Biology subjects

Morao, A. K.

Publications and source records attributed to Morao, A. K..

5 recordsLinked to original sources

bioRxiv2025

Nuclear Argonaute HRDE-1 Sustains Chromatin-Independent Transcriptional Silencing and Epigenetic Inheritance

Epigenetic inheritance of transcriptional silencing is traditionally attributed to chromatin-based mechanisms in which Argonaute-small RNA complexes recruit histone-modifying enzymes. Here we show 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 polyUG-dependent amplification of antisense 22G-RNAs in perinuclear condensates, thereby coupling nuclear transcriptional repression to small RNA biogenesis. HRDE-1 loss causes progressive erosion of 22G-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.

molecular biology↗

bioRxiv2025

DNA supercoiling modulates eukaryotic transcription in a gene-orientation dependent manner

DNA supercoiling is an intrinsic consequence of transcription that must be resolved to maintain proper gene expression. How DNA supercoiling shapes transcription dynamics in chromatinized animal genomes remains unclear. Here, we acutely depleted topoisomerases I and II in Caenorhabditis elegans and applied nascent transcription profiling, nuclear and total RNA-seq, histone modification mapping, and long-read sequencing to capture the immediate transcriptional and chromatin responses to topological stress. We show that the genomic context influences the effect of supercoiling on transcription initiation, elongation and coordinated expression of adjacent genes. The impact of supercoiling on transcription initiation is not uniformly repressive but instead depends on the orientation and proximity of neighboring genes. We find that negative supercoiling promotes coordinated expression of divergent gene pairs, while positive supercoiling inhibits and uncouples expression of convergent genes. DNA supercoiling hinders transcription elongation globally resulting in reduced production of longer transcripts and overall shortening of poly(A) tails. While DNA supercoiling generated by neighboring transcription impacts initiation, elongation defects are driven by local supercoiling generated by the genes own transcription. These elongation effects are not accompanied by global changes in elongation-associated histone modifications but coincide with modest reductions in promoter and enhancer marks. Our findings reveal a directional mechanism by which genome architecture shapes transcriptional responses to DNA supercoiling, uncovering a DNA topological basis for coordinated gene expression in a multicellular organism.

molecular biology↗

bioRxiv2022

Increased gene dosage and mRNA expression from chromosomal duplications in C. elegans

Isolation of copy number variations and chromosomal duplications at high frequency in the laboratory suggested that Caenorhabditis elegans tolerates increased gene dosage. Here we addressed if a general dosage compensation mechanism acts at the level of mRNA expression in C. elegans. We characterized gene dosage and mRNA expression in three chromosomal duplications and a fosmid integration strain using DNA-seq and mRNA-seq. Our results show that on average, increased gene dosage leads to increased mRNA expression, pointing to a lack of genome-wide dosage compensation. Different genes within the same chromosomal duplication show variable levels of mRNA increase, suggesting feedback regulation of individual genes. Somatic dosage compensation and germline repression reduce the level of mRNA increase from X chromosomal duplications. Together, our results show a lack of genome-wide dosage compensation mechanism acting at the mRNA level in C. elegans and highlight the role of epigenetic and individual gene regulation contributing to the varied consequences of increased gene dosage.

genomics↗

bioRxiv2021

Topoisomerases I and II facilitate condensin DC translocation to organize and repress X chromosomes in C. elegans

Condensin complexes are evolutionarily conserved molecular motors that translocate along DNA and form loops. While condensin-mediated DNA looping is thought to direct the chain-passing activity of topoisomerase II to separate sister chromatids, it is not known if topological constraints in turn regulate loop formation in vivo. Here we applied auxin inducible degradation of topoisomerases I and II to determine how DNA topology affects the translocation of an X chromosome specific condensin that represses transcription for dosage compensation in C. elegans (condensin DC). We found that both topoisomerases colocalize with condensin DC and control its movement at different genomic scales. TOP-2 depletion hindered condensin DC translocation over long distances, resulting in accumulation around its X-specific recruitment sites and shorter Hi-C interactions. In contrast, TOP-1 depletion did not affect long-range spreading but resulted in accumulation of condensin DC within expressed gene bodies. Both TOP-1 and TOP-2 depletions resulted in X chromosome transcriptional upregulation indicating that condensin DC translocation at both scales is required for its function in gene repression. Together the distinct effects of TOP-1 and TOP-2 on condensin DC distribution revealed two distinct modes of condensin DC association with chromatin: long-range translocation that requires decatenation/unknotting of DNA and short-range translocation across genes that requires resolution of transcription-induced supercoiling.

molecular biology↗

bioRxiv2021

A noncatalytic activity of the H4K20 demethylase DPY-21 regulates condensin DC binding

Condensin is a multi-subunit SMC complex that binds to and compacts chromosomes. Here we addressed the regulation of condensin binding dynamics using C. elegans condensin DC, which represses X chromosomes in hermaphrodites for dosage compensation. We established fluorescence recovery after photobleaching (FRAP) using the SMC4 homolog DPY-27 and showed that a well-characterized ATPase mutation abolishes its binding. Next, we performed FRAP in the background of several chromatin modifier mutants that cause varying degrees of X-chromosome derepression. The greatest effect was in a null mutant of the H4K20me2 demethylase DPY-21, where the mobile fraction of condensin DC reduced from [~]30% to 10%. In contrast, a catalytic mutant of dpy-21 did not regulate condensin DC mobility. Hi-C data in the dpy-21 null mutant showed little change compared to wild type, uncoupling Hi-C measured long-range DNA contacts from transcriptional repression of the X chromosomes. Together, our results indicate that DPY-21 has a non-catalytic role in regulating the dynamics of condensin DC binding, which is important for transcription repression.

cell biology↗