Phospholipase C-delta gene of the spontaneously hypertensive rat harbors point mutations causing amino acid substitutions in a catalytic domain.
Phospholipase C-delta gene of the spontaneously hypertensive rat harbors point mutations causing amino acid substitutions in a catalytic domain.
复制标题
自发性高血压大鼠的磷脂酶 C-delta 基因存在点突变，导致催化结构域中的氨基酸取代。
DOI：
10.1097/00004872-199111000-00004
复制
发表时间：
1991
影响因子：
4.9
通讯作者：
H. Nojima
中科院分区：
文献类型：
--
作者：
H. Yagisawa;H. Tanase;H. Nojima
本研究旨在从DNA序列水平上探讨新近发现的自发性高血压大鼠（SHR）特异性限制性片段长度多态性（RFLP）在（多聚）磷酸肌醇特异性磷脂酶C（PLC）-δ基因组位点的分布。本研究通过制备SHR主动脉cDNA文库、杂交克隆PLC-δ cDNA和聚合酶链反应分析基因组DNA，克隆了SHR PLC-δ cDNA，并对两个高血压大鼠品系SHR及其卒中易感亚系（SHR-SP）和三个正常血压大鼠品系Sprague-Dawley、Donryu和Wistar-Kyoto（WKY）的基因组DNA进行了分析。用限制性内切酶XhoI酶切后，发现一条仅在SHR和SHR-SP中出现的RFLP带，而在Sprague-Dawley、Donryu和WKY大鼠中没有。从SHR主动脉cDNA文库中克隆的PLC-δ cDNA的DNA测序揭示了总共三个SHR特异性点突变，其中两个导致氨基酸取代。在XhoI位点检测到第一个点突变（A至T），将苏氨酸（ACG）变为丝氨酸（TCG），在第一个点突变附近发现第二个点突变（A至G），将异亮氨酸（ATA）变为甲硫氨酸（ATG）。这是首次证明SHR基因组中的突变改变了氨基酸序列。这些位于PLC-δ的推定催化X结构域的氨基酸取代可能是在SHR中观察到的PLC活性增强的主要原因，可能导致高血压相关的表型瘤，例如异常的钙稳态和细胞内钙离子浓度增加。
This study was undertaken in order to investigate the newly discovered spontaneously hypertensive rat (SHR)-specific restriction fragment length polymorphism (RFLP) at the genomic locus of (poly)phosphoinositide-specific phospholipase C (PLC)-delta at a DNA sequence level. Our aim was to clone the PLC-delta complimentary DNA (cDNA) from SHR and analyse the genomic DNA obtained from two hypertensive rat strains such as SHR and its stroke-prone substrain (SHR-SP) and three normotensive rat strains such as Sprague-Dawley, Donryu and Wistar-Kyoto (WKY) by preparing an aortic cDNA library of SHR, hybridization cloning of PLC-delta cDNA and an analysis of the genomic DNA by polymerase chain reaction. By digesting with restriction enzyme XhoI, we discovered an RFLP band displaying only in SHR and SHR-SP, not in Sprague-Dawley, Donryu and WKY rats. DNA sequencing of PLC-delta cDNA cloned from an aortic cDNA library of SHR revealed a total of three SHR-specific point mutations, two of which resulted in amino acid substitutions. The first point mutation (A to T) was detected at the XhoI site, changing a threonine(ACG) to a serine(TCG), and the second point mutation (A to G) was discovered in the vicinity of the first one, changing an isoleucine(ATA) to a methionine(ATG). This is the first demonstration of the mutations in the SHR genome changing amino acid sequences. These amino acid substitutions, situated in the putative catalytic X domain of PLC-delta, may be the major cause of the augmented PLC activity observed in the SHR, possibly leading to hypertension-related phenonemoma such as abnormal calcium homeostasis and increased intracellular calcium ion concentrations.