Covalent modification of histone amino-terminal tails is an important mechanism used to modulate chromatin structure and regulate transcription. In particular, acetylation and methylation of Lys residues in the tails of histones H3 and H4 appear to establish a code that serves to recruit other proteins and alter chromatin structure. The role of histone H3 phosphorylation in Ser10 has been less studied and the mechanisms by which it alters chromatin structure are not understood. This modification appears to be a signal for either chromosome condensation during mitosis or transcription activation during interphase, but the basis for these apparently divergent roles is not known. Also unknown are the extent to which this modification is used to regulate gene expression, the aspects of transcription affected by H3 phosphorylation and the enzymatic or regulatory activities involved in controlling the levels of Ser10-phosphorylated H3. Proposed experiments address the question of how H3 phosphorylation is regulated and what its role is in the control of transcription and chromatin structure in Drosophila. The level of H3 Ser10 phosphorylation appears to be a determinant of transcription activity. The amount of phosphorylated H3 should depend on the activities of protein kinases and protein phosphatases acting on this histone. Preliminary results point to the JIL-1 kinase and PP2A phosphatase as two critical enzymes involved in this process. Experiments will attempt to confirm the involvement of these two enzymes and/or identify additional phosphatases involved in H3 phosphorylation. The proteins SET and pp32 have been shown to regulate PP2A activity in vitro, as well as to inhibit H3 acetylation. Therefore, these two proteins may be part of a switch that controls the interplay between histone H3 phosphorylation and acetylation. A combination of genetic and molecular approaches will be used to characterize these two proteins and study their in vivo function in the control of covalent histone modification. Finally, a genetic screen for modifiers of the JIL-1 mutant phenotype will be carried out in order to identify other factors involved in the control of H3 phosphorylation and chromatin condensation.
This research project will serve as the educational basis for an instructional program aimed at educating Baltimore public high school students in biomedical science careers. A series of educational activities will be carried out with two objectives: increasing the number of undergraduate students from underprivileged backgrounds at Johns Hopkins and training and mentoring these students while in high school and at Hopkins in cutting edge biological research through experiential learning in the laboratory.
