Constitutive activation of STAT5 by a point mutation in the SH2 domain
Constitutive activation of STAT5 by a point mutation in the SH2 domain
复制标题
DOI：
10.1074/jbc.m909771199
复制
发表时间：
2000-08-11
影响因子：
4.8
通讯作者：
Kitamura, T
中科院分区：
文献类型：
--
作者：
Ariyoshi, K;Nosaka, T;Kitamura, T
我们之前通过聚合酶链反应驱动的随机突变和逆转录病毒介导的表达筛选，鉴定出一种具有组成性活性的STAT（信号换能器和转录激活因子）5A，它在DNA结合和转录激活域有两个点突变，被命名为STAT5A1*6， STAT5A1*6表现出明显升高的DNA结合和反激活活性，具有稳定的酪氨酸磷酸化和核积累。并赋予依赖白细胞介素3的Ba/F3细胞自主细胞生长。我们现在报道了另一个组成型活性突变，STAT5A-N642H，它在SH2结构域有一个单点突变（N642H），使用与鉴定STAT5A1*G相同的策略鉴定，STAT5A-N642H在生化和生物学上与STAT5A1*6表现出相同的特性。有趣的是，STAT5A-N642H的突变导致保守的关键组氨酸的恢复，该组氨酸参与了大多数含sh2蛋白中磷酸酪氨酸的结合。在STAT5A- n642h中引入一个额外的突变（Y694F），该突变破坏了STATE二聚化所需的关键酪氨酸694，消除了突变STAT5A- n642h所表现出的所有活性，这表明STAT5A- n642h的活性需要二聚化，就像野生型STAT5A一样。目前的研究结果还表明，不同的突变通过一个共同的机制使STAT5A具有组成性活性。这和生理激活是相似的。
We previously identified a constitutively active form of STAT (signal transducer and activator of transcription) 5A by polymerase chain reaction-driven random mutagenesis followed by retrovirus-mediated expression screening, which had two point mutations in the DNA-binding and transcriptional activation domains, and was designated STAT5A1*6, STAT5A1*6 showed markedly elevated DNA binding and transactivation activities with stable tyrosine phosphorylation and nuclear accumulation, and conferred autonomous cell growth on interleukin 3-dependent Ba/F3 cells. We now report another constitutively active mutant, STAT5A-N642H which has a single point mutation (N642H) in its SH2 domain, identified using the same strategy as that used to identify STAT5A1*G, STAT5A-N642H showed identical properties to those of STAT5A1*6 both biochemically and biologically. Interestingly the mutation in STAT5A-N642H resulted in restoration of the conserved critical histidine which is involved in the binding of phosphotyrosine in the majority of SH2-containing proteins. Introduction of an additional mutation (Y694F) to STAT5A-N642H, which disrupted critical tyrosine 694 required for dimerization of STATE, abolished all the activities manifested by the mutant STAT5A-N642H, which indicates that dimerization is required for the activity of STAT5A-N642H as was the case for the wild-type STAT5A The present findings also show that different mutations rendered STAT5A constitutively active, through a common mechanism, which is similar to that of physiological activation.