Marciniak, D. M. | Search bioRxiv

bioRxiv2024

Nascent transcript O-MAP reveals the molecular architecture of a single-locus subnuclear compartment built by RBM20 and the TTN RNA

Eukaryotic nuclei adopt a highly compartmentalized architecture that influences nearly all genomic processes. Understanding how this architecture impacts gene expression has been hindered by a lack of tools for elucidating the molecular interactions at individual genomic loci. Here, we adapt oligonucleotide-mediated proximity-interactome mapping (O-MAP) to biochemically characterize discrete, micron-scale nuclear neighborhoods. By targeting O-MAP to introns within the TTN pre-mRNA, we systematically map the chromatin loci, RNAs, and proteins within a muscle-specific RNA factory organized around the TTN locus. This reveals an unanticipated compartmental architecture that organizes cis- and trans-interacting chromosomal domains, including a hub of transcriptionally silenced chromatin. The factory also recruits dozens of unique RNA-binding and chromatin-scaffolding factors, including QKI and SAFB, along with their target transcripts. Loss of the cardiac-specific splicing factor RBM20--a master regulator of TTN splicing that is mutated in dilated cardiomyopathy--remodels nearly every facet of this architecture. This establishes O-MAP as a pioneering method for probing single-locus, microcompartment-level interactions that are opaque to conventional tools. Our findings suggest new mechanisms by which coding genes can "moonlight" in nuclear-architectural roles.

genomics↗

bioRxiv2023

Cellular RNA interacts with MAVS to promote antiviral signaling

Immune signaling needs to be well-regulated to promote clearance of pathogens, while preventing aberrant inflammation. Interferons (IFNs) and antiviral genes are activated by the detection of viral RNA by RIG-I-like receptors (RLRs). Signal transduction downstream of RLRs proceeds through a multi-protein complex organized around the central adaptor protein MAVS. Recent work has shown that protein complex function can be modulated by RNA molecules providing allosteric regulation or acting as molecular guides or scaffolds. Thus, we hypothesized that RNA plays a role in organizing MAVS signaling platforms. Here, we show that MAVS, through its central intrinsically disordered domain, directly interacts with the 3' untranslated regions of cellular mRNAs. Importantly, elimination of RNA by RNase treatment disrupts the MAVS signalosome, including newly identified regulators of RLR signaling, and inhibits phosphorylation of the transcription factor IRF3. This supports the hypothesis that RNA molecules scaffold proteins in the MAVS signalosome to induce IFNs. Together, this work uncovers a function for cellular RNA in promoting signaling through MAVS and highlights a generalizable principle of RNA regulatory control of cytoplasmic immune signaling complexes.

immunology↗