Gαq Signaling in the Regulation of Autophagy and Heart Failure

REVIEW The adult heart undergoes distinct remodeling processes in response to either acute or chronic insults, which can lead to myocyte hypertrophy and ventricular wall thickening or myocyte elongation, death, and ventricular dilatation.1–3 Although an increase in cardiac tissue mass diminishes systolic wall stress to improve contractile performance in the short term, prolonged hypertrophy in response to pathological signaling is associated with a progression toward decompensated heart failure (HF).1–3 Accumulating evidence from studies in human patients and animal models suggests that in most instances hypertrophy is not a compensatory response to the change in mechanical load but rather is a maladaptive process.1,2,4 On the other hand, physiological hypertrophy, as occurs during normal postnatal development or in highly trained athletes, represents a beneficial form of cardiac growth.5 Numerous neuroendocrine and autocrine factors such as phenylephrine, angiotensin II, and endothelin-1 are well-known inducers of cardiomyocyte hypertrophy, and inhibition of their actions can be beneficial for the treatment of chronic HF after cardiac hypertrophy.2,3 Many such agonists act through G protein–coupled receptors that couple with Gαq (GqPCRs), which canonically activate phospholipase Cβ (PLCβ), generating the second messengers inositol 1,4,5-trisphosphate (IP3) and diacylglycerol , which then act to mobilize intracellular calcium and activate numerous downstream kinases, respectively. Chronic GqPCR activation results in cardiac hypertrophy and subsequently leads to cardiomyopathy with depressed contractile function.2,3 Although a low level of Gαq overexpression in mice does not significantly affect the heart, mild Gαq overexpression results in increased hypertrophic markers with minimal cardiac dysfunction6,7 and high Gαq overexpression results in marked hypertrophy, HF, and increased mortality.6 Further, Gαq-overexpressing hearts that are exposed to pressure overload show increased apoptosis,8 whereas overexpression of an inhibitory peptide that interferes with Gαq coupling prevents the development of cardiac hypertrophy and dysfunction in pressure-overloaded mice.9,10 Altogether, these data have highlighted the critical role of Gαq-signaling pathways in mediating the hypertrophic signaling and the transition to HF. Several studies have investigated the potential contributions of cardiomyocyte autophagy on protein turnover and cardiac hypertrophy. Autophagy is an evolutionally conserved mechanism for the degradation of cellular components and organelles by lysosomes. Autophagy also plays an important role in the maintenance of cellular energetics by recycling amino acids and fatty acids for ATP production.11 In eukaryotic cells, autophagy occurs constitutively at low levels to facilitate homeostatic functions such as protein and organelle turnover. However, autophagy is rapidly upregulated in conditions requiring the generation of high intracellular nutrients and energy, such as during starvation, growth factor withdrawal, or removal of damaging cytoplasmic components.11,12 Consistent with these functions, autophagy appears to play a protective role during normal conditions and in response to mild stress.13,14 However, excessive autophagy has been associated to cardiomyocyte death and the development of HF.13,14 Autophagy was observed in failing human hearts caused by dilated cardiomyopathy,15 by valvular disease,16 and by ischemic heart disease.17 Moreover, analysis of hearts from patients with end-stage HF indicated that cardiomyocytes died by a variety of mechanisms including necrosis, apoptosis, and autophagy.18 However, despite considerable evidence linking autophagic activity to HF progression, it is still debated whether autophagy is an adaptive response to protect cardiomyocyte from stress conditions or a maladaptive response that causes HF. In the current issue of the journal, Liu et al19 provide the first analysis of the role of Gαq activation on autophagy in the heart. Using a cardiomyocyte-specific transgenic mouse model of inducible GαqQ209L, a constitutive Gαq mutant deficient in GTP hydrolysis,7,20 the authors demonstrate that autophagic vacuoles, levels of proteins involved in autophagy, and autophagic activity were enhanced in GαqQ209L hearts at 7 days after transgene induction by tamoxifen treatment. GαqQ209L hearts also exhibit enhanced lysosomal degradation activity and elevated levels of the autophagy-initiation complex containing the class III phosphoinositide 3-kinase Vps34, which accounts for the higher abundance of autophagic vacuoles. This increase in autophagy in GαqQ209L mice was associated with a significant decrease in cardiac contractile function at 7 days after transgene induction. The study not only contributes to our understanding of the role of Gαq on autophagy in the heart, but also opens new directions by raising important questions as outlined below. How Does Gαq Activation Regulate Autophagy in the Heart? The study by Liu et al19 demonstrates that expression of GαqQ209L in the heart leads to increased assembly of the autophagy-initiation Beclin1/Vps34/Atg14 complex, Vps34 activity, and production of Phosphatidylinositol 3-phosphate(3)P, culminating in autophagosome formation. As discussed above, Gαq activation leads to stimulation of PLCβ to generate IP3 and diacylglycerol production, but has also been shown to activate the class 1A phosphoinositide 3-kinase PI3Kα, which is inhibited by binding to GαqQ209L.21 Thus, to determine which of these pathways contribute to the increase in autophagy in GαqQ209L hearts, Liu et al used a second cardiomyocyte-specific transgenic mouse line (GαqQ209L-AA) that expresses a tamoxifen-regulated GαqQ209L mutant in which Arg256 and Thr257 are changed to Ala. This protein cannot activate PLCβ but still inhibits PI3Kα.20,22 