Integrity and Stability of PTC Bearing CFTR mRNA and Relevance to Future Modulator Therapies in Cystic Fibrosis.

Luka A Clarke,Vanessa C C Luz,Sofia S Ramalho,Szymon Targowski,Carlos M Farinha,Margarida D Amaral

doi:10.3390/genes12111810

Luka A Clarke, Vanessa C C Luz + Show 4 more

Open Access

https://doi.org/10.3390/genes12111810

Copy DOI

Journal: Genes	Publication Date: Nov 18, 2021
Citations: 9	License type: CC BY 4.0

Affiliation: University of Lisbon

Abstract

Major advances have recently been made in the development and application of CFTR (cystic fibrosis transmembrane conductance regulator) mutation class-specific modulator therapies, but to date, there are no approved modulators for Class I mutations, i.e., those introducing a premature termination codon (PTC) into the CFTR mRNA. Such mutations induce nonsense-mediated decay (NMD), a cellular quality control mechanism that reduces the quantity of PTC bearing mRNAs, presumably to avoid translation of potentially deleterious truncated CFTR proteins. The NMD-mediated reduction of PTC-CFTR mRNA molecules reduces the efficacy of one of the most promising approaches to treatment of such mutations, namely, PTC readthrough therapy, using molecules that induce the incorporation of near-cognate amino acids at the PTC codon, thereby enabling translation of a full-length protein. In this study, we measure the effect of three different PTC mutations on the abundance, integrity, and stability of respective CFTR mRNAs, using CFTR specific RT-qPCR-based assays. Altogether, our data suggest that optimized rescue of PTC mutations has to take into account (1) the different steady-state levels of the CFTR mRNA associated with each specific PTC mutation; (2) differences in abundance between the 3′ and 5′ regions of CFTR mRNA, even following PTC readthrough or NMD inhibition; and (3) variable effects on CFTR mRNA stability for each specific PTC mutation.

Highlights

Cystic Fibrosis (CF) is an autosomal recessive condition caused by mutations in the CF transmembrane conductance regulator (CFTR) gene [1], which encodes a chloride/bicarbonate channel important for maintaining ion homeostasis across epithelial membranes, perhaps most essentially in the airways [2]
We have shown that while nonsense-mediated decay (NMD) is responsible for degradation of up to 80% of premature termination codon (PTC)-bearing CFTR transcripts, there is variation between different tissues, and among both different mutations and individuals with the same genotype [11]
PTC Mutations Cause a Reduction in CFTR mRNA Abundance

Summary

Introduction

Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. Cystic Fibrosis (CF) is an autosomal recessive condition caused by mutations in the CF transmembrane conductance regulator (CFTR) gene [1], which encodes a chloride/bicarbonate channel important for maintaining ion homeostasis across epithelial membranes, perhaps most essentially in the airways [2]. Of the more than 2100 potentially disease-causing mutations in CFTR, approximately 8.4% are nonsense mutations, i.e., single base pair substitutions, which introduce premature termination codons (PTCs) into the CFTR coding sequence [3]. Nonsense mutations, which are classified as Class I mutations in CF, are associated with severe disease phenotypes [4] and have two major effects upon

Methods

Results

Conclusion