Search for: All records

Abstract Heritable gene silencing has been proposed to rely on DNA methylation, histone modifications, and/or non-coding RNAs in different organisms. Here we demonstrate that multiple RNA-mediated mechanisms with distinct and easily detectable molecular signatures can underlie heritable silencing of the same open-reading frame in the nematodeC. elegans. Using two-gene operons, we reveal three cases of gene-selective silencing that provide support for the transmission of heritable epigenetic changes through different mechanisms of RNA silencing independent of changes in chromatin that would affect all genes of an operon equally. Different heritable epigenetic states of a gene were associated with distinct populations of stabilized mRNA fragments with untemplated poly-UG (pUG) tails, which are known intermediates of RNA silencing. These ‘pUG signatures’ provide a way to distinguish the multiple mechanisms that can drive heritable RNA silencing of a single gene. 

SUMMARY Progress in biology has generated numerous lists of genes that share some property. But, advancing from these lists of genes to understanding their roles is slow and unsystematic. Here we use RNA silencing inC. elegansto illustrate an approach for prioritizing genes for detailed study given limited resources. The partially subjective relationships between genes forged by both deduced functional relatedness and biased progress in the field was captured as mutual information and used to cluster genes that were frequently identified yet remain understudied. Studied genes in these clusters suggest regulatory links connecting RNA silencing with other processes like the cell cycle. Many proteins encoded by the understudied genes are predicted to physically interact with known regulators of RNA silencing. These predicted influencers of RNA-regulated expression could be used for feedback regulation, which is essential for the homeostasis observed in all living systems. Thus, among the gene products altered when a process is perturbed are regulators of that process, providing a way to use RNA sequencing to identify candidate protein-protein interactions. Together, the analysis of perturbed transcripts and potential interactions of the proteins they encode could help prioritize candidate regulators of any process. 

ABSTRACT RNAs in circulation carry sequence-specific regulatory information between cells in plant, animal, and host-pathogen systems. Such RNA can cross generational boundaries, as evidenced by somatic double-stranded RNA (dsRNA) in the nematodeC. eleganssilencing genes of matching sequence in progeny. Here we dissect the intergenerational path taken by dsRNA from parental circulation and discover that cytosolic import through the dsRNA importer SID-1 in the parental germline and/or developing progeny varies with developmental time and dsRNA substrates. Loss of SID-1 enhances initiation of heritable RNA silencing within the germline and causes changes in the expression of thesid-1-dependentgenesdg-1that last for more than 100 generations after restoration of SID-1. The SDG-1 protein is enriched in perinuclear Z granules required for heritable RNA silencing but is expressed from a retrotransposon targeted by such silencing. This auto-inhibitory loop reveals how retrotransposons could persist by hosting genes that regulate their own silencing. 

Since double-stranded RNA (dsRNA) is effective for silencing a wide variety of genes, all genes are typically considered equivalent targets for such RNA interference (RNAi). Yet, loss of some regulators of RNAi in the nematodeCaenorhabditis eleganscan selectively impair the silencing of some genes. Here, we show that such selective requirements can be explained by an intersecting network of regulators acting on genes with differences in their RNA metabolism. In this network, the Maelstrom domain-containing protein RDE-10, the intrinsically disordered protein MUT-16, and the Argonaute protein NRDE-3 work together so that any two are required for silencing one somatic gene, but each is singly required for silencing another somatic gene, where only the requirement for NRDE-3 can be overcome by enhanced dsRNA processing. Quantitative models and their exploratory simulations led us to find that (1) changingcis-regulatory elements of the target gene can reduce the dependence on NRDE-3, (2) animals can recover from silencing in non-dividing cells, and (3) cleavage and tailing of mRNAs with UG dinucleotides, which makes them templates for amplifying small RNAs, are enriched within ‘pUG zones’ matching the dsRNA. Similar crosstalk between pathways and restricted amplification could result in apparently selective silencing by endogenous RNAs. 

Interacting molecules create regulatory architectures that can persist despite turnover of molecules. Although epigenetic changes occur within the context of such architectures, there is limited understanding of how they can influence the heritability of changes. Here, I develop criteria for the heritability of regulatory architectures and use quantitative simulations of interacting regulators parsed as entities, their sensors, and the sensed properties to analyze how architectures influence heritable epigenetic changes. Information contained in regulatory architectures grows rapidly with the number of interacting molecules and its transmission requires positive feedback loops. While these architectures can recover after many epigenetic perturbations, some resulting changes can become permanently heritable. Architectures that are otherwise unstable can become heritable through periodic interactions with external regulators, which suggests that mortal somatic lineages with cells that reproducibly interact with the immortal germ lineage could make a wider variety of architectures heritable. Differential inhibition of the positive feedback loops that transmit regulatory architectures across generations can explain the gene-specific differences in heritable RNA silencing observed in the nematodeCaenorhabditis elegans. More broadly, these results provide a foundation for analyzing the inheritance of epigenetic changes within the context of the regulatory architectures implemented using diverse molecules in different living systems.