The RNA-binding protein QKI governs a muscle-specific alternative splicing program that shapes the contractile function of cardiomyocytes

Abstract

Aims In the heart, splicing factors orchestrate the functional properties of cardiomyocytes by regulating the alternative splicing of multiple genes. Work in embryonic stem cells has shown that the splicing factor Quaking (QKI) regulates alternative splicing during cardiomyocyte differentiation. However, the relevance and function of QKI in adult cardiomyocytes remains unknown. In this study we aim to identify the in vivo function of QKI in the adult mouse heart. Methods and Results We generated mice with conditional deletion of QKI in cardiomyocytes by the Cre-Lox system. Mice with cardiomyocyte-specific deletion of QKI died during the fetal period (E14.5), without obvious anatomical abnormalities of the heart. Adult mice with tamoxifen-inducible QKI deletion rapidly developed heart failure associated with severe disruption of sarcomeres, already 7 days after knocking out QKI. RNA sequencing revealed that QKI regulates the alternative splicing of more than 1000 genes, including sarcomere and cytoskeletal components, calcium handling genes and (post)transcriptional regulators. Many of these splicing changes corresponded to the loss of muscle-specific isoforms in the heart. Forced overexpression of QKI in cultured neonatal rat ventricular myocytes directed these splicing events in the opposite direction, and enhanced contractility of cardiomyocytes. Conclusion Altogether, our findings show that QKI is an important regulator of the muscle-specific alternative splicing program that builds the contractile apparatus of cardiomyocytes. Translational perspective Alternative splicing generates protein isoforms to maintain mechanical, structural, and metabolic properties of cardiomyocytes. We are the first to show that QKI is one of the essential splicing factors in the adult heart. During heart failure, alternative splicing of numerous genes is altered, thereby affecting cardiac function. Recent observations that QKI expression is downregulated in hearts of heart failure patients indicates that loss of QKI-mediated processes contributes to decreased sarcomere organization in these patients. Modulation of QKI activity may serve as a future therapeutic strategy to adapt cardiac isoform expression and improve cardiac function in heart failure patients.

RNA-Binding Proteins in Cardiomyopathies

“…It is also worth mentioning that many other RBPs, which are not discussed in this review, are involved in regulating the alternative splicing of cardiac genes, although no cardiomyopathy phenotypes were firmly associated with loss of their function. For example, the cardiomyocyte-specific deletion of Mbnl1 and Mbnl2 in mice causes heart spliceopathy [ 101 ], and the conditional knockout of QKI in the cardiomyocytes of adult mice leads to heart failure by disrupting the alternative splicing of cardiac genes with functions in the sarcomere and cytoskeleton [ 102 ]. Thus, we may expect that their combined actions coordinate alternative splicing events in heart development, function, and homeostasis.…”

Quaking regulates circular RNA production in cardiomyocytes

“…Numerous changes in alternative splicing were identified, mostly in sarcomeric, cytoskeletal and Ca 2+ -handling genes. We thus concluded that QKI is a major regulator of striated muscle identity by governing a muscle-specific alternative splicing program in cardiomyocytes ( Montañés-Agudo et al, 2023a ). This extended earlier studies, in which an essential role for QKI was demonstrated in cardiomyocyte differentiation of cardiac progenitors and in the specification of cardiac mesoderm ( Chen et al, 2021 ; Fagg et al, 2022 ).…”

“…Our laboratory has recently generated a mouse model with a conditional deletion of Qki in cardiomyocytes to study the function of QKI in the heart ( Montañés-Agudo et al, 2023a ). Removal of Qki in the adult heart induces dilation of the ventricles and a rapid decline in cardiac function, and this is associated with severe disruption of sarcomere organization.…”

“…Given the essential role of QKI in the alternative splicing program of cardiomyocytes ( Montañés-Agudo et al, 2023a ), and the identification of QKI as a major regulator of circRNA formation during EMT ( Conn et al, 2015 ), we hypothesized that QKI is also crucial for the formation of circRNAs in the heart. In this study, we explored QKI-dependent circRNA formation in the heart by using our previously characterized cardiomyocyte-specific Qki -knockout (KO) mouse model.…”

Therapeutic potential of alternative splicing in cardiovascular diseases

“…A spliceosome is composed of five small nuclear ribonucleoproteins (snRNPs) and over 100 auxiliary proteins and have been extensively studied and reviewed. 7 , 8 , 9 Non-snRNP factors involved in splicing or AS include serine and arginine-rich (SR) proteins 10 ; heterogeneous nuclear ribonucleoproteins (hnRNPs) 11 ; hnRNP-like proteins (such as RBFOX proteins, 12 CELF proteins, 13 NOVA, 14 TDP-43 15 and QKI 16 ); and other regulators, such as MBNL, 17 HuR 18 and RBM proteins. 19 , 20 Among them, SR proteins and hnRNPs are the most important splicing factors that regulate AS, inhibiting strong splice sites or amplifying the power of weak splice sites.…”