Investigation of interaction between SPAG6 and TCTE3 proteins

Shuang Chaofan; Liu Yunhao; Liu Junpin; Min Jie; Li Mengying; Gui Yuan; Huang Jinyu; Zhang Ling; Zhang Zhibing

doi:10.13406/j.cnki.cyxb.001194

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Investigation of interaction between SPAG6 and TCTE3 proteins

更新时间：2024-01-17

- Investigation of interaction between SPAG6 and TCTE3 proteins
- Journal of Chongqing Medical University Vol. 42, Issue 8, Pages: 948-952(2017)
- 作者机构：
  
  1. 武汉科技大学医学院公共卫生学院劳动卫生与环境卫生学教研室
  2. 弗吉利亚联邦大学妇产科学系
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.001194
  CLC： R698.2
- Published：2017，
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[1]双超凡,柳赟昊,刘晙玭,闵洁,李梦颖,桂媛,黄金宇,张玲,张志兵.精子相关抗原6(SPAG6)与TCTE3蛋白间相互作用探讨[J].重庆医科大学学报,2017,42(08):948-952.

Shuang Chaofan, Liu Yunhao, Liu Junpin, et al. Investigation of interaction between SPAG6 and TCTE3 proteins. [J]. Journal of Chongqing Medical University 42(8):948-952(2017)
[1]双超凡,柳赟昊,刘晙玭,闵洁,李梦颖,桂媛,黄金宇,张玲,张志兵.精子相关抗原6(SPAG6)与TCTE3蛋白间相互作用探讨[J].重庆医科大学学报,2017,42(08):948-952. DOI： 10.13406/j.cnki.cyxb.001194.

Shuang Chaofan, Liu Yunhao, Liu Junpin, et al. Investigation of interaction between SPAG6 and TCTE3 proteins. [J]. Journal of Chongqing Medical University 42(8):948-952(2017) DOI： 10.13406/j.cnki.cyxb.001194.

摘要

目的:探讨精子相关抗原6（sperm associated antigen 6

Spag6）和T复合体相关睾丸表达3（T-complex associated testis expressed 3

TCTE3）之间的相互作用。方法:PCR法观察Tcte3在雄性小鼠各组织中的表达;以小鼠睾丸c DNA为模板

扩增Tcte3序列

构建Tcte3/p GADT7、Tcte3/p EGFPN2重组质粒;以Tcte3/p EGFPN2转染CHO细胞和COS-1细胞

免疫荧光染色及Western blot检测TCTE3在细胞中的定位和表达;以Spag6/p GBKT7和Tcte3/p GADT7进行酵母双杂交实验

提取酵母蛋白后采用Western blot检测TCTE3与SPAG6之间是否存在直接的相互作用。结果:Tcte3在睾丸、脑、肝、肾、肺、脾、肌肉中均有表达;酶切和测序鉴定证实Tcte3/p GADT7和Tcte3/p EGFPN2重组质粒均构建成功;体外实验发现TCTE3主要在CHO和COS-1细胞质中表达

当与SPAG6质粒共转染后

TCTE3被募集至细胞微管中表达;酵母双杂交实验显示TCTE3/SPAG6组合后

在缺少SD/-Leu/-Trp/-His的培养板上有菌落生成

Western blot显示菌落中TCTE3、SPAG6蛋白表达显著

分子量正确。结论:精子相关抗原SPAG6与TCTE3之间存在直接的相互作用

SPAG6可募集TCTE3至细胞微管中表达。

Abstract

Objective:To test the interactions between SPAG6 and Tcte3 proteins. Methods:The expression of Tcte3 was observed in different tissues of mice by PCR. The testis c DNA was used as the template to amplify Tcte3 coding sequence by PCR

following with the Tcte3/p GADT7 and Tcte3/p EGFPN2 recombinant plasmids’ constructing. Immunofluorescent staining and Western blot were used to detect the location and protein express after transfecting Tcte3/p EGFPN2 into CHO cells and COS-1 cells. The interaction between SPAG6 and TCTE3 was detected by Western blot after yeast two hybrid for Tcte3/p GADT7 and SPAG6/PGBKT7. Results:Tcte3mainly expressed in testis

brain

liver

kidney

lung

spleen and muscle. The results of restriction enzyme digestion and sequencing showed the Tcte3/p GADT7 and Tcte3/p EGFPN2 recombinant plasmids were constructed successfully. TCTE3 mainly located in the whole cytoplasm in vitro

whereas it was recruited to microtubule after co-transfecting with SPAG6. In yeast two hybrid assay

the mating group of TCTE3/SPAG6 grow promoted colonies to grow in SD/-Leu/-Trp/-His plates

and Western blot assay had identified TCTE3 and SPAG6 proteins could express in these colonies. Conclusion:The directly interaction between SPAG6 and TCTE3 is confirmed

and SPAG6 could recruite TCTE3 to celluar microtubule.

关键词

男性不育精子相关抗原6T复合体相关睾丸表达3酵母双杂交蛋白表达

Keywords

male infertilitysperm associated antigen 6T-complex associated testis expressed 3yeast two hybridprotein expression

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