ER membrane-localized oxidoreductase Ero1 is required for disulfide bond formation in the rice endosperm.
The developing endosperm of rice (Oryza sativa, Os) synthesizes a large amount of storage proteins on the rough (r)ER. The major storage proteins, glutelins and prolamins, contain either intra or intermolecular disulfide bonds, and oxidative protein folding is necessary for the sorting of the proteins to the protein bodies. Here, we investigated an electron transfer pathway for the formation of protein disulfide bonds in the rER of the rice endosperm, focusing on the roles of the thiol-disulfide oxidoreductase, OsEro1. Confocal microscopic analysis revealed that N-glycosylated OsEro1 is localized to the rER membrane in the subaleurone cells, and that targeting of OsEro1 to the rER membrane depends on the N-terminal region from Met-1 to Ser-55. The RNAi knockdown of OsERO1 inhibited the formation of native disulfide bonds in the glutelin precursors (proglutelins) and promoted aggregation of the proglutelins through nonnative intermolecular disulfide bonds in the rER. Inhibition of the formation of native disulfide bonds was also observed in the seeds of the esp2 mutant, which lacks protein disulfide isomerase-like (PDIL)1;1, but shows enhanced OsEro1 expression. We detected the generation of H(2)O(2) in the rER of the WT subaleurone cells, whereas the rER-derived H(2)O(2) levels decreased markedly in EM49 homozygous mutant seeds, which have fewer sulfhydryl groups than the WT seeds. Together, we propose that the formation of native disulfide bonds in proglutelins depends on an electron transfer pathway involving OsEro1 and OsPDIL.
Onda
Yayoi
Y
Division of Plant Sciences, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602, Japan.
Kumamaru
Toshihiro
T
Kawagoe
Yasushi
Y
D004220:Disulfides / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000201:enzymology* / Q000378:metabolism; D017343:Genes, Plant; D006031:Glycosylation; D006720:Homozygote; D006861:Hydrogen Peroxide / Q000737:chemistry; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology*; D018613:Microscopy, Confocal / Q000379:methods; D012275:Oryza / Q000201:enzymology / Q000235:genetics*; D010455:Peptides / Q000737:chemistry; D010940:Plant Proteins / Q000235:genetics* / Q000502:physiology*; D011490:Protein Disulfide Reductase (Glutathione) / Q000235:genetics / Q000502:physiology*; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D013347:Subcellular Fractions
D004220:Disulfides; D008565:Membrane Proteins; D010455:Peptides; D010940:Plant Proteins; D006861:Hydrogen Peroxide; D011490:Protein Disulfide Reductase (Glutathione)