Functional association of cell death suppressor, Arabidopsis Bax inhibitor-1, with fatty acid 2-hydroxylation through cytochrome b₅.
Bax inhibitor-1 (BI-1) is a widely conserved cytoprotective protein localized in the endoplasmic reticulum (ER) membrane. We identified Arabidopsis cytochrome b(5)  (AtCb5) as an interactor of Arabidopsis BI-1 (AtBI-1) by screening the Arabidopsis cDNA library with the split-ubiquitin yeast two-hybrid (suY2H) system. Cb5 is an electron transfer protein localized mainly in the ER membrane. In addition, a bimolecular fluorescence complementation (BiFC) assay and fluorescence resonance energy transfer (FRET) analysis confirmed that AtBI-1 interacted with AtCb5 in plants. On the other hand, we found that the AtBI-1-mediated suppression of cell death in yeast requires Saccharomyces cerevisiae fatty acid hydroxylase 1 (ScFAH1), which had a Cb5-like domain at the N terminus and interacted with AtBI-1. ScFAH1 is a sphingolipid fatty acid 2-hydroxylase localized in the ER membrane. In contrast, AtFAH1 and AtFAH2, which are functional ScFAH1 homologues in Arabidopsis, had no Cb5-like domain, and instead interacted with AtCb5 in plants. These results suggest that AtBI-1 interacts with AtFAHs via AtCb5 in plant cells. Furthermore, the overexpression of AtBI-1 increased the level of 2-hydroxy fatty acids in Arabidopsis, indicating that AtBI-1 is involved in fatty acid 2-hydroxylation.
Nagano
Minoru
M
Institute of Molecular and Cellular Biosciences, the University of Tokyo, Bunkyo-ku, Tokyo 113-0032, Japan.
Ihara-Ohori
Yuri
Y
Imai
Hiroyuki
H
Inada
Noriko
N
Fujimoto
Masaru
M
Tsutsumi
Nobuhiro
N
Uchimiya
Hirofumi
H
Kawai-Yamada
Maki
M
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D016923:Cell Death; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D015786:Cytochromes b5 / Q000145:classification / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D015723:Gene Library; D005816:Genetic Complementation Test; D006900:Hydroxylation; D007425:Intracellular Membranes / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010802:Phylogeny; D025941:Protein Interaction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques
C491862:ATBI-1 protein, Arabidopsis; D029681:Arabidopsis Proteins; D005227:Fatty Acids; D008565:Membrane Proteins; D029701:Saccharomyces cerevisiae Proteins; D015786:Cytochromes b5; D003577:Cytochrome P-450 Enzyme System; C097486:CYP84A1 protein, Arabidopsis