Arabidopsis protein disulfide isomerase-8 is a type I endoplasmic reticulum transmembrane protein with thiol-disulfide oxidase activity

BMC Plant Biology

Research Article

David A. Christopher¹  Christen Y. L. Yuen¹  Kristie Matsumoto¹  Eun Ju Cho¹  Roger Shek¹  Byung-Ho Kang² 

[1] Department of Molecular Biosciences and Bioengineering, University of Hawaii, 1955 East-West Rd., Ag. Science Rm 218, 96822, Honolulu, HI, USA;The Chinese University of Hong Kong, School of Life Sciences, Shatin, Hong Kong, SAR, China;

关键词: Endoplasmic reticulum;    Transmembrane;    Protein disulfide isomerase;    Protein folding;   

DOI  :  10.1186/s12870-016-0869-2

 received in 2016-03-10, accepted in 2016-08-08,  发布年份 2016

来源: Springer

【 摘 要 】

BackgroundIn eukaryotes, classical protein disulfide isomerases (PDIs) facilitate the oxidative folding of nascent secretory proteins in the endoplasmic reticulum by catalyzing the formation, breakage, and rearrangement of disulfide bonds. Terrestrial plants encode six structurally distinct subfamilies of PDIs. The novel PDI-B subfamily is unique to terrestrial plants, and in Arabidopsis is represented by a single member, PDI8. Unlike classical PDIs, which lack transmembrane domains (TMDs), PDI8 is unique in that it has a C-terminal TMD and a single N-terminal thioredoxin domain (instead of two). No PDI8 isoforms have been experimentally characterized to date. Here we describe the characterization of the membrane orientation, expression, sub-cellular localization, and biochemical function of this novel member of the PDI family.ResultsHistochemical staining of plants harboring a PDI8 promoter:β-glucuronidase (GUS) fusion revealed that the PDI8 promoter is highly active in young, expanding leaves, the guard cells of cotyledons, and in the vasculature of several organs, including roots, leaves, cotyledons, and flowers. Immunoelectron microscopy studies using a PDI8-specific antibody on root and shoot apical cells revealed that PDI8 localizes to the endoplasmic reticulum (ER). Transient expression of two PDI8 fusions to green fluorescent protein (spGFP-PDI8 and PDI8-GFP-KKED) in leaf mesophyll protoplasts also resulted in labeling of the ER. Protease-protection immunoblot analysis indicated that PDI8 is a type I membrane protein, with its catalytic domain facing the ER lumen. The lumenal portion of PDI8 was able to functionally complement the loss of the prokaryotic protein foldase, disulfide oxidase (DsbA), as demonstrated by the reconstitution of periplasmic alkaline phosphatase in Escherichia coli.ConclusionThe results indicate that PDI8 is a type I transmembrane protein with its catalytic domain facing the lumen of the ER and functions in the oxidation of cysteines to produce disulfide bonds. It likely plays a role in folding newly-synthesized secretory proteins as they translocate across the ER membrane into the lumen. These foundational results open the door to identifying the substrates of PDI8 to enable a more thorough understanding of its function in plants.

【 授权许可】

CC BY   
© The Author(s). 2016