Biology subjects

Cortazar, M. A.

Publications and source records attributed to Cortazar, M. A..

3 recordsLinked to original sources

bioRxiv2025

Genomic stop codon scanning reveals quantitative principles of nonsense-mediated mRNA decay

Nonsense-mediated mRNA decay (NMD) degrades transcripts containing premature termination codons (PTCs), critically shaping the disease outcomes of protein-truncating variants. While existing NMD rules categorize PTCs as NMD-triggering or NMD-evading, they cannot quantitatively predict the degree of NMD activity for a given endogenous PTC variant. To provide quantitative insight into NMD, we used saturation genome editing (SGE) to systematically introduce all possible PTCs (TAA, TAG, TGA) at every codon position spanning the first, penultimate, and three internal exons of the Lamin A/C (LMNA) gene. Combining targeted sequencing with NMD inhibition, we measured mRNA expression and NMD activity for 722 PTCs and 211 single nucleotide variants (SNVs). Our data validate known positional trends in NMD activity but reveal unexpected complexity. In the penultimate exon, the PTC position effect extends beyond the binary 50-55 nucleotides (nt) rule, revealing a quantitative relationship between the PTC-EJC distance and NMD activity. At the 5 end, NMD is completely absent in the first 21 codons of LMNA, followed by sharp activation over the next 4 codons. Both patterns, at the 5 end and in the penultimate exon, are unexplained by current models. Finally, internal exons show robust NMD with outliers consistently mapping to predicted readthrough-permissive sequence contexts, including the conserved readthrough promoting TGA-CT motif. This comprehensive dataset provides an unprecedented resource for understanding the quantitative impact of PTC position and sequence context on NMD, with direct implications for the clinical interpretation of nonsense variants in the human population.

genomics↗

bioRxiv2024

Peptidyl-tRNA hydrolysis rate influences the efficiency of nonsense-mediated mRNA decay

Nonsense variants underlie many genetic diseases. The phenotypic impact of nonsense variants is determined by nonsense-mediated mRNA decay (NMD), which degrades transcripts with premature termination codons (PTCs). Despite its clinical importance, the factors controlling transcript-specific and context-dependent variation in NMD activity remain poorly understood. Through analysis of human genetic datasets, we discovered that the amino acid preceding the PTC strongly influences NMD activity. Notably, glycine codons promote robust NMD efficiency and show striking enrichment before PTCs but depletion before normal termination codons (NTCs). This glycine-PTC enrichment is particularly pronounced in genes tolerant to loss-of-function variants, suggesting evolutionary selection or neutrality conferred by efficient elimination of truncated proteins from non-essential genes. Using biochemical assays and massively parallel reporter analysis, we demonstrated that the peptide release rate during translation termination varies substantially with the identity of the preceding amino acid and serves as the primary determinant of NMD activity. We propose a "window of opportunity" model where translation termination kinetics modulate NMD efficiency. By revealing how sequence context shapes NMD activity through translation termination dynamics, our findings provide a mechanistic framework for improved clinical interpretation of nonsense variants.

molecular biology↗

bioRxiv2021

The myopathic transcription factor DUX4 induces the production of truncated RNA-bindingproteins in human muscle cells

Nonsense-mediated RNA decay (NMD) is a surveillance mechanism that degrades both canonical and aberrant transcripts carrying premature translation termination codons. NMD is thought to have evolved to prevent the synthesis of toxic truncated proteins. However, whether global inhibition of NMD results in widespread production of truncated proteins is unknown. A human genetic disease, facioscapulohumeral muscular dystrophy (FSHD) features acute inhibition of NMD upon expression of the disease-causing transcription factor, DUX4. Here, using a cell-based model of FSHD, we show the production of hundreds of truncated proteins from physiological NMD targets. Using ribosome profiling, we map the precise C-terminal end of these aberrant truncated proteins and find that RNA-binding proteins are especially enriched for aberrant truncations. The stabilized NMD isoform of one RNA-binding protein, SRSF3, is robustly translated to produce a stable truncated protein, which can also be detected in FSHD patient-derived myotubes. Notably, ectopic expression of truncated SRSF3 alone confers toxicity and its downregulation is cytoprotective. Our results demonstrate the genome-scale impact of NMD inhibition. This widespread production of potentially deleterious truncated proteins has implications for FSHD biology as well as other genetic diseases where NMD is therapeutically modulated.

genomics↗