Dominant negative mutants implicate STAT5 in myeloid cell proliferation and neutrophil differentiation
Dominant negative mutants implicate STAT5 in myeloid cell proliferation and neutrophil differentiation
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DOI：
10.1182/blood.v93.12.4154.412k15_4154_4166
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发表时间：
1999-06-15
期刊：
影响因子：
20.3
通讯作者：
Van Etten, RA
中科院分区：
文献类型：
--
作者：
Ilaria, RL;Hawley, RG;Van Etten, RA
STAT 5是信号转导和转录激活（STAT）家族的一员，在多种细胞因子信号传导途径中被激活的潜在转录因子。我们在小鼠STAT 5A的高度保守区域中引入丙氨酸取代突变，并研究了突变体的二聚化、DNA结合、反式激活和对促红细胞生成素诱导的STAT 5依赖性转录激活的显性负效应。这些突变包括两个靠近氨基末端的突变（W255 KR-->AAA和R（290）QQ-->AAA），两个在DNA结合结构域（E437 E-->AA和V466 VV-->AAA），以及一个类似于大鼠STAT 5 B天然亚型的STAT 5A羧基末端截短（STAT 5A/Delta 53 C）。除了WKR突变体外，所有STAT突变体蛋白均被JAK 2酪氨酸磷酸化，并与STAT 5 B异源二聚化，表明该区域在STAT 5中稳定二聚化的重要作用。WKR，EE，和VVV突变体没有检测到的DNA结合活性，和WKR和VVV突变体，但没有EE，在转录诱导缺陷。VVV突变体对促红细胞生成素诱导的STAT 5转录激活有中度显性负效应，这可能是由于DNA结合缺陷的异二聚体的形成。有趣的是，WKR突变体具有强有力的显性负效应，与反式激活结构域缺失突变体Delta 53 C相当。在鼠髓系精氨酸依赖性细胞系32 D中稳定表达WKR或Delta 53 C STAT 5突变体可抑制白细胞介素-3依赖性增殖和粒细胞集落刺激因子（G-CSF）依赖性分化，而不诱导凋亡。这些突变体在原代小鼠骨髓中的表达抑制体外G-CSF依赖性粒细胞集落形成。这些结果表明，STAT 5不同区域的突变可能通过不同的机制对细胞因子信号传导产生显性负效应，并表明STAT 5在骨髓细胞增殖和分化中的作用。(C)1999年，美国血液学会。
STAT5 is a member of the signal transducers and activation of transcription (STAT) family of latent transcription factors activated in a variety of cytokine signaling pathways. We introduced alanine substitution mutations in highly conserved regions of murine STAT5A and studied the mutants for dimerization, DNA binding, transactivation, and dominant negative effects on erythropoietin-induced STAT5-dependent transcriptional activation. The mutations included two near the amino-terminus (W255KR-->AAA and R(290)QQ-->AAA), two in the DNA-binding domain (E437E-->AA and V466VV-->AAA), and a carboxy-terminal truncation of STAT5A (STAT5A/Delta 53C) analogous to a naturally occurring isoform of rat STAT5B. All of the STAT mutant proteins were tyrosine phosphorylated by JAK2 and heterodimerized with STAT5B except for the WKR mutant, suggesting an important role for this region in STAT5 for stabilizing dimerization. The WKR, EE, and VVV mutants had no detectable DNA-binding activity, and the WKR and VVV mutants, but not EE, were defective in transcriptional induction. The VVV mutant had a moderate dominant negative effect on erythropoietin-induced STAT5 transcriptional activation, which was likely due to the formation of heterodimers that are defective in DNA binding. Interestingly, the WKR mutant had a potent dominant negative effect, comparable to the transactivation domain deletion mutant, Delta 53C. Stable expression of either the WKR or Delta 53C STAT5 mutants in the murine myeloid cytokine-dependent cell line 32D inhibited both interleukin-3-dependent proliferation and granulocyte colony-stimulating factor (G-CSF)-dependent differentiation, without induction of apoptosis. Expression of these mutants in primary murine bone marrow inhibited G-CSF-dependent granulocyte colony formation in vitro. These results demonstrate that mutations in distinct regions of STAT5 exert dominant negative effects on cytokine signaling, likely through different mechanisms, and suggest a role for STAT5 in proliferation and differentiation of myeloid cells. (C) 1999 by The American Society of Hematology.