Protein kinase D activation induces mitochondrial fragmentation and dysfunction in cardiomyocytes

Abstract

Regulation of mitochondrial morphology is crucial for the maintenance of physiological functions in many cell types including cardiomyocytes. Small and fragmented mitochondria are frequently observed in pathological conditions, but it is still unclear which cardiac signalling pathway is responsible for regulating the abnormal mitochondrial morphology in cardiomyocytes. Here we demonstrate that a downstream kinase of G_q protein-coupled receptor (G_q PCR) signalling, protein kinase D (PKD), mediates pathophysiological modifications in mitochondrial morphology and function, which consequently contribute to the activation of apoptotic signalling. We show that G_q PCR stimulation induced by α₁ -adrenergic stimulation mediates mitochondrial fragmentation in a fission- and PKD-dependent manner in H9c2 cardiac myoblasts and rat neonatal cardiomyocytes. Upon G_q PCR stimulation, PKD translocates from the cytoplasm to the outer mitochondrial membrane (OMM) and phosphorylates a mitochondrial fission protein, dynamin-like protein 1 (DLP1), at S637. PKD-dependent phosphorylation of DLP1 initiates DLP1 association with the OMM, which then enhances mitochondrial fragmentation, mitochondrial superoxide generation, mitochondrial permeability transition pore opening and apoptotic signalling. Finally, we demonstrate that DLP1 phosphorylation at S637 by PKD occurs in vivo using ventricular tissues from transgenic mice with cardiac-specific overexpression of constitutively active Gα_q protein. In conclusion, G_q PCR-PKD signalling induces mitochondrial fragmentation and dysfunction via PKD-dependent DLP1 phosphorylation in cardiomyocytes. This study is the first to identify a novel PKD-specific substrate, DLP1 in mitochondria, as well as the functional role of PKD in cardiac mitochondria. Elucidation of these molecular mechanisms by which PKD-dependent enhanced fission mediates cardiac mitochondrial injury will provide novel insight into the relationship among mitochondrial form, function and G_q PCR signalling.