Abstract
We previously identified several mRNAs encoding components of the secretory pathway, including signal recognition particle (SRP) subunit mRNAs, among transcripts associated with the RNA-binding protein CELF1. Through immunoprecipitation of RNAs crosslinked to CELF1 in myoblasts and in vitro binding assays using recombinant CELF1, we now provide evidence that CELF1 directly binds the mRNAs encoding each of the subunits of the SRP. Furthermore, we determined the half-lives of the Srp transcripts in control and CELF1 knockdown myoblasts. Our results indicate CELF1 is a destabilizer of at least five of the six Srp transcripts and that the relative abundance of the SRP proteins is out of balance when CELF1 is depleted. CELF1 knockdown myoblasts exhibit altered secretion of a luciferase reporter protein and are impaired in their ability to migrate and close a wound, consistent with a defect in the secreted extracellular matrix. Importantly, similar defects in wound healing are observed when SRP subunit imbalance is induced by over-expression of SRP68. Our studies support the existence of an RNA regulon containing Srp mRNAs that is controlled by CELF1. One implication is that altered function of CELF1 in myotonic dystrophy may contribute to changes in the extracellular matrix of affected muscle through defects in secretion.
Highlights
The concept of RNA regulons, in which functionally related genes are co-regulated by specific RNA-binding proteins (RBPs), was first proposed by Keene and Tenenbaum in 2002 [1]
Amongst the top 5% most enriched transcripts were several mRNAs encoding factors belonging to the secretory pathway, including three of the six protein subunits of the Signal Recognition Particle (Srp54, Srp72 and Srp68)
CELF1 CLIP-seq performed by others using mouse C2C12 cells identified binding sites in the 3’UTRs of Srp9, Srp14 and Srp72 mRNAs [5]
Summary
The concept of RNA regulons, in which functionally related genes are co-regulated by specific RNA-binding proteins (RBPs), was first proposed by Keene and Tenenbaum in 2002 [1]. The advent of RNA immunoprecipitation-based high throughput approaches including CLIP-seq [2] and PAR-CLIP [3] has facilitated the identification of large datasets of mRNA targets for a variety of RBPs, including CELF1 [4,5,6,7,8]. CELF1 Coordinates Decay of SRP Subunit mRNAs through National Science Foundation -NRT Award # 1450032 to Thomas Chen, Carol Wilusz & Asa Ben-Hur. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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