PhD Graduate: Lee Mei Chin
Thesis Title: Regulation of core histone gene expression by the Drosophila
melanogaster Oct-1 CoActivator in S-phase (dmOCA-S)
Drosophila histone genes, those encoding core histones in particular, are critical for development and homeostasis. How the expression of Drosophila core histone genes is regulated is largely unknown, with their promoters and associated transcription factors and/or cofactor(s) uncharacterized. In the current study, we reveal that all core histone genes of Drosophila melanogaster contain evolutionarily diversified (“cryptic”) octamer elements, which recruit a common transcription factor POU-domain protein in Drosophila melanogaster 1 (Pdm-1) that, in conjunction with the orthlogous D. melanogaster (dm) Oct-1 CoActivator in S-phase (dmOCA-S), exerts the gene-switching function of the core histone genes. Intriguingly, our studies suggest that dmOCA-S regulates the expression of all core histone genes in a directly coordinated manner, which is in accordance with a simpler regulation mode utilizing degenerate octamer elements. This is in line with the Duplication-Degeneration-Innovation (DDI) model and of significant implication in the metazoan evolution study.
Collectively, our studies broaden the roles of the multi-functional Pdm-1 as a transcription factor critical for histone biosynthesis, position dmOCA-S as a universal coactivator in the Drosophila core histone transcription pathway, and establish that dmOCA-S is recruited to the Drosophila histone locus bodies (HLB) during S phase to mark the S phase transcriptional activation of Drosophila core histone genes. Our biochemical approach also paves the way for genetic analysis of dmOCA-S, which is downstream of a cdk/cyclin signaling that promotes S-phase entry and progression.
Figure Legend: dmOCA-S is recruited to Drosophila Histone Locus Bodies during S phase.
(A) A transcriptional regulation pathway of D. melanogaster core histone genes. All the core histone genes contain multiple octamer sites in their promoters for recruitment of the common transcription factor Pdm-1, which in turn recruits co-activator dmOCA-S for the coordinated expression of all Drosophila core histone genes. (B-E) Random cells images. (F) Cell cycle profiles of random cells; ~20% of the cells were in the S-phase. A similar percentage was obtained when counting 100 randomly picked cells using HLB-foci-staining as a criterion. (G-J) Early S-phase cells images that show weak HLB foci and co-localization with nuclear dmGapdh. (K) Early S-phase cell cycle profiles. (L-O) Mid-S-phase cells images that show prominent HLB foci with strong co-localization with nuclear dmGapdh foci. (P) Mid-S-phase cell cycle profiles. (Q-T) Late S- and early G2-phase cells images that show decreased nuclear dmGapdh and HLB foci in size and number. (U) Late S- and early G2-phase cell cycle profiles. (V-Y) G2-phase cells, with no HLB and dmGapdh nuclear foci. (Z) G2-phase cell cycle profiles. When appropriate, arrows indicate HLB (B, G, L, Q, V) and nuclear dmGapdh (C, H, M, R, W) foci and their nuclear co-localization (D, I, N, S, X), which was confirmed by superimposing to DAPI nuclei-staining images (E, J, O, T, Y). HLB foci were stained with the MPM-2 antibody; nuclear dmGapdh foci were stained by anti-p38/GAPDH antibodies. Bar, 10 μM.
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