Despite significant advancement in treatments, the leading cause of morbidity and mortality worldwide continues to be cardiovascular diseases (CVDs) with highest incidence of heart failure (HF). It is widely known that epigenetic deregulation is associated with CVD pathophysiology. The epigenetic modification mainly N6-methyladenosine (m6A) is the most conserved internal modification in RNA, which functions as an additional layer of transcriptional and translational control. However, extensive research is required to understand the regulatory role of m6A methylation in the heart. Recent publication in our lab highlighted the deregulation of m6A methylation in cardiac hypertrophy and heart failure, which also imparts changes in protein translation independent of transcription. Further, the role of regulatory m6A in normal cardiac function were studied by cardiomyocyte specific knockout of m6A demethylase FTO (FTO-cKO) in rodents and our data showed that FTO depletion exacerbated cardiac phenotype by quickening heart failure progression with reduced hypertrophy. As a further step, here, we analysed the effect of regulatory m6A methylation in the normal and pressure overloaded FTO-cKO rodents using the widely available computational algorithms.

Firstly, the control and FTO-cKO group mice were subjected to sham and TAC (transverse aortic constriction) operation which is the model of cardiac hypertrophy and heart failure. Echocardiography were performed one week and eight weeks after the surgery to study the attributes of heart failure. Secondly, hearts were isolated from both the control and FTO-cKO mice which underwent sham and TAC; RNA extraction and m6A RNA immunoprecipitation (meRIP) were performed from the isolated mice heart tissue; and lastly, highthroughput m6A sequencing and analysis were performed in order to the study the effect of regulatory m6A methylation.

In consistent with our previous data, deregulation of m6A methylation observed in failing heart tissue compared to the healthy tissue. Notably, FTO demethylase depletion further increased the imbalance in m6A levels in both TAC and sham mice groups. Interestingly, more transcripts (397 with FC 1.5 FDR 0.05) were hypomethylated than hypermethylation in response to TAC surgery in FTO-cKO mice (FTO-cKO TAC vs FTO-cKO sham). Moreover, GO term analysis showed that the hypomethylated transcripts in failing heart are mainly involved in the pathways of apoptosis and collagen formation, whereas the hypermethylated transcripts are part of RNA metabolism and cell cycle progression.

Our data insists that the altered changes in the regulatory m6A levels are responsible for the reduced hypertrophy and faster progression of heart failure. Overall, this indicates the significance of epitranscriptome in CVD pathophysiology and paves way for interesting therapeutic targets for the treatment of heart failure.