Key Points
- To characterize the CELF family of RNA binding proteins and determine their role in regulating cell-specific, developmental alternative splicing of cardiac troponin T.
- Assayed RNA-protein interactions in vitro using muscle-specific splicing enhancer (MSE) sequences flanking cardiac troponin T (cTNT) exon 5.
- Tested MSE-dependent exon inclusion in vivo using minigene constructs in transfected cell lines.
- Profiled expression levels and protein isoforms of CELF family members, including ETR-3, across differentiating C2C12 myoblasts and developing heart tissue.
- Identified the CELF family (including CUG-BP, ETR-3, and CELF4) as RNA binding proteins that specifically bind MSE targets in vitro and activate MSE-dependent cTNT exon inclusion in vivo.
- Found that the developmental switch from cTNT exon skipping to inclusion tightly correlates with the induction of ETR-3 protein expression during C2C12 myoblast differentiation.
- Demonstrated that developmentally regulated transitions in ETR-3 protein isoforms correlate with the cTNT splicing switch during cardiac organogenesis.
Structured PICO
PPopulationPreclinical models including in vitro assays, C2C12 myoblasts, and developing heart models
IInterventionCELF family of RNA binding proteins (including CUG-BP and ETR-3)
OOutcomeAlternative splicing of cardiac troponin T (cTNT) exon 5surrogate
The CELF family of RNA binding proteins, particularly ETR-3, are identified as major regulators of developmentally regulated alternative splicing of cardiac troponin T.