Titin Circular RNAs Create a Back-Splice Motif Essential for SRSF10 Splicing

Area of Science:

• Cardiovascular Biology
• Molecular Genetics
• RNA Biology

Background:

• Titin (TTN) protein provides cardiomyocyte elasticity; TTN mutations cause hereditary heart failure.
• TTN transcripts produce circular RNAs (circRNAs) dependent on RBM20.
• These circRNAs possess a unique motif that binds SRSF10, regulating splicing.

Purpose of the Study:

• Investigate the function of TTN-derived circRNAs in cardiomyocyte regulation.
• Determine the mechanism by which TTN circRNAs interact with splicing factors RBM20 and SRSF10.
• Assess the impact of TTN circRNA loss on cardiac function and gene splicing.

Main Methods:

• Calculated genetic constraint of the circRNA motif in the gnomAD database.
• Focused on cTTN1, the highest expressed TTN circRNA in the human heart.
• Used shRNAs to selectively deplete cTTN1 in human iPSC-derived cardiomyocytes.

Main Results:

• Human genetics indicates reduced tolerance for mutations in the circRNA motif.
• Selective loss of cTTN1 caused cardiac structural abnormalities, apoptosis, and reduced contractility.
• Loss of cTTN1 led to aberrant splicing of cardiomyocyte genes (MEF2A, CASQ2, TTN) and distorted RBM20 localization and splicing.

Conclusions:

• TTN-derived circRNAs are essential for normal splicing of muscle genes via RBM20 and SRSF10.
• The TTN transcript has regulatory roles beyond its structural function.
• The unique sequence of TTN circRNA back-splice junctions represents a novel source of functional sequence variation.