As our understanding of epigenetic regulation grows, it is becoming clear that such pathways are involved in many complex cellular processes that have far-reaching impacts on organisms and populations. Using Caenorhabditis elegans as our model of choice, we investigate the molecular mechanisms underpinning epigenetic processes, with a particular focus on germline processes. One area of interest is the involvement of granules – biomolecular condensates of RNA and protein which have been implicated in germline integrity and epigenetic regulation.
In the search for readers of H3K23me3, a recently identified histone mark of interest, we performed a peptide-based pulldown coupled with proteomic analysis by mass spectrometry. After curation of the resulting list of proteins, we have selected two genes for further investigation, tag-250 and Y50D4C.3, that we predict are involved in epigenetic regulation. By observing the AlphaFold-predicted structures of these genes’ protein products, we have highlighted previously unannotated structural features, which in turn provide novel insights into their potential functions. Relevant features include the presence of Tudor domains, which commonly act as readers for histone modifications. TAG-250 additionally contains a LOTUS domain, often found in germ-granule associated proteins, while Y50D4C.3 contains a large intrinsically disordered region (IDR), which are often important for phase separation. Furthermore, Y50D4C.3 shows structural homology to a well-characterised human gene, TDRD3, which has known roles in transcriptional activation and cytoplasmic stress granules.
Through investigating how the knockouts of these genes affect C. elegans at the epigenomic, transcriptomic and phenotypic levels, and tracking the expression and localisation of the gene in vivo, we can obtain an overall picture of the genes’ functions within both individual cells as well as the whole organism. We will present our results on characterising the functions of tag-250 and Y50D4C.3 and their involvement in granules thus far, as well as our future directions.