Generation of induced cardiomyocytes (iCMs) directly from fibroblasts offers a great opportunity for cardiac disease modeling and cardiac regeneration. A major challenge of iCM generation is the low conversion rate. To address this issue, we attempted to identify small molecules that could potentiate the reprogramming ability towards cardiac fate by removing inhibitory roadblocks. Using mouse embryonic fibroblasts as the starting cell source, we first screened 47 cardiac development related epigenetic and transcription factors, and identified an unexpected role of H3K4 methyltransferase Mll1 and related factor Men1 in inhibiting iCM reprogramming. We then applied small molecules (MM408 and MI503) of Mll1 pathway inhibitors and observed an improved efficiency in converting embryonic fibroblasts and cardiac fibroblasts into functional cardiomyocyte-like cells. We further observed that these inhibitors directly suppressed the expression of Mll1 target gene Ebf1 involved in adipocyte differentiation. Consequently, Mll1 inhibition significantly decreased the formation of adipocytes during iCM induction. Therefore, Mll1 inhibitors likely increased iCM efficiency by suppressing alternative lineage gene expression. Our studies show that targeting Mll1 dependent H3K4 methyltransferase activity provides specificity in the process of cardiac reprogramming. These findings shed new light on the molecular mechanisms underlying cardiac conversion of fibroblasts and provide novel targets and small molecules to improve iCM reprogramming for clinical applications.

中文翻译：

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靶向Mll1 H3K4甲基转移酶活性，以指导成纤维细胞心脏谱系特异性重编程。

直接从成纤维细胞中产生诱导型心肌细胞（iCM），为心脏疾病建模和心脏再生提供了巨大的机会。iCM产生的主要挑战是低转换率。为了解决这个问题，我们试图鉴定出一些小分子，这些小分子可以通过消除抑制性障碍来增强针对心脏命运的重编程能力。使用小鼠胚胎成纤维细胞作为起始细胞来源，我们首先筛选了47种与心脏发育相关的表观遗传和转录因子，并确定了H3K4甲基转移酶Mll1和相关因子Men1在抑制iCM重编程中的意外作用。然后，我们应用了Mll1途径抑制剂的小分子（MM408和MI503），并观察到将胚胎成纤维细胞和心脏成纤维细胞转化为功能性心肌样细胞的效率提高。我们进一步观察到这些抑制剂直接抑制了参与脂肪细胞分化的Mll1靶基因Ebf1的表达。因此，Mll1抑制显着减少了iCM诱导过程中脂肪细胞的形成。因此，Mll1抑制剂可能通过抑制替代谱系基因表达来提高iCM效率。我们的研究表明，靶向Mll1依赖的H3K4甲基转移酶活性在心脏重编程过程中提供了特异性。