The Evolution of Polycomb Repressive Complex 2: Uncovering gene duplications in eukaryotes and investigating function post-duplication in moss
- Author(s): Yee, Stephen Francis
- Advisor(s): Specht, Chelsea D
- et al.
Gene duplication plays an important role in the evolution of novel protein functions. Gene duplications in Polycomb Repressive Complex 2 (PRC2) proteins could have led to novel biochemical functions. However, since PRC2 regulates gene expression by marking its target genes with Histone 3 Lysine 27 trimethylation (H3K27me3), gene duplication might not alter the complex’s biochemical function, but could instead affect its set of target genes. Thus, gene duplication could result in new genomic targets for one of the duplicate PRC2s, subdivision of the genomic targets between the duplicate PRC2s, or some combination of these two scenarios. To determine where gene duplication, and possible functional divergence, occurred we first reconstructed the evolutionary history of the four core PRC2 proteins in eukaryotes. Gene duplications were detected in each of the four PRC2 core proteins: Enhancer of zeste [E(z)]; extra sex combs (esc); Suppressor of zeste 12 [Su(z)12]; and Chromatin assembly factor 1, p55 subunit (Caf1-55). Vertebrates and plants experienced duplications in their E(z), Su(z)12, and Caf1-55 homologs, whereas invertebrates only underwent a small-scale esc duplication in the Schizophora clade of flies. The fungal Caf1-55 genes duplicated at least twice, but these duplications were only found in fungal clades that had lost the other PRC2 core proteins. The consequences of these PRC2 duplications are discussed—these duplications may have resulted in novel biochemical activity, new genes targeted for repression, or partitioning of PRC2 activity throughout development.
Three different PRC2 complexes exist in Arabidopsis. While some of the three Arabidopsis E(z) homologs can be used interchangeably in PRC2s, each of the three Su(z)12 homologs is unique to its PRC2. The model moss, Physcomitrella patens, also possesses three Su(z)12 homologs, although they arose from independent gene duplications. Unlike Arabidopsis, there is only a single copy of the other PRC2 proteins, so we investigated the evolution and function of the only multi-copy P. patens PRC2 core protein. We amplified five conserved exons from the Su(z)12 genes in mosses and detected multiple lineage-specific Su(z)12 duplication events. To determine the functional consequence of the duplications, we made single gene knockouts for each of the three P. patens Su(z)12 genes and then characterized the phenotypes of the mutants. Although there exists some degree of redundancy among the three Su(z)12 copies, each regulates development of the haploid gametophyte generation differently.
While the P. patens Su(z)12 genes affect gametophyte growth, moss E(z) and esc homologs are required to prevent sporophyte development during the gametophyte generation. A handful of other candidate genes important in gametophyte and sporophyte development have been identified. To identify additional genes required for either gametophyte or sporophyte development, we performed RNA-Seq on RNA from both generations. We found that a large proportion of genes and transcripts were differentially expressed between the two tissue types. These differentially expressed genes were enriched for Gene Ontology categories related to translation and photosynthesis in the gametophyte and DNA binding and vesicle-mediated transport in the sporophyte. The most highly differentially expressed genes between gametophyte and sporophyte tissues could be investigated in future studies to determine their roles in moss development.