Nuclear factor-κB regulates expression of platelet phospholipase C-β2 (PLCB2).
Nuclear factor-κB regulates expression of platelet phospholipase C-β2 (PLCB2).
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DOI：
10.1160/th15-09-0749
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发表时间：
2016-10-28
影响因子：
6.7
通讯作者：
Rao AK
中科院分区：
文献类型：
--
作者：
Mao G;Jin J;Kunapuli SP;Rao AK
磷脂酶C(PLC)-PLCB2(基因β2)是血小板活化反应的重要调节因子。PLC-β-2在血小板/单核细胞中表达的调控机制尚不清楚。我们对一例血小板PLC-β2缺乏症患者的研究发现，PLCB2编码序列正常，并降低了血小板PLC-β2mR NA，提示转录调控缺陷。−基因5‘-上游区含有13bp的杂合性缺失(−1645/PLCB1633bp)，包含核因子-κB(NF-κB)的共同序列。随后，在50名健康受试者中有3人检测到了这一点。为了了解PLC-β2的调控机制，我们研究了这种变异对PLCB2的影响。用人红白血病细胞核提取液或重组p65的凝胶位移研究表明，NF-κB与带有NF-κB位点的寡核苷酸结合；在荧光素酶研究中，它的缺失降低了PLCB2启动子的活性。核转录因子κB p65亚基的siRNA下调后，PLCB2的表达降低，而p65过表达则使其表达增加。免疫印迹法检测17例健康受试者血小板PLC-β2与p65的相关性(r=0.76，p=0.0005)。这些研究首次证明了NF-κB调节MK/血小板PLC-β-2的表达。这种相互作用很重要，因为PLC-β2在血小板激活中起主要作用，而核因子-κB在包括炎症和动脉粥样硬化在内的过程中发挥主要作用，两者都密切相关。
Phospholipase C (PLC)-β2 (gene PLCB2) is a critical regulator of platelet responses upon activation. Mechanisms regulating of PLC-β2 expression in platelets/MKs are unknown. Our studies in a patient with platelet PLC-β2 deficiency revealed the PLCB2 coding sequence to be normal and decreased platelet PLC-β2 mRNA, suggesting a defect in transcriptional regulation. PLCB2 5’- upstream region of the patient revealed a heterozygous 13 bp deletion (−1645/−1633 bp) encompassing a consensus sequence for nuclear factor-κB (NF-κB). This was subsequently detected in 3 of 50 healthy subjects. To understand the mechanisms regulating PLC-β2 we studied the effect of this variation in the PLCB2. Gel-shift studies using nuclear extracts from human erythroleukemia (HEL) cells or recombinant p65 showed NF-κB binding to oligonucleotide with NF-κB site; in luciferase reporter studies its deletion reduced PLCB2 promoter activity. PLCB2 expression was decreased by siRNA knockdown of NF-κB p65 subunit and increased by p65 overexpression. By immunoblotting platelet PLC-β2 in 17 healthy subjects correlated with p65 (r=0.76, p=0.0005). These studies provide the first evidence that NF-κB regulates MK/platelet PLC-β2 expression. This interaction is important because of the major role of PLC-β2 in platelet activation and of NF-κB in processes, including inflammation and atherosclerosis, where both are intimately involved.