The long term objective of this study is to gain a molecular understanding of epigenetic processes of gene silencing and chromatin remodeling. PEV in Drosophila has served as a major paradigm for the identification and genetic analysis of evolutionary conserved determinants of epigenetic regulation of chromatin structure and gene silencing. We present evidence that the JIL-1 histone H3S10 chromosomal kinase functions to maintain euchromatic regions by antagonizing heterochromatization and gene silencing at ectopic locations by methyltransferase mediated histone H3K9 dimethylation and HP1 recruitment. Consequently, to understand regulation of heterochromatin formation and gene silencing in Drosophila it will be crucial to determine the molecular mechanisms of JIL-1's role in this process and JIL-Ts placement in the genetic hierarchy. Thus, we will test the hypothesis that JIL-1 functions to counterbalance the spread of heterochromatic factors by 1) determining the changes in the distribution of chromatin markers that are diagnostic for active (euchromatic) or silenced (heterochromatic) chromatin in JIL-1 mutant backgrounds, by 2) conducting genetic experiments to define the in vivo pathway in which JIL-1 functions to maintain chromosomal domains and determine JIL-Ts place in the genetic hierarchy, and by 3) determining the molecular mechanisms underlying the loss of viability and perturbation of chromatin structure in JIL-1 mutant backgrounds. Furthermore, we propose that changes in PEV observed in JIL-1 loss-of-function mutants will be accompanied by changes in chromatin structure at the nucleosomal level in a chromatin context-dependent manner. To test these hypotheses we will conduct a series of genetic and biochemical experiments designed to examine how JIL-1 affects nucleosome organization and PEV using chromosomal inversions and insertions of a reporter gene into different chromatin environments. We will directly test the hypothesis that alterations in chromatin structure are caused by the phosphorylation state of histone H3S10 by targeting JIL-1 to ectopic chromosome locations. Finally, we will define the specific domains of JIL-1 that are necessary for chromatin structure regulation in PEV and determine the role of kinase activity in this process by generating """"""""kinase dead"""""""" constructs and expressing the mutated constructs of JIL-1 transgenically in null mutant flies. Gene silencing is a critical developmental process relevant to many human health problems that include cancer. Thus, the proposed studies of JIL-1 will provide important new insights into the molecular mechanisms of how kinase activity modulates chromatin structure and gene regulation that are directly relevant to humans. ? ? ?

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Molecular Genetics B Study Section (MGB)
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Carter, Anthony D
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Iowa State University
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Li, Yeran; Wang, Chao; Cai, Weili et al. (2017) H2Av facilitates H3S10 phosphorylation but is not required for heat shock-induced chromatin decondensation or transcriptional elongation. Development 144:3232-3240
Wang, Chao; Li, Yeran; Cai, Weili et al. (2014) Histone H3S10 phosphorylation by the JIL-1 kinase in pericentric heterochromatin and on the fourth chromosome creates a composite H3S10phK9me2 epigenetic mark. Chromosoma 123:273-80
Cai, Weili; Wang, Chao; Li, Yeran et al. (2014) Genome-wide analysis of regulation of gene expression and H3K9me2 distribution by JIL-1 kinase mediated histone H3S10 phosphorylation in Drosophila. Nucleic Acids Res 42:5456-67
Girton, Jack; Wang, Chao; Johansen, Jørgen et al. (2013) The effect of JIL-1 on position-effect variegation is proportional to the total amount of heterochromatin in the genome. Fly (Austin) 7:129-33
Wang, Chao; Yao, Changfu; Li, Yeran et al. (2013) Evidence against a role for the JIL-1 kinase in H3S28 phosphorylation and 14-3-3 recruitment to active genes in Drosophila. PLoS One 8:e62484
Li, Yeran; Cai, Weili; Wang, Chao et al. (2013) Domain requirements of the JIL-1 tandem kinase for histone H3 serine 10 phosphorylation and chromatin remodeling in vivo. J Biol Chem 288:19441-9
Yao, Changfu; Ding, Yun; Cai, Weili et al. (2012) The chromodomain-containing NH(2)-terminus of Chromator interacts with histone H1 and is required for correct targeting to chromatin. Chromosoma 121:209-20
Wang, Chao; Cai, Weili; Li, Yeran et al. (2012) H3S10 phosphorylation by the JIL-1 kinase regulates H3K9 dimethylation and gene expression at the white locus in Drosophila. Fly (Austin) 6:93-7
Wang, Chao; Cai, Weili; Li, Yeran et al. (2011) The epigenetic H3S10 phosphorylation mark is required for counteracting heterochromatic spreading and gene silencing in Drosophila melanogaster. J Cell Sci 124:4309-17
Wang, Chao; Girton, Jack; Johansen, Jorgen et al. (2011) A balance between euchromatic (JIL-1) and heterochromatic [SU(var)2-5 and SU(var)3-9] factors regulates position-effect variegation in Drosophila. Genetics 188:745-8

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