The goal of this study is to broaden our understanding of a gene regulatory network that plays key roles during embryogenesis. Recent studies argue strongly that the transcription factors Sox2 and Oct-3/4 each exhibit important hallmarks of master regulators that orchestrate mammalian embryogenesis and the self-renewal of embryonic stem (ES) cells. Sox2 and Oct-3/4 have been shown to work together cooperatively to coordinate the transcription of at least 6 genes (FGF-4, UTF1, Fbx15, Nanog, Sox2 and Oct-3/4) expressed during embryogenesis and by ES cells. These genes are referred to as Sox2:Oct-3/4 target genes. The work proposed here is based on 3 novel findings and hypotheses. First, we recently identified 19 additional genes that are likely to be part of the Sox2:Oct-3/4 gene regulatory network. Given that 4, and possibly 5, of the 6 known Sox2:Oct-3/4 target genes are essential for embryogenesis it is our central hypothesis that the majority of Sox2:Oct-3/4 target genes play critical roles during embryogenesis, and that many are required for the self-renewal of ES cells. Second, the expression of the 6 known Sox2:Oct-3/4 target genes was shown to be very heavily dependent on closely spaced DNA binding sites for Sox2 and Oct-3/4 in the enhancer of each gene. These and other studies led us to hypothesize that the Sox2:Oct-3/4 gene regulatory network is controlled coordinately by a shared set of co-activators, at least some of which are themselves regulated during differentiation and, thus, are key players in cell fate determination. Third, although Sox2, in combination with Oct-3/4, is known to stimulate the expression of Sox2:Oct-3/4 target genes in stem cells, we determined that at least 5 of the 6 Sox2:Oct-3/4 target genes, including the Sox2 gene itself, are inhibited when Sox2 is overexpressed. These findings led to our hypothesis that Sox2:Oct-3/4 target genes are regulated carefully by a negative, as well as a positive, feedback loop to ensure that these genes are expressed at levels required for embryogenesis.
Three Specific Aims are proposed to test these hypotheses. 1) Examine a newly identified set of putative Sox2:Oct-3/4 target genes for their effects on the self-renewal of ES cells and mammalian development. 2) Examine the roles of specific co-activators in the expression of Sox2:Oct-3/4 target genes. 3) Determine the molecular mechanisms by which Sox2 overexpression inhibits the expression of Sox2:Oct-3/4 target genes. Together, these studies will provide a mechanistic understanding of a critical gene regulatory network that orchestrates embryogenesis and controls the self-renewal of ES cells. As such, these studies will provide novel insights and have significant impact on developmental biology, regenerative medicine and cancer biology.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM080751-23
Application #
7759527
Study Section
Development - 2 Study Section (DEV2)
Program Officer
Haynes, Susan R
Project Start
1985-08-01
Project End
2013-01-31
Budget Start
2010-02-01
Budget End
2013-01-31
Support Year
23
Fiscal Year
2010
Total Cost
$265,592
Indirect Cost
Name
University of Nebraska Medical Center
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
168559177
City
Omaha
State
NE
Country
United States
Zip Code
68198
Rizzino, Angie (2013) Concise review: The Sox2-Oct4 connection: critical players in a much larger interdependent network integrated at multiple levels. Stem Cells 31:1033-9
Ormsbee Golden, Briana D; Wuebben, Erin L; Rizzino, Angie (2013) Sox2 expression is regulated by a negative feedback loop in embryonic stem cells that involves AKT signaling and FoxO1. PLoS One 8:e76345
Wuebben, Erin L; Mallanna, Sunil K; Cox, Jesse L et al. (2012) Musashi2 is required for the self-renewal and pluripotency of embryonic stem cells. PLoS One 7:e34827
Mallanna, Sunil K; Rizzino, Angie (2012) Systems biology provides new insights into the molecular mechanisms that control the fate of embryonic stem cells. J Cell Physiol 227:27-34
Cox, Jesse L; Mallanna, Sunil K; Ormsbee, Briana D et al. (2011) Banf1 is required to maintain the self-renewal of both mouse and human embryonic stem cells. J Cell Sci 124:2654-65
Chakravarthy, Harini; Ormsbee, Briana D; Mallanna, Sunil K et al. (2011) Rapid activation of the bivalent gene Sox21 requires displacement of multiple layers of gene-silencing machinery. FASEB J 25:206-18
Ji, Ming; Rao, Enyu; Ramachandrareddy, Himabindu et al. (2011) The miR-17-92 microRNA cluster is regulated by multiple mechanisms in B-cell malignancies. Am J Pathol 179:1645-56
Mallanna, Sunil K; Rizzino, Angie (2010) Emerging roles of microRNAs in the control of embryonic stem cells and the generation of induced pluripotent stem cells. Dev Biol 344:16-25
Mallanna, Sunil K; Ormsbee, Briana D; Iacovino, Michelina et al. (2010) Proteomic analysis of Sox2-associated proteins during early stages of mouse embryonic stem cell differentiation identifies Sox21 as a novel regulator of stem cell fate. Stem Cells 28:1715-27
Cox, Jesse L; Rizzino, Angie (2010) Induced pluripotent stem cells: what lies beyond the paradigm shift. Exp Biol Med (Maywood) 235:148-58

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