The goal of this study is to understand the underlying mechanisms by which a uniform population of early mesoderm cells gives rise to a variety of different tissues Our findings indicate that Twist, a basic helix-loop-helix (bHLH) transcription factor, plays a central role in specification of the mesoderm, its subdivision into different tissues, patterning of the somatic muscles, and differentiation of the adult muscle progenitors. We will focus on how Twist is able to execute these different roles. Based on our findings that, in vivo and in vitro, Twist can form homodimers and heterodimers with the bHLH protein, Daughterless (Da), we predict that Twist activity is tightly regulated by interactions with other HLH proteins in the mesoderm at different points in development. We have defined four HLH candidate proteins that may interact with Twist. Based on expression and loss of function data, we propose that interactions between Twist, Da, and Extramachrochaete are critical for early mesodermal events, whereas interactions between Twist, L'Scute,Da and Hairy are decisive for later events.To test this hypothesis, we will use genetic techniques to manipulate the dosage of Twist and each of these potential partners. If these proteins interact, we will see defects in mesodermal development. We will then examine the relationship between Twist and the candidate HLH protein, using a combination of in vitro [e.g., gel shift] and in vivo [e.g., overexpresssion with tethered dimers] techniques to assess if the proteins interact directly to activate or repress target genes. Our preliminary findings indicate that Twist homodimers activate the myogenic pathway, whereas Twist/Da heterodimers repress the myogenic fate. We will use both in vivo and in vitro techniques to map the domains of Twist required for activation. We will test whether these domains transform Da and vertebrate Twist, Wingless, Decapentaplegic, Notch and Ras signaling pathways affect mesoderm development, at least in part, by altering ratios of Twist homodimers and heterodimers. We will test in vivo, if overexpression of particular dimers in embryos mutant for a signaling pathway rescues the mesodermal defects. Since interactions with the known HLH proteins and Twist will not completely explain the scope of Twist's functions, we will also screen for unknown HLH proteins and co-regulators that interact with Twist on Twist-dependent enhancers. With the strong conservation of genes and mechanisms between drosophila and other species, our data will help understand similar decisions that occur in muscle development in all species. Indeed, mutations in homologues of Twist, Wg, and N in mouse and human lead to developmental abnormalities and our work will have a direct impact on understanding these defects and diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM056989-02
Application #
6138629
Study Section
Genetics Study Section (GEN)
Program Officer
Greenberg, Judith H
Project Start
1999-01-01
Project End
2003-12-31
Budget Start
2000-01-01
Budget End
2000-12-31
Support Year
2
Fiscal Year
2000
Total Cost
$256,704
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
Bothe, Ingo; Baylies, Mary K (2016) Drosophila myogenesis. Curr Biol 26:R786-91
Dobi, Krista C; Schulman, Victoria K; Baylies, Mary K (2015) Specification of the somatic musculature in Drosophila. Wiley Interdiscip Rev Dev Biol 4:357-75
Kumar, Ram P; Dobi, Krista C; Baylies, Mary K et al. (2015) Muscle cell fate choice requires the T-box transcription factor midline in Drosophila. Genetics 199:777-91
Dobi, Krista C; Halfon, Marc S; Baylies, Mary K (2014) Whole-genome analysis of muscle founder cells implicates the chromatin regulator Sin3A in muscle identity. Cell Rep 8:858-70
Wong, Ming-Ching; Dobi, Krista C; Baylies, Mary K (2014) Discrete levels of Twist activity are required to direct distinct cell functions during gastrulation and somatic myogenesis. PLoS One 9:e99553
Yu, Shannon F; Baylies, Mary K (2013) Cell biology: Death brings new life to muscle. Nature 497:196-7
Nowak, Scott J; Baylies, Mary K (2012) Akirin: a context-dependent link between transcription and chromatin remodeling. Bioarchitecture 2:209-13
Metzger, Thomas; Gache, Vincent; Xu, Mu et al. (2012) MAP and kinesin-dependent nuclear positioning is required for skeletal muscle function. Nature 484:120-4
Nowak, Scott J; Aihara, Hitoshi; Gonzalez, Katie et al. (2012) Akirin links twist-regulated transcription with the Brahma chromatin remodeling complex during embryogenesis. PLoS Genet 8:e1002547
Dobi, Krista C; Metzger, Thomas; Baylies, Mary K (2011) Characterization of early steps in muscle morphogenesis in a Drosophila primary culture system. Fly (Austin) 5:68-75

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