Epigenetic inheritance and evolution in nematodes

Abstract

Piwi-interacting RNAs (piRNAs) are 21-30 nt small RNAs that play key roles in germline development and transposable element silencing in metazoans. The piRNA pathway shows extreme diversity and high rates of evolution across metazoans, with few similarities between the nematode and insect systems where characterization of the pathway is most advanced.

In the first part of this thesis, I focus on the evolution of piRNAs across multiple nematode species, gaining insight on several aspects of piRNA biogenesis. First, I show that nematode piRNA loci are likely to be derived from snRNA promoters. I then describe two alternative modes of piRNA organization in nematode genomes: either clustered at high density in intergenic regions of repressive H3K27me3 chromatin, as found in the model nematode Caenorhabditis elegans; or more broadly distributed across the genome, located within the introns of transcriptionally active genes enriched in H3K36me3 chromatin, as found in the satellite model nematode Pristionchus pacificus. Finally, I study the mechanisms of piRNA transcription termination, showing that promoter-proximal Pol II pausing and premature termination of Pol II contribute to the generation of 28 nt capped piRNA precursors in C. elegans. This data exemplifies how understanding the origin and evolution of a molecular system can illuminate its mechanistic basis.

C. elegans piRNAs can initiate an extremely stable form of transgene silencing in the germline that is stably inherited transgenerationally for multiple generations. Once initiated, silencing can become independent of piRNAs due to the generation of secondary 22G-RNAs downstream of piRNA targeting. 22G-RNAs are inherited through gametes and reamplified each generation by RNA-dependent RNA polymerase activities to enforce silencing. While the mechanisms of epigenetic inheritance of transgene silencing are being intensively studied, little is known about the transgenerational dynamics of 22G-RNA populations targeting endogenous genes. In the second part of this thesis, I show that stochastic changes in 22G-RNAs targeting endogenous genes - epimutations - occur during C. elegans growth in controlled environmental conditions in the laboratory. I show that epimutations arise at a rate 25x higher than genetic mutations, but their stability is limited, with a median duration of 2-3 generations. This data demonstrates that while heritable variation in 22G-RNA levels does exist, it is short lived. Therefore, epimutations may contribute to short-term evolutionary processes, but are unlikely to generate long-term divergence in the absence of DNA sequence changes.