Regulation of platelet myosin light chain (MYL9) by RUNX1: implications for thrombocytopenia and platelet dysfunction in RUNX1 haplodeficiency
Regulation of platelet myosin light chain (MYL9) by RUNX1: implications for thrombocytopenia and platelet dysfunction in RUNX1 haplodeficiency
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DOI：
10.1182/blood-2010-06-289850
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发表时间：
2010-12-23
期刊：
影响因子：
20.3
通讯作者：
Rao, A. Koneti
中科院分区：
文献类型：
--
作者：
Jalagadugula, Gauthami;Mao, Guangfen;Rao, A. Koneti
转录因子 RUNX1 的突变与家族性血小板疾病、血小板减少症和白血病易感性相关。我们描述了一名患有血小板减少症和激动剂诱导的血小板聚集、分泌和糖蛋白 (GP) IIb-IIIa 激活受损的患者，这些患者与 RUNX1 突变相关。血小板肌球蛋白轻链（MLC）磷酸化及其基因 MYL9 的转录水平降低。肌球蛋白 IIA 和 MLC 磷酸化在血小板对激活的反应中很重要，并通过对过早前血小板形成的负调节作用来调节血小板生成。我们提出了 MYL9 是 RUNX1 转录靶标的假设。使用巨核细胞的染色质免疫沉淀 (ChIP) 显示 RUNX1 与 MYL9 启动子区域 -729/-542 碱基对 (bp) 结合，其中包含 4 个 RUNX1 位点。电泳迁移率变动分析显示 RUNX1 与每个位点结合。在瞬时 ChIP 测定中，这些位点的突变消除了 RUNX1 与 MYL9 启动子构建体的结合。在报告基因检测中，每个 RUNX1 位点的缺失都会降低活性。 RUNX1 短干扰 RNA (siRNA) 抑制 MYL9 表达，而 RUNX1 过表达则增强 MYL9 表达。 RUNX1 siRNA 减少了细胞在胶原蛋白和纤维蛋白原上的铺展。我们的结果首次证明 MYL9 基因是 RUNX1 的直接靶标，并提供了与 RUNX1 突变相关的血小板 MYL9 表达减少、MLC 磷酸化、血小板减少和血小板功能障碍的机制。 （血。2010；116（26）：6037-6045）
Mutations in transcription factor RUNX1 are associated with familial platelet disorder, thrombocytopenia, and predisposition to leukemia. We have described a patient with thrombocytopenia and impaired agonist-induced platelet aggregation, secretion, and glycoprotein (GP) IIb-IIIa activation, associated with a RUNX1 mutation. Platelet myosin light chain (MLC) phosphorylation and transcript levels of its gene MYL9 were decreased. Myosin IIA and MLC phosphorylation are important in platelet responses to activation and regulate thrombopoiesis by a negative regulatory effect on premature proplatelet formation. We addressed the hypothesis that MYL9 is a transcriptional target of RUNX1. Chromatin immunoprecipitation (ChIP) using megakaryocytic cells revealed RUNX1 binding to MYL9 promoter region -729/-542 base-pairs (bp), which contains 4 RUNX1 sites. Electrophoretic mobility shift assay showed RUNX1 binding to each site. In transient ChIP assay, mutation of these sites abolished binding of RUNX1 to MYL9 promoter construct. In reporter gene assays, deletion of each RUNX1 site reduced activity. MYL9 expression was inhibited by RUNX1 short interfering RNA (siRNA) and enhanced by RUNX1 overexpression. RUNX1 siRNA decreased cell spreading on collagen and fibrinogen. Our results constitute the first evidence that the MYL9 gene is a direct target of RUNX1 and provide a mechanism for decreased platelet MYL9 expression, MLC phosphorylation, thrombocytopenia, and platelet dysfunction associated with RUNX1 mutations. (Blood. 2010; 116(26):6037-6045)