Overlapping and Distinct Roles of Two C. Elegans H3 Lysine 36 Histone Methyltransferases
Establishment and maintenance of cell type-specific gene expression patterns is essential for development and normal tissue function. A growing number of studies demonstrate that epigenetic information contributes to cell fate specification and maintenance, and can be transmitted though mitotic divisions as well as from parents to progeny. Yet, the mechanisms involved in establishing and maintaining epigenetic information, as well as the consequences to gene expression in cells inheriting epigenetic information are not well understood. One form of epigenetic information, post-translational modifications of histones, can regulate gene expression patterns and provide a long-term memory of expression patterns established by transient transcription factor activity during early development. In C. elegans, two antagonistic histone methyltransferases (HMTs) are essential for germline development in a maternal effect manner. MES-2 is part of a PRC2-like complex that methylates H3 lysine 27 (H3K27me3), and MES-4 is one of two H3K36me3 HMTs. This thesis focuses on H3K36me3 and the two enzymes that generate this mark, MES-4 and MET-1. While MES-4 is required for germline development in all conditions, MET-1 is only required at elevated temperatures. Our mass spectrometry analysis of histone tails from C. elegans early embryos confirmed that both MET-1 and MES-4 catalyze H3K36me3, a modification that is generated by only a single enzyme in other organisms. We performed immunostaining studies to investigate the generation of H3K36me3 in the adult germline, its transmission from parents to progeny, and its maintenance during early embryogenesis. Our data show that MET-1 and MES-4 serve unique roles in the generation and transmission of H3K36me3. In the germline, MET-1 co-transcriptionally catalyzes H3K36me3 and is solely responsible for H3K36me3 on the oocyte X chromosome. MES-4 also contributes to H3K36me3 in the germline and is solely responsible for maintenance of H3K36me3 on chromosomes in early embryos, where it operates in a transcription-independent manner.
We discovered that both oocytes and sperm transmit chromosomes carrying H3K36me3 to the embryo. This observation supports the hypothesis that epigenetic information generated in the adult germline can be transmitted to progeny from either parent. To determine if inherited H3K36me3 is required for MES-4 to associate with chromosomes, we generated embryos in which only a subset of chromosomes carry H3K36me3. In these embryos, MES-4 is recruited to H3K36me3-positive chromosomes but not to H3K36me3-negative chromosomes, suggesting MES-4 is recruited to chromosomes by pre-existing H3K36me3. Additionally, as these embryos divide, MES-4 and H3K36me3 are maintained on only a subset of chromosomes until at least the 32-cell stage, likely because MES-4 is propagating H3K36me3 in regions of chromatin with pre-existing H3K36me3. This observation suggests that MES-4 maintains an epigenetic memory of inherited H3K36me3. Together, these data support the model that MET-1 is primarily responsible for generating H3K36me3 on genes expressed in the germline, and that MES-4 is primarily responsible for maintaining an epigenetic memory of inherited H3K36me3 through early embryogenesis, likely to guide gene expression patterns in nascent germ cells.