Characterization of a Potential Gene Interaction between Chromatin Modifiers Spr-5, Met-2, and Mep-1 in Determining Germline Versus Soma in C. Elegans

In C. elegans, histone modifying enzymes aid in the activation and repression of genes that are required to distinguish germline from soma. For example, SPR-5 is a histone demethylase that removes the activating histone modification H3K4me1/2 and MET-2 is a histone methyltransferase that adds the repressive histone modification H3K9me2. Together, these chromatin modifiers work synergistically to establish a totipotent ground state by shutting down the transcription of germline specific genes that were expressed in the germline of the parents. Previous studies have shown that spr-5;met-2 double mutants produce progeny that experience a severe developmental delay. Transcriptomic analysis indicated that the delay is largely due to inappropriate germline gene expression in somatic tissues. Here we explore the possibility that the chromatin state established in the early embryo by SPR-5 and MET-2 is reinforced in the later embryo by ATP-dependent chromatin remodeling activity. MEP-1 is a component of the ATP-dependent epigenetic deacetylase complex, MEC. Similar to spr-5;met-2 double mutants, mep-1 mutants fail to suppress germline genes in somatic tissues, as indicated by the misexpression germline proteins in the soma of L1 larvae. We tested a potential interaction between these pathways by subjecting spr-5, met-2 and spr-5;met-2 mutants to mep-1 RNAi. mep-1 knockdown results in an exacerbated developmental delay phenotype in these mutant backgrounds. The “triple” mutant is most severe, displaying a full L1 arrest. To fully understand the molecular basis of this synergy, we performed RNA-sequencing. We find very little evidence of new expression changes induced in the triple mutants compared to controls. Instead, we find that expression changes in the spr-5;met-2 double mutants are exacerbated in triple mutants. This suggests that MEP-1 specifically reinforces the repressed chromatin state established by SPR-5 and MET-2 reprogramming at fertilization. These results help establish how chromatin modifying enzymes work together to establish developmental cell fates.