The correlation between expression and localization of a foreign gene product in rice endosperm.
Glucagon-like peptide 1 (GLP-1) is a 30 amino acid peptide hormone involved in insulin stimulation that is dependent upon blood glucose levels. We have previously reported that when this short peptide gene was directly expressed under the control of a glutelin promoter and its signal peptide, it was not accumulated in transgenic rice seed due to gene silencing. However, when the modified GLP-1 (mGLP-1) gene was enlarged to 5xmGLP-1 (mGLPx5) by tandem repeat, no silencing was observed. The mGLPx5 peptide could be accumulated in rice seed and its localization was mainly limited to the endoplasmic reticulum (ER). We also investigated alternative cellular localization sites that would increase accumulation. The relationship between the expression level and localization was examined by attaching the chitinase signal peptide to mGLPx5 to direct it into the intercellular space (apoplast), or by expression as a fusion protein with glutelin by insertion into a variable region of the acidic subunit, thus directing the peptide to protein body II (PB II). Attachment of the KDEL ER retention signal to the 6xmGLP-1 (mGLPx6) or its fusion to the C-terminus of the 13 kDa prolamin directed the peptide to the ER or PB I, respectively. Unexpectedly, these results indicated that mGLPx5 without any signal except for the glutelin signal peptide was accumulated to the greatest extent in rice endosperm. It can thus be concluded that the ER is a suitable intracellular organelle for accumulation of mGLPx5 peptide.
Yasuda
Hiroshi
H
Department of Plant Biotechnology, National institute of Agrobiological Sciences, Tsukuba, Ibaraki, 305-8602 Japan.
Hayashi
Yuji
Y
Jomori
Takahito
T
Takaiwa
Fumio
F
D002688:Chitinases / Q000032:analysis / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000648:ultrastructure; D018506:Gene Expression Regulation, Plant / Q000235:genetics* / Q000502:physiology; D020868:Gene Silencing / Q000502:physiology; D052216:Glucagon-Like Peptide 1 / Q000032:analysis* / Q000235:genetics*; D005983:Glutens / Q000032:analysis / Q000235:genetics; D007150:Immunohistochemistry; D008854:Microscopy, Electron; D012275:Oryza / Q000235:genetics* / Q000502:physiology; D010940:Plant Proteins / Q000032:analysis / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics; D055315:Prolamins; D012639:Seeds / Q000737:chemistry* / Q000502:physiology / Q000648:ultrastructure; D020080:Tandem Repeat Sequences / Q000235:genetics
D018744:DNA, Plant; D010940:Plant Proteins; D055315:Prolamins; D005983:Glutens; D052216:Glucagon-Like Peptide 1; D002688:Chitinases