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The formation of a complex comprising PLSCR1, NP, and importin α inhibits the incorporation of importin β into the complex.

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posted on 2018-01-19, 19:06 authored by Weiyu Luo, Jie Zhang, Libin Liang, Guangwen Wang, Qibing Li, Pengyang Zhu, Yuan Zhou, Junping Li, Yuhui Zhao, Nan Sun, Shanyu Huang, Chenchen Zhou, Yu Chang, Pengfei Cui, Pucheng Chen, Yongping Jiang, Guohua Deng, Zhigao Bu, Chengjun Li, Li Jiang, Hualan Chen

(A–D) PLSCR1 formed a complex with NP and different members of the importin α family: importin α1 (A), importin α3 (B), importin α5 (C), and importin α7 (D). HEK293T cells were transfected with plasmids expressing V5-WSNNP and Myc-tagged importin α proteins, together with gradual increasing amounts (0–0.6 μg) of Flag-PLSCR1. The cell lysates were immunoprecipitated with a mouse anti-Myc mAb, and the bound proteins were detected by western blotting with a rabbit anti-V5 pAb, a rabbit anti-Flag pAb, or a rabbit anti-Myc pAb to detect NP, PLSCR1, and importin α family members, respectively. (E) Validation of complex formation among NP, PLSCR1, and importin α1 by including MOV10 as a control. HEK293T cells were transfected with plasmids expressing V5-WSNNP and Myc-tagged importin α1, together with Flag-PLSCR1 or Flag-MOV10. The cell lysates were immunoprecipitated with a mouse anti-Myc mAb, and the bound proteins were detected by western blotting with a rabbit anti-V5 pAb, a rabbit anti-Flag pAb, or a rabbit anti-Myc pAb to detect NP, PLSCR1 or MOV10, and importin α1, respectively. (F) Complex formation among PLSCR1, NP, and importin α1 inhibited the incorporation of importin β into the complex. HEK293T cells were transfected with plasmids expressing V5-WSNNP, Myc-importin α1 and importin β, together with Flag-PLSCR1. The cell lysates were immunoprecipitated with a mouse anti-Myc mAb, and the bound proteins were detected by western blotting with rabbit pAb against V5, Myc or the Flag tag, or importin β.

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