VCPIP1 drives diabetic cardiomyopathy by deubiquitinating AMPKγ1 and preventing AMPKα-γ subunit assembly in cardiomyocytes

Protein ubiquitination modifications contribute to cardiomyocyte homeostasis and pathophysiology in diabetic cardiomyopathy (DCM). Yet the roles of deubiquitinating enzymes (DUBs) in DCM remain poorly defined. This study sought to delineate how valosin-containing protein interacting protein 1 (VCPIP1), a DUB, regulates DCM and to explore the molecular basis involved. We identify that VCPIP1 was significantly elevated in diabetic hearts, and upregulated VCPIP1 was mainly distributed in cardiomyocytes. Knockout of VCPIP1 specifically in cardiomyocytes ameliorated cardiac damage in mouse models of both type 2 and type 1 diabetes. Ubiquitinome and interactome profiling revealed AMPKγ1 as a substrate of VCPIP1 in cardiomyocytes. Mechanistically, VCPIP1 binds the cystathionine-β-synthase 2 domain of AMPKγ1 via its UBX-L domain and then catalyzes the K63-linked deubiquitination of AMPKγ1-K234 site through its catalytic residue C218. This VCPIP1-mediated AMPKγ1 deubiquitination disrupts AMPKα-γ heterodimer integrity, allosterically impairing AMPKα2-LKB1 interaction and limiting LKB1-mediated AMPKαT172 phosphorylation. RNA-seq analysis showed that AMPKγ1-K234 deubiquitination impaired mitochondrial respiration through inactivating AMPKα in cardiomyocytes, while VCPIP1 deficiency reversed hyperglycemia-induced mitochondrial dysfunction. Finally, AMPKγ1-K234R mutant phenocopied VCPIP1-mediated cardiac pathology in db/db mice. In conclusion, our findings unveil the VCPIP1-AMPKγ1 axis as a non-canonical regulatory mechanism for AMPKαT172 phosphorylation in cardiomyocytes, suggesting that inhibition of VCPIP1 represents a novel treatment approach for DCM.