Regulation of Nucleocytoplasmic Shuttling of Bruton's Tyrosine Kinase (Btk) through a Novel SH3-Dependent Interaction with Ankyrin Repeat Domain 54 (ANKRD54)
Regulation of Nucleocytoplasmic Shuttling of Bruton's Tyrosine Kinase (Btk) through a Novel SH3-Dependent Interaction with Ankyrin Repeat Domain 54 (ANKRD54)
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DOI：
10.1128/mcb.06620-11
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发表时间：
2012-07-01
影响因子：
5.3
通讯作者：
Nore, Beston F.
中科院分区：
文献类型：
--
作者：
Gustafsson, Manuela O.;Hussain, Alamdar;Nore, Beston F.
Bruton‘s酪氨酸激酶(BTK)属于酪氨酸激酶(TFK)家族，对B淋巴细胞的发育起重要作用。BTK信号的取消会导致人类X连锁无丙种球蛋白血症(XLA)和小鼠X连锁免疫缺陷(XID)。我们采用亲和纯化Flag标记的BTK结合串联质谱学的方法来捕获和鉴定新的相互作用蛋白。在这里，我们研究了与Ankryin重复结构域54蛋白(ANKRD54)的相互作用，ANKRD54也被称为Lyn相互作用Ankyrin重复蛋白(LIAR)。虽然BTK是一种核质蛋白，但人们发现说谎者池的穿梭速度比BTK更快。重要的是，我们的结果表明，Liar介导了BTK和另一种TFK，TXK/Rlk的核输出。Liar介导的BTK穿梭富含激活环，非磷酸化的BTK，完全依赖于BTK的SH3结构域。Liar还显示出与天冬氨酸拟磷酸突变体SH3的结合减少。另外三个被研究的核定位蛋白Abl、雌激素受体β(ERβ)和转录因子T-bet都不受Liar的影响。我们将相互作用位点映射到BTK SH3结构域的C末端。生物素化的合成BTK多肽ARDKNGQEGYIPSNYVTEAEDS足以进行这种相互作用。Liar是第一个被发现专门影响BTK和TXK核质穿梭的蛋白质，属于一组罕见的已知蛋白质，以CRM1依赖的方式执行这一活动。
Bruton's tyrosine kinase (Btk), belonging to the Tec family of tyrosine kinases (TFKs), is essential for B-lymphocyte development. Abrogation of Btk signaling causes human X-linked agammaglobulinemia (XLA) and murine X-linked immunodeficiency (Xid). We employed affinity purification of Flag-tagged Btk, combined with tandem mass spectrometry, to capture and identify novel interacting proteins. We here characterize the interaction with ankryin repeat domain 54 protein (ANKRD54), also known as Lyn-interacting ankyrin repeat protein (Liar). While Btk is a nucleocytoplasmic protein, the Liar pool was found to shuttle at a higher rate than Btk. Importantly, our results suggest that Liar mediates nuclear export of both Btk and another TFK, Txk/Rlk. Liar-mediated Btk shuttling was enriched for activation loop, nonphosphorylated Btk and entirely dependent on Btk's SH3 domain. Liar also showed reduced binding to an aspartic acid phosphomimetic SH3 mutant. Three other investigated nucleus-located proteins, Abl, estrogen receptor beta (ER beta), and transcription factor T-bet, were all unaffected by Liar. We mapped the interaction site to the C terminus of the Btk SH3 domain. A biotinylated, synthetic Btk peptide, ARDKNGQEGYIPSNYVTEAEDS, was sufficient for this interaction. Liar is the first protein identified that specifically influences the nucleocytoplasmic shuttling of Btk and Txk and belongs to a rare group of known proteins carrying out this activity in a Crm1-dependent manner.