JOSD2 promotes cardiomyocyte apoptosis and exacerbates cardiac injury after myocardial infarction via LKB1-AMPK suppression

Abstract

Aim: This study aimed to identify key ubiquitin-regulating molecules involved in cardiomyocyte survival under ischemic injury and to investigate the role of Josephin domain-containing protein 2 (JOSD2) in ischemic stress-induced cardiomyocyte death after myocardial infarction (MI).

Methods: A ubiquitin-related single-guide RNA (sgRNA) library targeting 903 genes was screened in AC16 cells under oxygen-glucose deprivation (OGD), followed by sgRNA sequencing. Integrated analysis of multiple transcriptomic datasets was performed to identify JOSD2 as a candidate. Stable JOSD2 knockdown and overexpression cell models were established. Reverse transcription quantitative polymerase chain reaction (RT-qPCR), Western blot, flow cytometry, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining, and Cell Counting Kit-8 (CCK-8) assays were performed to evaluate OGD-induced cardiomyocyte apoptosis and survival. Transcriptomic analysis, co-immunoprecipitation (co-IP), and deubiquitination assays were performed to explore the underlying mechanism. Adeno-associated virus 9 (AAV9)-mediated cardiomyocyte-specific JOSD2-overexpressing mice were used to evaluate cardiac remodeling and function after MI.

Results: The CRISPR-Cas9 screen identified a series of ubiquitin-regulating candidate genes associated with cardiomyocyte survival under ischemic stress. Combined transcriptomic and experimental analyses showed that JOSD2 was significantly upregulated in cardiomyocytes and mouse hearts after ischemic injury. JOSD2 knockdown attenuated OGD-induced cardiomyocyte apoptosis, whereas JOSD2 overexpression increased apoptosis. Mechanistically, JOSD2 reduced K27-, K29-, K33-, and K63-linked polyubiquitin chains on liver kinase B1 (LKB1), suppressed its activity, and inhibited downstream AMPK (AMP-activated protein kinase) phosphorylation. In vivo, cardiomyocyte-specific JOSD2 overexpression promoted cardiomyocyte death and aggravated cardiac remodeling and dysfunction after MI.

Conclusion: JOSD2 is upregulated in cardiomyocytes after MI and may promote post-infarction cell death and cardiac remodeling by suppressing the LKB1-AMPK pathway. Targeting JOSD2 may represent a potential therapeutic strategy for MI.

Keywords

JOSD2 / myocardial infarction / cardiomyocyte apoptosis / LKB1-AMPK pathway / deubiquitination

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