Argonaute2 Mediates Transcriptional Silencing of Spermatogenesis Genes in Drosophila melanogaster

NAZER EZEQUIEL; CHINEN, MADOKA; DALE, RYAN K.; LEI, ELISSA P

Congreso; FASEB meeting on Transcription, Chromatin and Epigenetics; 2015

Argonauteproteins are commonly known as core components of RNA silencing pathways.However, recent work has shown that Argonaute proteins also possess otherfunctions, such as regulation of transcription, splicing and chromatinarchitecture. In particular, Drosophila Argonaute2 (AGO2) has been shownto exhibit substantial genome-wide colocalization with the chromatin insulatorproteins CTCF and CP190. Insulatorproteins possess the ability to mediate inter- and intra-chromosomalinteractions, thus connecting distant regulatory elements in the genome. Forexample, looping between insulator sites could bring an enhancer in proximityto a promoter, thus activating transcription of a gene. Recently, our labprovided the first evidence for an Argonaute protein functioning directly oneuchromatin and affecting enhancer-promoter interaction at the homeotic AbdB-locus independently of the RNAinterference (RNAi) pathway. These results raise the interesting possibilitythat AGO2 could modulate global chromatin architecture, which in turn affectstranscription regulation. To test this hypothesis,mRNA-Seq libraries from null (AGO2[51B]) and RNA slicing activity (AGO2[V966M])AGO2 mutant larvae were constructed.Our preliminary data suggest that the absence of AGO2 de-repressestranscription of a set of spermatogenesis genes, independently of its catalyticactivity. In this regard, AGO2 knockdown in embryonic and larval female celllines also showed to promote up-regulation of spermatogenesis genes. Inaddition, one of the genes de-repressed is nht,a spermatogenesis transcription factor. Interestingly, the nht null mutant suppresses the up-regulation of spermatogenesisgenes observed in AGO2[51B] mutants, thus suggesting a hierarchical role ofAGO2 over nht to silence aspermatogenesis gene program. Overall, these results raise the interestingpossibility that AGO2 is involved in transcriptional silencing of atestis-specific gene expression program throughout female development, thuscontributing to proper sexual dimorphism. Currently, a set of experimentalapproaches including chromatin immunoprecipitation (ChIP), chromosomalconformation assays (3C) and DNA-FISH are underway in order to provide molecularinsights about the AGO2 role for mediating transcriptional silencing ofspermatogenesis genes in Drosophila.