Objectives:
MicroRNAs (miRs) have been identified as chief post-transcriptional regulators of cardiac disease progression. In addition, a critical role of the adenosine monophosphate-activated kinase (AMPK) pathway in the development of myocardial hypertrophy has been revealed. Yet, regulation of the AMPK pathway by miRs in the heart has not been addressed. We hypothesized that components of the AMPK pathway are targeted by miRs and alter AMPK signaling in a mouse model of hypertrophic cardiomyopathy (HCM).
Methods and results:
Using real-time PCR, a candidate miR screen that included 22 miRs implicated in pathological cardiac disease and/or metabolic dysregulation was performed on hearts from 60-, 120-, and 240-day-old transgenic HCM male mice harboring an R403Q mutation in the myosin heavy gene. Among early (60 day) elevated miRs were miR-195 and -451. Both miR-195 and -451 have conserved target sites in the 3′ UTR of CAB39 (MO25), a central component of the MO25/STRAD/LKB1 complex that acts as an upstream kinase for AMPK and its subsequent activation. We further confirmed the elevation of miR-195 and -451 by Northern blotting. Next, we demonstrated specific expression and a similar distribution pattern of miR-195 and -451 in cardiomyocytes of R403Q HCM hearts by in situ hybridization. To determine whether the conserved sites in MO25 3′ UTR acted as functional targets, either the miR-195 or miR-451 target sequence was cloned into a luciferase expression vector. MiR-195 but not miR-451 suppressed luciferase activity compared to the missense sequence control vector in C2C12 cells. In addition, over-expression of miR195 in C2C12 cells knocked down MO25 expression levels and downstream AMPK signaling (phosphorylation of Acetyl CoA carboxylase). Finally, parallel changes were measured in 60 day R403Q HCM male hearts that included reduced MO25 expression and lowered phosphorlation of AMPK and Acetyl CoA carboxylase.
Conclusion:
Our findings indicate that miR-195 targets the LKB1/AMPK signaling axis and suggest a functional role for miR-195 elevation in R403Q HCM disease progression.
The Impact of the Interferon/TNF-Related Apoptosis-Inducing Ligand Signaling Axis on Disease Progression in Respiratory Viral Infection and Beyond