title: Accumulation of foreign polypeptides to rice seed protein body type I using prolamin portion sequences.
abstract: Rice prolamins are accumulated in endoplasmic reticulum (ER)-derived proteins bodies, although conserved sequences retained in ER are not confirmed. We investigated portion sequences of prolamins that must accumulate in PB-Is. Rice seed prolamins are accumulated in endoplasmic reticulum (ER)-derived protein body type I (PB-I), but ER retention sequences in rice prolamin polypeptides have not been confirmed. Here we investigated the lengths of the prolamin portion sequences required for accumulation in PB-Is. Of the rice prolamins, we compared 13a and 13b prolamins because the amino acid sequences of these prolamins are quite similar except for the presence or absence of Cys-residues. We also generated and analyzed transgenic rice expressing several prolamin portion sequence-GFP fusion proteins. We observed that in 13a prolamin, when the portion sequences were extended more than the 68th amino acid residue from the initiating methionine, the prolamin portion sequence-GFP fusion proteins were accumulated in PB-Is. In 13b prolamin, when the portion sequences were extended by more than the 82nd amino acid residue from the initiating methionine, the prolamin portion sequence-GFP fusion proteins were accumulated in PB-Is. When those fusion proteins were extracted under non-reduced or reduced conditions, the 13a prolamin portion sequence-GFP fusion proteins in PB-Is were soluble under only the reduced condition. In contrast, 13b prolamin portion sequence-GFP fusion proteins were soluble under both non-reduced and reduced conditions. These results suggest that the accumulation of 13a prolamin in PB-Is is associated with the formation of disulfide bonds and/or hydrophobicity in 13a prolamin polypeptide, whereas the accumulation of 13b prolamin in PB-Is was less involved in the formation of disulfide bonds.
authors.affiliation: Laboratory of Genetic Engineering, Graduate School of Life and Environmental Sciences, Kyoto Prefectural University, Shimogamo, Kyoto, 606-8522, Japan.
authors.forename: Ai
authors.initials: A
authors.lastname: Sasou
authors.affiliation: Laboratory of Genetic Engineering, Graduate School of Life and Environmental Sciences, Kyoto Prefectural University, Shimogamo, Kyoto, 606-8522, Japan.
authors.forename: Takanari
authors.initials: T
authors.lastname: Shigemitsu
authors.affiliation: Laboratory of Genetic Engineering, Graduate School of Life and Environmental Sciences, Kyoto Prefectural University, Shimogamo, Kyoto, 606-8522, Japan.|Biotechnology Research Department, Kyoto Prefectural Agriculture, Forestry and Fisheries Technology Research Center, Kitainayazuma, Seika-cho, Soraku-gun, Kyoto, 619-0244, Japan.
authors.forename: Shigeto
authors.initials: S
authors.lastname: Morita
authors.affiliation: Laboratory of Genetic Engineering, Graduate School of Life and Environmental Sciences, Kyoto Prefectural University, Shimogamo, Kyoto, 606-8522, Japan. masumura@kpu.ac.jp.|Biotechnology Research Department, Kyoto Prefectural Agriculture, Forestry and Fisheries Technology Research Center, Kitainayazuma, Seika-cho, Soraku-gun, Kyoto, 619-0244, Japan. masumura@kpu.ac.jp.
authors.forename: Takehiro
authors.initials: T
authors.lastname: Masumura
mesh_terms: D000595:Amino Acid Sequence; D002021:Buffers; D049452:Green Fluorescent Proteins / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D012275:Oryza / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D030821:Plants, Genetically Modified; D055315:Prolamins / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012639:Seeds / Q000235:genetics / Q000378:metabolism*; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology
chemical_list: D002021:Buffers; D010455:Peptides; D055315:Prolamins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D012967:Sodium Dodecyl Sulfate
keywords: Green fluorescent protein; Prolamin; Protein body type I; Rice