miR-106a promotes cardiac hypertrophy by targeting mitofusin 2

Highlights

• As a new molecular target miR-106a induces hypertrophy in cardiomyocytes.

• Mfn2 as a cardiac protective factor alleviates hypertrophy.

• miR-106a promotes cardiac hypertrophy by inhibiting Mfn2.

Abstract

Pathological cardiac hypertrophy is a main factor leading to heart failure and associated sudden death. Improved understanding of the underlying molecular mechanisms should aid better treatment of the disease. This study aimed to test our hypothesis that a microRNA miR-106a played an important role in the development of cardiac hypertrophy through targeting mitofusin 2 (Mfn2), a mitochondrial fusion protein known to be critical in regulating cardiac function. miR-106a was robustly upregulated in hypertrophied myocardium both in vivo and in vitro. Forced transient expression of miR-106a in otherwise healthy cardiomyocytes induced the hypertrophic phenotypes resembling those produced by angiotensin II (AngII) exposure. Knockdown of miR-106a by its specific inhibitor nearly completely reversed the hypertrophic phenotypes induced by AngII pretreatment and pressure overload. On the other hand, Mfn2 was markedly downregulated in hypertrophic heart and cardiomyocytes, which was in reciprocal to expression of miR-106a. Mfn2 was experimentally validated as a direct target gene for miR-106a. Overexpression of Mfn2 counteracted the hypertrophic responses induced by miR-106a, whereas silence of Mfn2 by its siRNA abolished the anti-hypertrophic property of miR-106a inhibitor. Furthermore, overexpression of Mfn2 alleviated the hypertrophic phenotypes induced by AngII in cultured cardiomyocytes, while Mfn2 siRNA alone was able to induce hypertrophic changes in cultured cardiomyocytes. Moreover, AngII and miR-106a treatment cultured cardiomyocytes mitochondria presented cristae defects, considerable depolarization of mitochondrial membrane and increased ROS production. These alterations were reversed by miR-106a inhibitor or overexpression of Mfn2. Taken together, our findings indicate miR-106a as an important factor to promote hypertrophic progress and suggest miR-106a as a new molecular target for the treatment of pathological hypertrophy. The present study also uncovered a novel relationship between miR-106a and Mfn2, with Mfn2 as a downstream signaling mediator of miR-106a.

Introduction

Cardiac hypertrophy was thought to be an adaptive response to various stresses to maintain normal cardiac functions. It is now believed that cardiac hypertrophy in many cases is a pathological process, as it can lead to heart failure, arrhythmia and even sudden death [1], [2]. Cardiac hypertrophy is characterized by cardiomyocyte growth in size at the cellular level, increased protein synthesis and re-expression of many fetal genes at the molecular level, and increased left ventricular (LV) wall thickness and impaired cardiac function at the organ level [3]. It is known that cardiac hypertrophy can be induced by a variety of cardiovascular disorders such as long-standing hypertension, valvular insufficiency and stenosis, myocarditis, and diabetic cardiomyopathy.

An increasing body of evidence indicates that microRNAs (miRNAs) serve as an important layer of the regulatory network in cardiovascular disease including cardiac hypertrophy. miRNAs are a class of small, ~ 22-nucleotides-long, non-coding RNAs that are believed to regulate the expression of up to 30% of human genes by binding to the 3′-UTR of mRNAs [4], [5]. By regulating expression of protein-coding genes, miRNAs can often result in well-defined phenotypes of pathophysiological processes in humans and animal models. For example, ectopic expression of miR-1, miR-328 and miR-21 has been reported to be critically involved in acute myocardial infarction, arrhythmogenesis and cardiac hypertrophy [6], [7], [8]. The miR-17-92 cluster has been shown to participate in cardiac ischemic/reperfusion injury [9], and deregulation of miR-17-92 causes lethal hypertrophic cardiomyopathy and arrhythmogenesis [10]. Moreover, these miRNAs have also been found to be required for cardiomyocyte proliferation in postnatal and adult hearts [11]. In our pilot study aiming to investigate the role of the miR-17-5p seed family (the miRNA sharing the same seed site as miR-17-5p) in cardiac hypertrophy, we found that miR-106a was substantially up-regulated. Our theoretical analysis further indicated that this miR-106a has the potential to target a gene called Mfn2 which is reportedly important in regulating cardiac hypertrophy [12], [13]. However, the role of miR-106a in the heart has not been studied.

Mfn2 is a key member of mitofusin protein family, which is located in outer membrane of mitochondrial and acts to maintain the structure of mitochondrial [14]. Mfn2 is highly expressed in tissues and cells with high metabolism rates such as heart, skeletal muscle, brain, kidney, and liver [15], [16], [17], [18], [19]. It promotes mitochondrial fusion to interchange mitochondrial DNA or energy with nearby mitochondria. It also interlinks mitochondrial and sarcoplasmic reticulum to promote mitochondrial respiration [20], [21]. Mfn2 is known as a cell proliferation suppressor gene, probably by inhibiting the MAPK/ERK pathway [15], [22]. Recent studies indicate that Mfn2 deficiency is involved in cardiovascular disease [17], [23], [24].

On the basis of these facts, we hypothesized that miR-106a might be involved in hypertrophic growth of the heart through targeting Mfn2. This study was designed to examine this notion in a mouse model of cardiac hypertrophy and a cellular model of cardiomyocyte hypertrophy.