Histone methyltransferase KMT2D contributes to the protection of myocardial ischemic injury

Abstract

Histone H3 lysine 4 (H3K4) methyltransferase 2D (KMT2D) plays an important role in cell development in early life. However, the function of KMT2D in adult cells such as cardiomyocytes or neurons has not been reported. In this study, cardiomyocyte-specific KMT2D knockout (KMT2D-cKO) and control (KMT2D-Ctl) mice were exposed to sham or myocardial ischemia (MI) surgery. Depletion of KMT2D aggravated the ischemic area, led to the increased mortality (26.5% in KMT2D-cKO vs 12.5% in KMT2D-Ctl) of the mice, and weakened the left ventricular systolic function. RNA-seq analysis in cardiac tissues identified genes whose expression was changed by MI and KMT2D deletion. Combined with the genome-wide association study (GWAS) analysis, cardiac disease-associated genes Rasd1, Thsd7a, Ednra, and Tns1 were identified. The expression of the Rasd1 was significantly decreased by MI or the loss of KMT2D in vivo. Meanwhile, ChIP assays demonstrated that either MI or loss of KMT2D attenuated monomethylated H3K4 (H3K4me1) enrichment on the enhancer of Rasd1. By generating a KMT2D knockout (H9C2-KO) H9C2 monoclone, we verified that the expression of Rasd1 was controlled by KMT2D, and the expression of Rasd1 was decreased by serum starvation but not low-(O2) treatment in H9C2 cells. KMT2D has a protective effect on ischemic myocardium by regulating cardiac disease-associated genes including Rasd1. KMT2D is required for the H3K4me1 deposition on the enhancer of Rasd1. Our data for the first time suggest that KMT2D-mediated Rasd1 expression may play an important protective effect on adult cells during nutritional deficiency caused by ischemic injury.

Loss of Histone Methyltransferase KMT2D Attenuates Angiogenesis in the Ischemic Heart by Inhibiting the Transcriptional Activation of VEGF-A

“…Suppressed vascular density of Kmt2d cardiomyocyte-specific knockout mice after MI may be attributed to the inhibition of the paracrine effects of cardiomyocytes on ECs mediated by KMT2D deficiency. To this end, Kmt2d gene was deleted in H9c2 cardiomyocytes using the CRISPR/Cas9 gene-editing knockout system as previously reported [ 14 ], after which the CM was collected from the cardiomyocytes cultured for 24 h under normoxia or hypoxia (1% O 2 ) and added into EC culture. After determining that KMT2D protein was significantly knocked down in H9c2 cells (Fig.…”

“…Loss of KMT2D protein significantly reduces the transcription activity of target genes [ 35 ]. Our previous studies demonstrated that cardiomyocyte-specific KMT2D knockout mice exhibited larger infarct size after myocardial infarction surgery with reduced expression of functional genes [ 14 ]. In the current study, we detected impaired angiogenesis and reduced level of H3K4me1 in the Vegf-a promoter region during MI in the cardiomyocyte-specific Kmt2d knockout mice and KMT2D silenced H9c2 cardiomyocyte respectively.…”

“…Left anterior descending coronary artery (LAD) was ligated to create the MI mice model, as we applied before [ 14 ]. All mice were anesthetized in the chamber at 2% isoflurane, under sterile conditions, LAD was tied by a 7–0 silk suture.…”

“…The generation of KMT2D knockout H9c2 cardiomyocytes cell lines based on CRISPR/Cas9 gene-editing knockout system has been reported in our previous study [ 14 ]. Briefly, the oligodeoxynucleotides of gRNAs were annealed and cloned into PX458 plasmids (48138, Addgene), named PX458- Kmt2d -gRNA1 and PX458- Kmt2d -gRNA2, respectively, and confirmed by sequencing.…”

“…Canonically, KMT2D is an essential regulator of the promoter regions or enhancer regions to induce H3K4me1 or H3K4me2 modification, which is involved in regulating chromatin accessibility and transcriptional activation of target genes [ 12 ].Previous studies have shown that KMT2D is essential for heart development [ 13 ]. We recently demonstrated the critical function of KMT2D in the adult MI mouse model [ 14 ]. Our present observations show impaired angiogenesis in the ischemic mouse heart when KMT2D was deleted specifically in the cardiomyocytes.…”