The aim of this research is to understand how stable states of gene expression are maintained during development. Molecular, genetic, and biochemical methods will be used to study proteins that control transcription of homeotic genes in Drosophila melanogaster. The homeotic genes determine the identities of segments in the fly. Expression of homeotic genes, in restricted patterns along the anterior-posterior axis, is needed throughout embryonic, larval and pupal development. Segmentation gene products control the initial Polycomb group (PcG) products are transcriptional repressors that then maintain restricted homeotic patterns throughout the rest of development. Current models suggest that PcG products recognize the initial off states of homeotic genes established in early embryos and they maintain repression by assembling stable protein- DNA complexes in the local chromatin. This research will investigate the individual roles of PcG repressor proteins. The work will focus on two of the PcG products, extra sex combs (esc) protein and Sex comb on midleg (Scm) protein, which are likely to play different roles in repression. esc may play a short-term role in assembling or guiding other objectives of this work are to define the functional domains in esc and Scm and to determine if and how they interact with other PcG proteins. Site-specific mutagenesis, coupled with germline transformation and transient rescue assays, will test the role of WD40 repeats in esc protein, which are likely used for protein-protein contact. DNA sequence analysis of mutant alleles and site-directed mutagenesis will be used to test the functional domains in Scm, including several zinc fingers. Isolation of esc and Scm homologs in other insect species will be used to identify functional domains. These studies will be extended to test for mammalian homologs of these PcG genes. Gel shift assays will test if Scm protein binds to DNA. In vitro binding assays, yeast wi-hybrid tests, and co-immune precipitations will check for physical interactions between esc, Scm and other PcG proteins. These interactions will also be tested in vivo using PcG proteins fused to a DNA-binding domain. esc and Scm fusion proteins will be tethered to specific sites in the genome and chromosome immunostaining will test for co-association of other PcG proteins. This work will address how cell fate decision are maintained during fly development. In addition, the mammalian Hox genes control anterior- posterior patterning in ways that resemble the fly homeotic genes. There are several mouse genes that are related to fly PcG genes, including at least one that is a functional homolog. Thus, an understanding of PcG repression in files should provide clues about similar controls in higher organisms. Since the mouse PcG homolog is an oncogene, this work may also provide insight into the basis of uncontrolled cell proliferation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM049850-03
Application #
2749946
Study Section
Genetics Study Section (GEN)
Project Start
1996-08-01
Project End
2000-07-31
Budget Start
1998-08-01
Budget End
1999-07-31
Support Year
3
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
168559177
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Wang, Liangjun; Joshi, Preeti; Miller, Ellen L et al. (2018) A Role for Monomethylation of Histone H3-K27 in Gene Activity in Drosophila. Genetics 208:1023-1036
Herzog, Veronika A; Lempradl, Adelheid; Trupke, Johanna et al. (2014) A strand-specific switch in noncoding transcription switches the function of a Polycomb/Trithorax response element. Nat Genet 46:973-981
Simon, Jeffrey A; Kingston, Robert E (2013) Occupying chromatin: Polycomb mechanisms for getting to genomic targets, stopping transcriptional traffic, and staying put. Mol Cell 49:808-24
Rai, Aswathy N; Vargas, Marcus L; Wang, Liangjun et al. (2013) Elements of the polycomb repressor SU(Z)12 needed for histone H3-K27 methylation, the interface with E(Z), and in vivo function. Mol Cell Biol 33:4844-56
O'Meara, M Maggie; Simon, Jeffrey A (2012) Inner workings and regulatory inputs that control Polycomb repressive complex 2. Chromosoma 121:221-34
Smith, Matthew; Mallin, Daniel R; Simon, Jeffrey A et al. (2011) Small ubiquitin-like modifier (SUMO) conjugation impedes transcriptional silencing by the polycomb group repressor Sex Comb on Midleg. J Biol Chem 286:11391-400
Wang, Liangjun; Jahren, Neal; Miller, Ellen L et al. (2010) Comparative analysis of chromatin binding by Sex Comb on Midleg (SCM) and other polycomb group repressors at a Drosophila Hox gene. Mol Cell Biol 30:2584-93
Chen, Shuai; Bohrer, Laura R; Rai, Aswathy N et al. (2010) Cyclin-dependent kinases regulate epigenetic gene silencing through phosphorylation of EZH2. Nat Cell Biol 12:1108-14
Zhu, Changqi C; Bornemann, Douglas J; Zhitomirsky, David et al. (2008) Drosophila histone deacetylase-3 controls imaginal disc size through suppression of apoptosis. PLoS Genet 4:e1000009
Wang, Liangjun; Jahren, Neal; Vargas, Marcus L et al. (2006) Alternative ESC and ESC-like subunits of a polycomb group histone methyltransferase complex are differentially deployed during Drosophila development. Mol Cell Biol 26:2637-47

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