Glyceraldehyde-3-Phosphate Dehydrogenase Interacts with Proapoptotic Kinase Mst1 to Promote Cardiomyocyte Apoptosis

Bei You,Shengdong Huang,Bing Yi,Zhiyun Xu,Yang Yuan,Qing Qin,Jianxin Sun,Anindita Das

doi:10.1371/journal.pone.0058697

Abstract

Mammalian sterile 20-like kinase 1 (Mst1) is a critical component of the Hippo signaling pathway, which regulates a variety of biological processes ranging from cell contact inhibition, organ size control, apoptosis and tumor suppression in mammals. Mst1 plays essential roles in the heart disease since its activation causes cardiomyocyte apoptosis and dilated cardiomyopathy. However, the mechanism underlying Mst1 activation in the heart remains unknown. In a yeast two-hybrid screen of a human heart cDNA library with Mst1 as bait, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was identified as an Mst1-interacting protein. The interaction of GAPDH with Mst1 was confirmed by co-immunoprecipitation in both co-transfected HEK293 cells and mouse heart homogenates, in which GAPDH interacted with the kinase domain of Mst1, whereas the C-terminal catalytic domain of GAPDH mediated its interaction with Mst1. Moreover, interaction of Mst1 with GAPDH caused a robust phosphorylation of GAPDH and markedly increased the Mst1 activity in cells. Chelerythrine, a potent inducer of apoptosis, substantially increased the nuclear translocation and interaction of GAPDH and Mst1 in cardiomyocytes. Overexpression of GAPDH significantly augmented the Mst1 mediated apoptosis, whereas knockdown of GAPDH markedly attenuated the Mst1 activation and cardiomyocyte apoptosis in response to either chelerythrine or hypoxia/reoxygenation. These findings reveal a novel function of GAPDH in Mst1 activation and cardiomyocyte apoptosis and suggest that disruption of GAPDH interaction with Mst1 may prevent apoptosis related heart diseases such as heart failure and ischemic heart disease.

Highlights

Mammalian sterile 20—like kinase 1 (Mst1) is an ubiquitously expressed serine/threonine kinase with a similarity to the Hippo kinase from Drosophila and it is a critical component of the Hippo signaling pathway, which regulates a variety of biological processes ranging from cell contact inhibition, cell growth, organ size control, apoptosis and tumor suppression in mammals [1,2]
Hybrid screening was performed with human dominant negative Mst1 (DN-Mst1) as bait in conjunction with a human heart cDNA library
The interaction of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) with wild-type Mst1 was further supported by coimmunoprecipitation studies showing that in both cotransfected HEK293T cells and cardiomyocytes, GAPDH interacts with Mst1

Summary

Introduction

Mammalian sterile 20—like kinase 1 (Mst1) is an ubiquitously expressed serine/threonine kinase with a similarity to the Hippo kinase from Drosophila and it is a critical component of the Hippo signaling pathway, which regulates a variety of biological processes ranging from cell contact inhibition, cell growth, organ size control, apoptosis and tumor suppression in mammals [1,2]. MST1 has been shown to phosphorylate FOXO and promote FOXO nuclear translocation, thereby inducing apoptosis in neuronal cells [7,8]. Several phosphorylation sites have been identified in Mst, namely Thr175, Thr177, Thr183, Thr187, Ser327 and Thr387, of which, Thr183 and Thr187 appear to be essential for kinase activation [14,15,16]. Protein-protein interactions have been shown to play critical roles in the regulation of Mst activity. Our recent results demonstrated that Hsp decreases Mst activity through promoting Mst degradation via a CHIP dependent pathway, thereby preventing cancer cells from cisplatin induced apoptosis [21]

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