Regulation of the interaction between glycogen synthase kinase 3 and the Kaposi's sarcoma-associated herpesvirus latency-associated nuclear antigen
Regulation of the interaction between glycogen synthase kinase 3 and the Kaposi's sarcoma-associated herpesvirus latency-associated nuclear antigen
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DOI：
10.1128/jvi.79.16.10429-10441.2005
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发表时间：
2005-08-01
影响因子：
5.4
通讯作者：
Hayward, SD
中科院分区：
文献类型：
--
作者：
Fujimuro, M;Liu, JY;Hayward, SD
卡波西肉瘤相关疱疹病毒（KSHV）编码的潜伏相关核抗原（拉娜）蛋白通过与负调节因子糖原合成酶激酶3（GSK-3）结合并消耗细胞质GSK-3水平的新机制稳定β-连环蛋白。进一步表征了与GSK-3相互作用所需的拉娜的两个结构域。使用GSK-3和拉娜突变体获得了C-末端拉娜相互作用结构域和轴蛋白GSK-3相互作用结构域之间相似性的证据。GSK-3（F291 L）不与轴蛋白相互作用，也不能与拉娜结合，并且拉娜的轴蛋白同源结构域中的突变L1132 P破坏了与GSK-3的结合。发现N-末端拉娜相互作用结构域通过充当GSK-3的底物来介导相互作用。GSK-3（R96 A）是一种引发口袋突变体，不与拉娜结合，表明拉娜是引发GSK-3的底物。GSK-3抑制剂LiCl可抑制从原发性渗出性淋巴瘤细胞沉淀的内源性拉娜的磷酸化。在体外，GST-LANA（1340）被GSK-3磷酸化，丝裂原活化蛋白激酶（MAPK）和酪蛋白激酶I起着引发激酶的作用。GSK-3共有位点的突变揭示了拉娜氨基酸219和268之间的位点对于GSK-3磷酸化是重要的。免疫沉淀试验表明，GSK-3磷酸化的损失与GSK-3相互作用的损失相关的N-末端结构域。尽管LANA相关的GSK-3主动磷酸化拉娜，但与拉娜共沉淀的GSK-3不能磷酸化外源肽底物。GSK-3的拉娜隔离可以解释KSHV感染细胞耐受核GSK-3水平增加的能力。
The Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded latency-associated nuclear antigen (LANA) protein stabilizes beta-catenin by the novel mechanism of binding to the negative regulator, glycogen synthase kinase 3 (GSK-3), and depleting cytoplasmic GSK-3 levels. The two domains of LANA required for interaction with GSK-3 were further characterized. Evidence for similarity between the C-terminal LANA interaction domain and the axin GSK-3 interaction domain was obtained using GSK-3 and LANA mutants. GSK-3(F291L), which does not interact with axin, also failed to bind to LANA, and a mutation in the axin homology domain of LANA, L1132P, destroyed binding to GSK-3. The N-terminal LANA interaction domain was found to mediate interaction by acting as a substrate for GSK-3. GSK-3(R96A), a priming pocket mutant, did not bind to LANA, suggesting that LANA was a primed GSK-3 substrate. Phosphorylation of endogenous LANA precipitated from primary effusion lymphoma cells was inhibited by the GSK-3 inhibitor LiCl. GST-LANA(1340) was phosphorylated by GSK-3, and mitogen-activated protein kinase (MAPK) and casein kinase I functioned as priming kinases in vitro. Mutation of consensus GSK-3 sites revealed that sites between LANA amino acids 219 and 268 were important for GSK-3 phosphorylation. Immunoprecipitation assays revealed that loss of GSK-3 phosphorylation of this N-terminal domain correlated with loss of GSK-3 interaction. Although LANA-associated GSK-3 actively phosphorylated LANA, GSK-3 coprecipitated with LANA was unable to phosphorylate an exogenous peptide substrate. LANA sequestration of GSK-3 may explain the ability of KSHV-infected cells to tolerate increased levels of nuclear GSK-3.