BACH2 ameliorates cell apoptosis and autophagy as a molecular switch of BCL-2/Beclin-1 complex in myocardial infarction.

Apoptosis and autophagy are the two dominant programmed cell death subtypes occurring in myocardial infarction models.

• Various forms of programmed cell death were detected in myocardial infarction models both in vivo and in vitro.
• Among the PCD subtypes examined, apoptosis and autophagy were identified as the predominant forms in this context.
• Detection methods included western blot, immunofluorescent staining, and other molecular assays across multiple model systems.

BACH2 protein translocates from the nucleus to the cytoplasm and its total protein levels decrease upon hypoxia/reoxygenation stimulation.

• BACH2 expression and cellular localization were monitored using western blot, qRT-PCR, immunofluorescent staining, and isolation of nuclear and cytoplasmic proteins.
• The translocation and protein level changes were characterized in a time- and dose-dependent manner under hypoxia/reoxygenation (H/R) conditions.
• Both the subcellular redistribution and overall reduction in BACH2 protein were observed in response to H/R stimulation.

Cardiac-specific knockout of BACH2 aggravated cardiac dysfunction caused by myocardial infarction, while knockin of BACH2 ameliorated it.

• Cardiac-specific knockout mice were generated by crossing BACH2flox/flox mice with MYH6-Cre mice followed by Tamoxifen injection.
• Cardiac-specific knockin mice were generated by crossing BACH2 Rosa26flox/flox mice with MYH6-Cre mice followed by Tamoxifen injection.
• Functional effects were assessed in both mouse models and cultured cells, confirming consistent results across in vivo and in vitro systems.

BACH2 binds to a specific site on the BCL2 promoter and transcriptionally increases BCL2 mRNA levels, suppressing cardiomyocyte apoptosis.

• Binding of BACH2 to the BCL2 promoter was confirmed using chromatin immunoprecipitation (ChIP) assays.
• Mechanism studies employed immunoprecipitation, site-specific mutation, dual luciferase activity assays, and specific antagonists.
• Increased BCL2 mRNA expression downstream of BACH2 was identified as a mechanism for apoptosis suppression.
• RNA-seq analysis combined with CUT&Tag assays were performed to identify the underlying molecular targets.

BACH2 enhances the formation of the BCL2/Beclin-1 complex, resulting in inhibition of autophagy in cardiomyocytes.

• The interaction between BCL2 and Beclin-1 was examined using immunoprecipitation assays.
• BACH2 promoted formation of the BCL2/Beclin-1 complex as a distinct mechanism separate from its transcriptional regulation of BCL2.
• This dual function positions BACH2 as a molecular switch controlling both apoptosis and autophagy simultaneously.
• Specific antagonists and site-specific mutations were used to confirm the mechanistic relationship.

The natural small compound Myricetin was identified as an efficient activator of BACH2 and protected the heart from myocardial infarction in a BACH2-dependent manner.

• Myricetin was identified as a BACH2 agonist through screening approaches.
• Myricetin was injected into both wild-type and BACH2 conditional knockout mice with myocardial infarction to assess its cardioprotective effects.
• The cardioprotective effect of Myricetin was confirmed to be BACH2-dependent, as protection was lost in BACH2 conditional knockout mice.
• These findings were used to assess the clinical potential of targeting BACH2 with small molecules.