role for FACT in Repopulation of Nucleosomes at Inducible Genes. PLoS ONE. 9, e84092.
  7. Stillman, D.J. (2013) Dancing the cell cycle two-step: regulation of yeast G1-cell-cycle genes by chromatin structure. Trends Biochem. Sci. 38:467-475.
  8. Zhang, Q., Y. Yoon, Y. Yu, E.J. Parnell, J.A. Garay, M.M. Mwangi, F.R. Cross, D.J. Stillman, and L. Bai. (2013) Stochastic expression and epigenetic memory at the yeast HO promoter. Proc. Natl. Acad. Sci. USA. 110:14012-14017.
  9. Takahata, S., Y. Yu, and D.J. Stillman. (2011) Repressive Chromatin Affects Factor Binding at the Yeast HO (Homothallic Switching) Promoter. J. Biol. Chem. 286:34809-34819.
  10. McCullough, L., R. Rawlins, A.E. Olsen, H. Xin, D.J. Stillman, and T. Formosa. (2011) Insight into the Mechanism of Nucleosome Reorganization from Histone Mutants that Suppress Defects in the FACT Histone Chaperone. Genetics. 188:835-846.
  11. Stillman, D.J. (2010) Nhp6: A Small but Powerful Effector of Chromatin Structure in Saccharomyces cerevisiae. Biochim. Biophys. Acta. 1799:175-180.
  12. Takahata, S., Y. Yu, and D.J. Stillman. (2009) The E2F functional analog SBF recruits the Rpd3(L) HDAC, via Whi5 and Stb1, and the FACT chromatin reorganizer, to yeast G1 cyclin promoters. EMBO J. 28:3378-3389
  13. Pondugula, S., D.W. Neef, W.P. Voth, A. Dhasarathy, M.M. Reynolds, S. Takahata, D.J. Stillman, and M.P. Kladde. (2009) Coupling Phosphate Homeostasis to Cell Cycle-Specific Transcription: Mitotic Activation of S. cerevisiae PHO5 by Mcm1 and Forkhead Proteins. Mol. Cell Biol. 29:4891-4905.
  14. Xin, H., S. Takahata, M. Blanksma, L. McCullough, D.J. Stillman, and T. Formosa. (2009) yFACT Induces Global Accessibility of Nucleosomal DNA Without H2A-H2B Displacement. Mol. Cell. 35:365-376.
  15. Takahata, S., Y. Yu, and D.J. Stillman. (2009) FACT and Asf1 regulate nucleosome dynamics and movement of coactivators at the HO promoter. Mol. Cell. 34:405-415.
  16. Biswas, D., S. Takahata, and D.J. Stillman. (2008) Different Genetic Functions for the Rpd3(L) and Rpd3(S) Complexes Suggest Competition Between NuA4 and Rpd3(S) Mol. Cell Biol. 28:4445-48.
  17. Sbia, M., E.J. Parnell, Y. Yu, A.E. Olsen, K.L. Kretschmann, W.P. Voth, and D.J. Stillman. (2008) Regulation of the Yeast Ace2 Transcription Factor During the Cell Cycle. J. Biol. Chem. 283:11135-45.
  18. Biswas, D., S. Takahata, H. Xin, R. Dutta-Biswas, Y. Yu, T. Formosa, and D.J. Stillman. (2008) A role for Chd1 and Set2 in negatively regulating DNA replication in Saccharomyces cerevisiae. Genetics. 178:649-59.
  19. Voth, W.P., Y. Yu, S. Takahata, K.L. Kretschmann, J.D. Lieb, R.L. Parker, B. Milash, and D.J. Stillman. (2007) Forkhead proteins control the outcome of transcription factor binding by antiactivation. EMBO J. 26: 4324-34.
  20. Biswas, D., R. Dutta-Biswas, and D.J. Stillman. (2007) Chd1 and yFACT act in opposition in regulating transcription. Mol. Cell Biol. 27: 6279-87.
  21. Biswas, D., R. Dutta-Biswas, D. Mitra, Y. Shibata, B.D. Strahl, T. Formosa, and D.J. Stillman. (2006) Opposing roles for Set2 and yFACT in regulating TBP binding at promoters. EMBO J. 25:4479-4489.
  22. Mitra, D., E.J. Parnell, J.W. Landon, Y. Yu, and D.J. Stillman. (2006) Swi/Snf binding requires histone acetylation and stimulates TBP recruitment to the HO promoter. Mol. Cell Biol. 26:4095-4110.
  23. Biswas, D., Y. Yu, M. Prall, T. Formosa, and D.J. Stillman. (2005) The Yeast FACT Complex Has a Role in Transcriptional Initiation. Mol. Cell Biol. 25: 5812-5822.
  24. Biswas, D., Y. Yu, P. Eriksson, A.N. Imbalzano, and D.J. Stillman. (2004) A role for Nhp6, Gcn5, and the Swi/Snf complex in stimulating formation of the TBP-TFIIA-DNA complex. Mol. Cell Biol. 24: 8312-8321.
  25. Eriksson, P., D. Biswas, Y. Yu, J.M. Stewart, and D.J. Stillman. (2004) TATA-binding protein (TBP) mutants that are lethal in the absence of the Nhp6 HMG Protein. Mol. Cell Biol. 24: 6419-6429.
  26. Laabs, T.L., D.D. Markwardt, M. G. Slattery, L. L. Newcomb,  D.J. Stillman, and W. Heideman. (2003) ACE2 is required for daughter cell-specific G1 delay in Saccharomyces cerevisiae. Proc. Natl. Acad. Sci. USA. 100:10275-10280.
  27. Yu, Y., P. Eriksson, L.T. Bhoite, and D.J. Stillman. (2003) Regulation of TBP Binding by the SAGA Complex and the Nhp6 HMG Protein. Mol. Cell Biol. 23:1910-1921.
  28. Bhoite, L.T., Y. Yu, and D.J. Stillman. (2001) The Swi5 activator recruits the Mediator complex to the HO promoter without RNA polymerase II. Genes Dev. 15: 2457-2469.
  29. Formosa, T., P. Eriksson, J. Wittmeyer, J. Ginn, Y. Yu, and D.J. Stillman. (2001) Spt16-Pob3 and the HMG protein Nhp6 combine to form the nucleosome-binding factor SPN. EMBO J. 20: 3506-3517.

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Last Updated: 8/15/17