Transposable elements are segments of non-self DNA that have the ability to copy and move themselves within the host genome and hence create a threat to the genome stability and integrity. To avoid the spreading of these selfish elements, organisms have evolved different defence mechanisms such as small RNAs with the piRNA pathway being one of them in the nematode C. elegans germline. To identify novel proteins involved in the spatial and temporal regulation of the chromatin state and dependent on the piRNA pathway, we have developed a germline specific single locus ChIP system (piChIP) and successfully identified an uncharacterized protein with a catalytically dead PARP (poly-ADP ribosyl polymerase) domain that resembles to the TASOR and MPP8 proteins of the recently discovered HUSH complex in humans.
Interestingly, null mutants f this catalytically dead PARP protein showed synthetic lethality when crossed to a piRNA pathway mutant, hrde-1, at 20C, therefore indicating a parallel pathway. Hence, we started to characterize its role together with catalytically active C. elegans PARP-1 by comparing transcriptome and genome wide binding profiles of wild type and null mutant animals. We found by total RNA sequencing that null mutants desilenced retrotransposons, such as CELE45, to a similar extent, suggesting that these two proteins might function in the same pathway. Additionally, these RNA-seq data sets showed no correlation with those of other known piRNA or nuclear RNAi pathway mutants, demonstrating that they might silence a specific subset of targets. Furthermore, PARP-1 ChIP-seq suggested specific binding on a set of SINE retrotransposons including CELE45 and this enrichment was consistent with PARP-1 ChIP-seq performed in a piRNA pathway mutant.
This new protein with a catalytically dead PARP domain shows domain similarity to the HUSH complex components as well as to the TONSL protein, which interacts with BRCA-1 in humans. Since BRCA-1, together with TONSL, recognize R-loops and single strand DNA breaks as a result of transcription deficiencies and PARP-1 mediates DNA damage repair for single and double strand breaks, we speculate that their specific enrichment on retrotransposons could be transcription-dependent. We are now trying to characterize proteomics analysis in relation to RNA pol II function and DNA damage response on retrotransposons.
