Biology subjects

Uzonyi, A.

Publications and source records attributed to Uzonyi, A..

3 recordsLinked to original sources

bioRxiv2023

Dissecting the basis for differential substrate specificity of ADAR1 and ADAR2

Millions of adenosines are deaminated throughout the transcriptome by ADAR1 and ADAR2, modulating double-stranded RNA (dsRNA) immunogenicity and recoding mRNA. The high variability in the susceptibility of different adenosines to editing begs the question of what are the determinants of substrate specificity. Here, we systematically monitor how secondary structure modulates ADAR2 vs ADAR1 substrate selectivity, on the basis of systematic probing of thousands of synthetic sequences transfected into ADAR1-deleted cell lines exogenously expressing either ADAR2 or ADAR1. In both cases, structural disruptions gave rise to symmetric, strand-specific induced editing at a fixed offset, but of varying length: -26 nt for ADAR2, and -35 nt for ADAR1. We dissect the basis for the differences in offset between ADAR1 and ADAR2 via diverse mutants, domain-swaps, and ADAR evolutionary homologs, and reveal that it is encoded by the differential RNA binding domain architecture. We demonstrate that this offset-enhanced editing can allow an improved design of ADAR2-recruiting therapeutics, with proof-of-concept experiments suggestive of increased on-target and potentially decreased off-target editing. Our findings provide novel insight into the determinants guiding ADAR2 substrate selectivity and into the roles of the RNA binding domains of ADAR1 and ADAR2 in mediating differential targeting, and should facilitate the design of improved ADAR-recruiting therapeutics.

biochemistry↗

bioRxiv2022

The lncRNA landscape of cardiac resident macrophages and identification of Schlafenlnc as a regulator of macrophage migratory function

Cardiac resident macrophages (crMPs) were recently shown to exert pivotal functions in cardiac homeostasis and disease, but the underlying molecular mechanisms are largely unclear. Long non-coding RNAs (lncRNAs) are increasingly recognized as important regulatory molecules in a number of cell types, but neither the identity nor the molecular mechanisms of lncRNAs in crMPs are known. Here, we have employed deep RNA-seq and single cell RNA sequencing to resolve the crMP lncRNA landscape from healthy and diseased murine myocardium. CrMPs express previously unknown and highly cell type-specific lncRNAs, among which one lncRNA, termed Schlafenlnc, was particularly abundant and enriched in crMPs. We found Schlafenlnc to be necessary for migration-associated gene expression in macrophages in vitro and in vivo and essential for their adhesion and migration. Collectively, our data provide a basis to the systematic characterization of lncRNAs in crMPs and establish Schlafenlnc as a critical regulator of macrophage migratory functions.

cell biology↗

bioRxiv2022

Exon-intron architecture determines mRNA stability by dictating m6A deposition

N6-methyladenosine (m6A), a widespread destabilizing mark on mRNA, is non-uniformly distributed across the transcriptome, yet the basis for its selective deposition is unknown. Here, we uncover that m6A deposition is not selective. Instead, m6A distribution is exclusion-based: m6A-consensus harboring sites are methylated by default, unless they are within a window of up to [~]200 nt from an exon-intron junction. A simple model, relying exclusively on presence of m6A motifs and exon-intron architecture allows high accuracy recapitulation of experimentally-measured m6A profiles and of all m6A hallmarks. We further establish that m6A serves as the long-sought mechanism underlying the strong association between exon-intron architecture and mRNA stability. Our findings establish a mechanism by which the memory of nuclear RNA splicing is covalently etched on an mRNA, in the form of m6A, and determines its cytoplasmic stability, with broad implications on the regulation, function, and evolution of both m6A and mRNA stability.

molecular biology↗