p35 interacts with alpha-tubulin and organelle proteins: nuclear translocation of p35 in dying cells.
We identified heterogeneous nuclear ribonucleoprotein (hnRNP) C1/C2, hnRNP A1, the translocase of the transporter outer membrane 40 (TOM40), and alpha-tubulin as new interaction partners of anti-apoptotic protein p35 using MS-based functional proteomics with GST-p35 fusion protein as a bait, and using a pull-down assay with p35-6His followed by Western blot analysis. p35 was localized in the cytoplasm and in distinct organelles such as the nucleus and mitochondria. p35 was more abundant in the cytoplasm than it was in the nucleus. It co-localized with alpha-tubulin in the cytoplasm in the absence of a death stimulus. However, while cells were undergoing death induced by actinomycin D, cytoplasmic p35 was translocated into the nucleus; this process was inhibited by deletions of the N- and C-terminal domains containing leucine-rich motifs. Gene delivery of p35 using recombinant adenoviruses inhibited cytoplasmic compartmentalization of hnRNP C1/C2 and hnRNP A1 in dying cells. This study demonstrated translocation of p35 into the nuclei, as well as protection of the hnRNPs from redistribution in cells undergoing death. We propose an active role for p35 in maintaining the integrity of nuclear proteins during cell death.
Son
Yonghae
Y
Department of Pharmacology, School of Medicine, Pusan National University, Yangsan, Kyeongnam 626-770, Republic of Korea.
Kim
Sunmi
S
Choi
Kyungha
K
Park
Youngchul
Y
Eo
Seongkug
S
Kim
Youngkook
Y
Rhim
Byungyong
B
Kim
Koanhoi
K
D020816:Amino Acid Motifs; D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D002846:Chromatography, Affinity; D006224:Cricetinae; D003412:Cricetulus; D003609:Dactinomycin / Q000494:pharmacology; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B / Q000378:metabolism; D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C / Q000378:metabolism; D006801:Humans; D018613:Microscopy, Confocal; D011485:Protein Binding; D021381:Protein Transport / Q000187:drug effects; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014404:Tubulin / Q000378:metabolism*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C; D014404:Tubulin; D014764:Viral Proteins; C081618:p35 protein, Nucleopolyhedrovirus; D003609:Dactinomycin