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Phospho-Ser727 triggers a multistep inactivation of STAT3 by rapid dissociation of pY705-SH2 through C-terminal tail modulation.
International Immunology ( IF 4.4 ) Pub Date : 2020-02-07 , DOI: 10.1093/intimm/dxz061
Junhao Yang 1 , Hiroyuki Kunimoto 1 , Bumpei Katayama 2 , Hong Zhao 1 , Takashi Shiromizu 3 , Lingyu Wang 1 , Toshiyuki Ozawa 2 , Takeshi Tomonaga 3 , Daisuke Tsuruta 2 , Koichi Nakajima 1
Affiliation  

Signal transducer and activator of transcription 3 (STAT3) is involved in many biological processes, including immunity and cancer. STAT3 becomes phosphorylated at Tyr705 and Ser727 on IL-6 stimulation. Phospho-Tyr705 (pY705) stabilizes the STAT3 dimer with reciprocal interactions between pY705 and the SH2 of the other molecule and phospho-Ser727 (pS727) accelerates pY705 dephosphorylation. We study how pS727 regulates STAT3 in both structural and biological perspectives. Using STAT3 reconstituted in HepG2-stat3-knockout cells, we show that pS727, together with a handshake N-terminal domain (NTD) interaction, causes rapid inactivation of STAT3 for pY705 dephosphorylation and a chromosome region maintenance 1 (CRM1)-independent nuclear export, which is critical for faithful STAT3 response to the cellular signals. The various N-terminal tags, GFP-related Ruby and FLAG, rendered the export CRM1-dependent and especially FLAG-tag caused nuclear accumulation of STAT3, indicating the presence of conformational changes in inactivation. Impaired reactivation of STAT3 by S727A or FLAG-tag delayed or inhibited the IL-6-induced saa1 mRNA expression, respectively. The detailed analysis of the pY705-SH2 structure identified the C-terminal tail (CTT) from L706 to P715 as a key regulator of the CTT-CTT intermolecular and the CTT-SH2 intramolecular interactions that support pY705-SH2 association. The functional studies using multiple STAT3 mutants indicated that the degree of the two interactions determines the stability of pY705-SH2 interaction. Importantly, Pro715 was critical for the pS727's destabilizing activity and the known phosphorylation and acetylation at the CTT structurally inhibited the pY705-SH2 interaction. Thus, pS727 triggers pY705-SH2 dissociation by weakening the supportive interactions likely through CTT modulation, inducing rapid cycles of STAT3 activation-inactivation for proper function of STAT3.

中文翻译:

Phospho-Ser727通过C端尾部调制快速解离pY705-SH2,从而触发STAT3的多步失活。

信号转导和转录激活因子3(STAT3)参与许多生物过程,包括免疫和癌症。STAT3在IL-6刺激下在Tyr705和Ser727处被磷酸化。Phospho-Tyr705(pY705)通过pY705与其他分子的SH2之间的相互作用而稳定STAT3二聚体,而phospho-Ser727(pS727)则促进pY705的去磷酸化。我们从结构和生物学角度研究pS727如何调节STAT3。使用在HepG2-stat3-敲除细胞中重构的STAT3,我们显示pS727与握手N末端域(NTD)相互作用一起,导致STAT3迅速失活,导致pY705去磷酸化和染色体区域维持1(CRM1)独立的核输出。 ,这对于忠实的STAT3对细胞信号的响应至关重要。各种N端子标签,GFP相关的Ruby和FLAG使出口CRM1依赖,尤其是FLAG标签引起STAT3的核积累,表明灭活中存在构象变化。S727A或FLAG标签对STAT3的再激活受损,分别延迟或抑制了IL-6诱导的saa1 mRNA表达。对pY705-SH2结构的详细分析确定了从L706到P715的C末端尾巴(CTT)是CTT-CTT分子间和支持PY705-SH2缔合的CTT-SH2分子内相互作用的关键调节剂。使用多个STAT3突变体的功能研究表明,两种相互作用的程度决定了pY705-SH2相互作用的稳定性。重要的是,Pro715对于pS727'至关重要 的去稳定活性和CTT处的已知磷酸化和乙酰化在结构上抑制了pY705-SH2相互作用。因此,pS727通过减弱可能通过CTT调节产生的支持性相互作用来触发pY705-SH2解离,从而诱导STAT3激活-失活的快速循环,从而发挥STAT3的正常功能。
更新日期:2019-11-01
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