Reinvestigation of Disulfide-bonded Oligomeric Forms of the Unfolded Protein Response Transducer ATF6

• Koba Hibiki

  Department of Biophysics, Graduate School of Science, Kyoto University

• Jin Shengyu

  Department of Biophysics, Graduate School of Science, Kyoto University

• Imada Nanami

  Department of Biophysics, Graduate School of Science, Kyoto University

• Ishikawa Tokiro

  Department of Biophysics, Graduate School of Science, Kyoto University

• Ninagawa Satoshi

  Department of Biophysics, Graduate School of Science, Kyoto University

• Okada Tetsuya

  Department of Biophysics, Graduate School of Science, Kyoto University

• Sakuma Tetsushi

  Division of Integrated Sciences for Life, Graduate School of Integrated Sciences for Life, Hiroshima University

• Yamamoto Takashi

  Division of Integrated Sciences for Life, Graduate School of Integrated Sciences for Life, Hiroshima University

• Mori Kazutoshi

  Department of Biophysics, Graduate School of Science, Kyoto University

Abstract

ATF6α is an endoplasmic reticulum (ER)-embedded transcription factor which is rapidly activated by ER stress, and a major regulator of ER chaperone levels in vertebrates. We previously suggested that ATF6α occurs as a monomer, dimer and oligomer in the unstressed ER of Chinese hamster ovary cells due to the presence of two evolutionarily conserved cysteine residues in its luminal region (C467 and C618), and showed that ATF6α is reduced upon ER stress, such that only reduced monomer ATF6α is translocated to the Golgi apparatus for activation by proteolysis. However, mutagenesis analysis (C467A and C618A) revealed that the C618A mutant behaves in an unexpected manner (monomer and oligomer) during non-reducing SDS-PAGE, for reasons which remained unclear. Here, we used human colorectal carcinoma-derived HCT116 cells deficient in ATF6α and its relevant ATF6β, and found that ATF6α dimer and oligomer are both dimers, which we designated C618-dimer and C467-dimer, respectively. We demonstrated that C467-dimer (previously considered an oligomer) behaved bigger than C618-dimer (previously considered a dimer) during non-reducing SDS-PAGE, based on their disulfide-bonded structures. Furthermore, ATF6α monomer physically associates with another ATF6α monomer in the absence of disulfide bonding, which renders two C467 residues in close proximity so that formation of C467-dimer is much easier than that of C618-dimer. In contrast, C618-dimer is more easily reduced upon ER stress. Thus, our analysis revealed that all forms of ATF6α, namely monomer, C618-dimer and C467-dimer, are activated by single reduction of a disulfide bond in response to ER stress, ensuring the rapidity of ATF6α activation. Key words: disulfide-bonded structure, endoplasmic reticulum, membrane-bound transcription factor, non-reducing SDS-PAGE, unfolded protein response

Journal

• Cell Structure and Function

  Cell Structure and Function 45 (1), 9-21, 2020

  Japan Society for Cell Biology

Keywords

• disulfide-bonded structure
• endoplasmic reticulum
• membrane-bound transcription factor
• non-reducing SDS-PAGE
• unfolded protein response

Details

• CRID: 1390846609800961024
• NII Article ID: 130007792116
• NII Book ID: AA0060007X
• DOI: 10.1247/csf.19030
• ISSN: 13473700
• ISSN: 03867196
• NDL BIB ID: 031311424
• PubMed: 31852864
• Web Site: https://ndlsearch.ndl.go.jp/books/R000000004-I031311424
• Web Site: https://www.jstage.jst.go.jp/article/csf/45/1/45_19030/_pdf
• Web Site: https://search.jamas.or.jp/link/ui/2022034673
• Text Lang: en
• Data Source:
  • JaLC
  • NDL
  • Crossref
  • PubMed
  • CiNii Articles
  • KAKEN
• Abstract License Flag: Disallowed