The overarching goal of this proposal is to the explore the role of stochastic gene expression during development and cell differentiation using the nematode worm Caenorhabditis elegans and mammalian cell lines as experimental model systems. In the first program of this proposal I describe a single molecule mRNA detection technique that allows accurate measurements of stochastic expression of endogenous genes in developmental and differentiation pathways. We will use this technique to determine expression fluctuations in the C. elegans endodermal specification pathway. Next, we will use this technology to better understand the phenomenon of incomplete penetrance. Many C. elegans mutants exhibit the phenomenon of incomplete penetrance, where only a certain fraction of genetically identical mutants exhibit the defective mutant phenotype, while the remaining organisms are often indistinguishable from the wild-type. We will test the hypothesis that these mutants allow stochastic events in the gene expression program controlling development to influence the organism's ultimate developmental outcome. Finally we will use the mRNA counting method to determine the stochastic component of mammalian cell fate decisions using the glucocorticoid pathway and hemapoietic progenitor cells as experimental model systems. The second program focuses on elucidating the roles of microRNAs in controlling gene expression fluctuations. During the last decade it has become clear that microRNAs play crucial roles in diverse biological processes including development and differentiation. Program 2 will explore the potential roles for microRNAs controlling gene expression fluctuations. First, we will experimentally determine the relationship between the concentration of a microRNA and its target in a single cell. Second, we will test the hypothesis that in order to dampen translational amplification of mRNA fluctuations, miRNAs may be employed to selectively lower the translation rate for genes that can fluctuate significantly. Finally, we will explore if microRNA-mediated feedback networks can lower fluctuations in target gene expression.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
NIH Director’s Pioneer Award (NDPA) (DP1)
Project #
8DP1CA174420-05
Application #
8306912
Study Section
Special Emphasis Panel (ZGM1-NDPA-B (P2))
Program Officer
Mietz, Judy
Project Start
2008-09-30
Project End
2014-01-31
Budget Start
2012-08-01
Budget End
2014-01-31
Support Year
5
Fiscal Year
2012
Total Cost
$831,600
Indirect Cost
$336,600
Name
Massachusetts Institute of Technology
Department
Physics
Type
Schools of Arts and Sciences
DUNS #
001425594
City
Cambridge
State
MA
Country
United States
Zip Code
02139
Semrau, Stefan; Crosetto, Nicola; Bienko, Magda et al. (2014) FuseFISH: robust detection of transcribed gene fusions in single cells. Cell Rep 6:18-23
Mentink, Remco A; Middelkoop, Teije C; Rella, Lorenzo et al. (2014) Cell intrinsic modulation of Wnt signaling controls neuroblast migration in C. elegans. Dev Cell 31:188-201
Slavov, Nikolai; Budnik, Bogdan A; Schwab, David et al. (2014) Constant growth rate can be supported by decreasing energy flux and increasing aerobic glycolysis. Cell Rep 7:705-14
Klemm, Sandy; Semrau, Stefan; Wiebrands, Kay et al. (2014) Transcriptional profiling of cells sorted by RNA abundance. Nat Methods 11:549-51
Kim, Dong hyun; Grun, Dominic; van Oudenaarden, Alexander (2013) Dampening of expression oscillations by synchronous regulation of a microRNA and its target. Nat Genet 45:1337-44
Fang, Miaoqing; Xie, Huangming; Dougan, Stephanie K et al. (2013) Stochastic cytokine expression induces mixed T helper cell States. PLoS Biol 11:e1001618
Teytelman, Leonid; Thurtle, Deborah M; Rine, Jasper et al. (2013) Highly expressed loci are vulnerable to misleading ChIP localization of multiple unrelated proteins. Proc Natl Acad Sci U S A 110:18602-7
Ji, Ni; Middelkoop, Teije C; Mentink, Remco A et al. (2013) Feedback control of gene expression variability in the Caenorhabditis elegans Wnt pathway. Cell 155:869-80
Hansen, Clinton H; van Oudenaarden, Alexander (2013) Allele-specific detection of single mRNA molecules in situ. Nat Methods 10:869-71
Munsky, Brian; Neuert, Gregor; van Oudenaarden, Alexander (2012) Using gene expression noise to understand gene regulation. Science 336:183-7

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