In contrast to GαqQ209L mice, the GαqQ209L-AA mice did not develop a contractility defect after 7 days of tamoxifen treatment and there was no observable difference in the levels of p62, LC3-II, Vps34, Beclin1, Atg7, or Atg14 between wild-type and GαqQ209L-AA mice, suggesting that enhanced autophagy requires Gαq activation of PLCβ.19 However, investigation of canonical Gαq signaling pathway molecules have reported mixed effects on autophagy. Knockdown of Gαq/11 in HeLa cells using siRNAs have been associated with increased autophagy,23 whereas elevation of intracellular IP3 in cardiac and noncardiac cells decreased autophagy independently of mammalian target of rapamycin complex regulation.24 The mechanisms by which IP3 reduced autophagy are still debated with studies suggesting that elevated IP3 receptor levels inhibit autophagy by either sequestering Beclin 1 away from other autophagic machinery,25 or by increasing intracellular Ca2+ and activating calcium-dependent cysteine proteases (calpains).26 The increase in Ca2+ release by IP3 receptor activation can also maintain mammalian target of rapamycin complex1 activity and therefore reduce autophagy.27 However, it is noteworthy that elevation of cytosolic Ca2+ can also increase autophagy through the Ca2+-dependent activation of PKCβ, the Erk1/2 or the calcium/calmodulin-dependent protein kinase kinase β which can activate AMP-activated protein kinase and initiate autophagy by multiple mechanisms.28 The role of other signaling pathways in mediating Gαq-induced autophagy cannot be ruled out as Gαq may also induce mitochondrial oxidative stress,29 which is closely related to mitochondrial dysfunction and autophagy.30 Therefore, more detailed mechanistic information is needed to dissect the role of Gαq-induced PLCβ signaling on autophagy, and specifically in the heart. What Is the Impact of Gαq Activity on Autophagy-Induced HF Progression? The work by Liu et al19 provides new insight into the role of Gαq-mediated autophagy in the heart. However, whether autophagy is activated as part of a compensatory pathway to protect the heart against Gαq-induced cardiac decompensation or actually contributes to Gαq protein-mediated HF remains to be resolved. Liu et al performed their experiments on mice injected with tamoxifen for 7 days, a timepoint at which the heart has been shown to display a mild contractile defect and abnormal sarcomere structure in cardiomyocytes,19,20 but they did not assess whether the autophagy markers are still elevated in GαqQ209L hearts after 14–28 days post-tamoxifen treatment, when pronounced cardiac contractile dysfunction and transition to HF would be detected.19 Previous studies delineated sequential and ordered changes in autophagy and mitochondrial autophagy that occurred at different phases of development of hypertrophy and cardiac dysfunction, wherein mixed results were observed depending on the duration and the severity of the stress stimuli.31–33 Autophagic activity was reduced during compensatory hypertrophic response induced by moderate thoracic transverse aortic constriction (TAC) at 1 week postsurgery in wild-type mice.33 However, autophagy was upregulated at 4 weeks post-TAC when left ventricular dilation and cardiac dysfunction developed during the transition from hypertrophy to HF.33 In contrast, autophagic activity increases from 24 hours after severe TAC and persists for at least 2 weeks in wild-type or autophagy reporter mice.32 This load-induced pathologic remodeling was significantly reduced in beclin 1 haploinsufficient mice, although compensatory ventricular growth was not altered.32 Conversely, cardiac-specific Beclin 1 transgenic mice exhibit cardiac dilatation and dysfunction 3 weeks after moderate TAC. These findings suggest that autophagy may increase in hypertrophied hearts under conditions of severe stress, such as occurs with Gαq overexpression, and that an excessive increase in autophagy may play a detrimental role in the transition from cardiac hypertrophy to HF. Does Gαq-Induced Autophagy Offer a Potential Target for Treatment of Chronic HF? Although the study by Liu et al19 suggests that Gαq-mediated autophagic flux may contribute to HF, caution is in order when considering translational implications. The view that Gαq-coupled receptor signaling is toxic is based in large part on a transgenic mouse model with Gαq overexpression that exceeds the 2-fold increase found in human HF34–36 and thus cannot be considered to simulate human pathophysiology. In human HF, the maximal increase in Gαq abundance is 2-fold,35 and transgenic mice with 2-fold cardiomyocyte-specific Gαq overexpression have no discernible cardiac phenotype.34,36 Moreover, the study of Liu et al19 used transgenic mice that express recombinant (GαqQ209L-hbER) fusion proteins to activate Gαq signaling selectively in cardiac myocytes. This inducible model, in contrast to the standard transgenic models, was found to cause a dilated cardiomyopathy and HF that does not appear to progress through a hypertrophic stage.20 These discrepancies in Gαq-induced hypertrophy and failure could be related to the transgene being expressed (wild-type Gαq vs. GαqQ209L mutant), their levels of expression or to the overexpression approaches that, even in transgenic mice with tissue-specific promoters do not provide an ideal means of annotating specificity compared with loss-of-function experiments. We also lack a clear understanding of the role of autophagy in noncardiac cells during Gαq-induced cardiac hypertrophy and failure, and how inhibition of autophagy in the failing heart could affect the overall role of Gαq-dependent autophagy in maintaining cellular homeostasis during cardiac remodeling. Therefore understanding the characteristics of autophagy and the effects of Gαq signaling on its induction is important to translate this body of evidence derived from mouse models into pathological conditions.