Center for Cardiovascular Research, John A. Burns School of Medicine, University of Hawaii , Honolulu, Hawaii.
Bioinformatics Core, John A. Burns School of Medicine, University of Hawaii , Honolulu, Hawaii.
Physiol Genomics. 2018 Jul 1;50(7):479-494. doi: 10.1152/physiolgenomics.00130.2017. Epub 2018 Apr 13.
Alternative splicing of RNA is an underexplored area of transcriptional response. We expect that early changes in alternatively spliced genes may be important for responses to cardiac injury. Hypoxia inducible factor 1 (HIF1) is a key transcription factor that rapidly responds to loss of oxygen through alteration of metabolism and angiogenesis. The goal of this study was to investigate the transcriptional response after myocardial infarction (MI) and to identify novel, hypoxia-driven changes, including alternative splicing. After ligation of the left anterior descending artery in mice, we observed an abrupt loss of cardiac contractility and upregulation of hypoxic signaling. We then performed RNA sequencing on ischemic heart tissue 1 and 3 days after infarct to assess early transcriptional changes and identified 89 transcripts with altered splicing. Of particular interest was the switch in Pkm isoform expression (pyruvate kinase, muscle). The usually predominant Pkm1 isoform was less abundant in ischemic hearts, while Pkm2 and associated splicing factors (hnRNPA1, hnRNPA2B1, Ptbp1) rapidly increased. Despite increased Pkm2 expression, total pyruvate kinase activity remained reduced in ischemic myocardial tissue. We also demonstrated HIF1 binding to PKM by chromatin immunoprecipitation, indicating a direct role for HIF1 in mediating this isoform switch. Our study provides a new, detailed characterization of the early transcriptome after MI. From this analysis, we identified an HIF1-mediated alternative splicing event in the PKM gene. Pkm1 and Pkm2 play distinct roles in glycolytic metabolism and the upregulation of Pkm2 is likely to have important consequences for ATP synthesis in infarcted cardiac muscle.
RNA 的可变剪接是转录反应中一个尚未充分探索的领域。我们预计，可变剪接基因的早期变化可能对心脏损伤的反应很重要。缺氧诱导因子 1（HIF1）是一种关键的转录因子，通过改变代谢和血管生成，对缺氧迅速做出反应。本研究的目的是研究心肌梗死（MI）后的转录反应，并确定新的、受缺氧驱动的变化，包括可变剪接。在小鼠的左前降支结扎后，我们观察到心脏收缩力突然丧失和缺氧信号的上调。然后，我们在梗塞后 1 天和 3 天对缺血性心脏组织进行 RNA 测序，以评估早期转录变化，并确定 89 个具有可变剪接的转录本。特别有趣的是 Pkm 同工型表达（丙酮酸激酶，肌肉）的转变。通常占主导地位的 Pkm1 同工型在缺血性心脏中含量较少，而 Pkm2 和相关剪接因子（hnRNPA1、hnRNPA2B1、Ptbp1）迅速增加。尽管 Pkm2 表达增加，但缺血性心肌组织中的总丙酮酸激酶活性仍然降低。我们还通过染色质免疫沉淀证明了 HIF1 与 PKM 的结合，表明 HIF1 在介导这种同工型转换中起直接作用。我们的研究为 MI 后早期转录组提供了新的、详细的描述。从这项分析中，我们确定了 PKM 基因中 HIF1 介导的可变剪接事件。Pkm1 和 Pkm2 在糖酵解代谢中发挥不同的作用，上调 Pkm2 可能对梗塞心肌中的 ATP 合成产生重要影响。
