Exo70二聚化的关键序列及其在乳腺癌细胞迁移过程中的作用

Abstract

Exo70是胞外分泌复合体Exocyst的关键亚基,可通过参与Exocyst复合体或自身聚合促进细胞迁移。我们在研究中发现,Exo70自身形成二聚体的能力显著强于与Exocyst其它7个亚基的结合,提示Exo70二聚化的重要作用。进一步构建Exo70缺失突变体,并通过一系列免疫共沉淀实验确定了Exo70形成二聚体的关键位置是位于氨基酸31~35与505~509的两段同样的序列SLEKS。进而利用这两个序列的缺失突变体,通过transwell迁移实验,证实这两段SLEKS序列在Exo70二聚化及乳腺癌细胞迁移过程中的关键作用。以上结果为Exo70独立于Exocyst复合体功能之外的新功能研究提供了依据,也为乳腺癌及其转移的治疗提供潜在的靶点。

Exo70,the key subunit of Exocyst protein complex,promotes cell metastasisby self-dimerization or by involving in the Exocyst complex formation.In this research,we found that Exo70 was dramatically more capable of forming self-dimerization than binding to the other subunits of exocyst complex,and aindicative of the important roles of its self-dimerization.Then we constructed a set of truncation mutants of Exo70 in order to determine the core sequence for its dimerization.With co-immun oprecipitation analysis,we identified two identical protein sequences"SLEKS"located at amino acid 31~35 and 505~509 as critical sequencesfor the dimerization of Exo70.By "cell transwell"essay,we further found that the full length Exo70 significantly promotes breast cancer metastasis,while the two truncation mutants of Exo70(Δ31~35 and Δ505~509) don't process such ability,indicating that these two "SLEKS"sequences play pivotal role in breast cancer cell metastasis and invasion.Together,these results provide fundamental basis for researches on novel functions of Exo70 besides its well-known role in Exocyst complex formation and provide potential drug target for treating breast cancer and its metastasis.