Arantxa Rojas Mechanisms of Epigenetic Inheritance, Department of Developmental and Stem Cell Biology, Institut Pasteur, CNRS UMR3738, Paris. Postdoc in Germano Cecere’s lab

Title: Nuclear Argonaute HRDE-1 Maintains Chromatin-Independent Transcriptional Silencing through a Transgenerational Small RNA Feedback Loop

Abstract: Epigenetic inheritance of transcriptional silencing is traditionally attributed to chromatin- based mechanisms in which Argonaute–small RNA complexes recruit histone-modifying enzymes. In this project, we found that in Caenorhabditis elegans, the nuclear Argonaute HRDE-1 maintains germline transcriptional repression independently of canonical heterochromatin marks. Using inducible, germline-specific HRDE-1 depletion combined with nuclear sorting, CUT&Tag, and nascent transcription profiling, we identified endogenous genes that become transcriptionally activated upon loss of HRDE-1 despite retaining H3K9me3 and H3K23me3. HRDE-1 directly restrains RNA polymerase II (Pol II) while simultaneously promoting antisense small RNA production in perinuclear condensates, thereby coupling nuclear transcriptional repression to small RNA biogenesis. HRDE-1 loss causes progressive erosion of antisense small RNAs and delays silencing re-establishment, revealing a transgenerational feedback loop in which small RNAs, and not chromatin modifications, constitute the primary heritable signal. These findings redefine nuclear Argonautes as active drivers of RNA-based epigenetic inheritance and broaden our understanding of how small RNA pathways maintain transcriptional silencing across generations.