Phylogenetic And Functional Analysis Of The Ubiquitin Proteasome System in Pristionchus pacificus

Abstract

Transgenerational Epigenetic Inheritance (TEI) is an often overlooked aspect of biological inheritance, whose underlying mechanisms still remain poorly understood. The model nematode Pristionchus pacificus, with its two discrete mouthforms which can be environmentally induced and transgenerationally inherited, offers an excellent model to study this process. Previous work in the lab demonstrated that a ubiquitin ligase gene, ebax-1, is necessary for TEI and that a long term environmental induction of the predatory mouthform can lead to the worms becoming permanently predatory, a phenomenon that was termed ‘canalisation’. It was also seen that hypermutator strains undergo ‘canalisation’ more often. Sequencing of these canalised lines revealed multiple mutations in genes coding for ubiquitin ligases, which were also seen in an EMS mutagenesis screen for mutants defective in TEI. This prompted us to carry out a bioinformatic analysis of the genes coding for the ubiquitin proteasome system in P. pacificus and their phylogenetic relationship with their counterparts in C. elegans, and investigate the role of selected candidate genes in TEI. It was found that most of the core components of the ubiquitin proteasome system are strongly conserved across P. pacificus and C. elegans, some classes of genes have undergone extreme divergence and diversification, including between different strains of P. pacificus, indicating ongoing diversification and evolution. We selected the ubiquitin ligase genes eel-1, ubr-4, ubr-5 and bath-38 as targets for CRISPR mediated knockouts, following which we saw that eel-1 deletion mutants displayed a Transgenerational Inheritance Defective (tid) phenotype. No canalisation was seen for the duration of the assay.