Background:
SLM2 was found to be elevated in the ventricular biopsies from patients with dilated cardiomyopathy (DCM), which might be a compensatory mechanism to adapt to the increased wall stress and the hypertrophic response of the heart. Although SLM2's upregulation in DCM suggests a therapeutic potential, its functional roles remain underexplored. We aim to investigate the impact of SLM2 knockout on cardiac physiology and splicing regulation.
Methodology:
In order to identify the molecular mechanisms through alternative splicing, we investigated the expression of TTN mRNA isoforms N2B and N2BA in cultured mouse cardiomyocytes upon exogenous expression of SLM2, RBM20, and dominant negative RS domain mutant of RBM20. We also investigated the functional interaction of SLM2 and RBM20 splicing factors by overexpressing the SLM2 together with wild-type and mutant RBM20.
Results:
We observed significant alterations in the expression of TTN alternative splicing isoforms in mouse HL1 cells upon SLM2 and RBM20 overexpression. The combination of exogenous SLM2 expression with Wt and mutant RBM20 simultaneously, supported our observation that SLM2 and RBM20 converge on TTN transcript alternative splicing regulation. SLM2 heart specific KO mice exhibited no significant change in cardiac morphology or function, with echocardiographic parameters comparable to controls. While transcriptomic analysis showed no relevant changes in overall gene expression, it revealed dysregulation in the alternative splicing of sarcomeric genes, suggesting a latent effect of SLM2 depletion. We observed significant overexpression of SLM2 and Sam68 in heart tissue upon ORAB surgery, suggesting a compensatory function of these splicing factors in our pressure overload mouse model. Stress-induced vulnerability was manifested after ORAB surgery, with KO mice displaying a more pronounced decline in ventricular function and higher mortality rates compared to controls, thus revealing a critical role of SLM2 in cardiac stress. Finally, in order to identify the novel target mRNA transcripts that are SLM2 regulated under this condition, we conducted (RNA immunoprecipitation) RIP experiments with different splicing factors in wild type and SLM2 KO mice 8 weeks after ORAB surgery and the results are currently being analyzed.
Conclusion:
Our preliminary data demonstrates that the alternative splicing patterns of mRNAs display differences in SLM2 knockout mice that have undergone ORAB surgery. In conclusion, SLM2 affects sarcomere gene splicing under stress conditions and SLM2 deficiency leads to cardiac vulnerability in a pressure overload model of heart failure.
