modified storage protein in synthesized, processed, and degraded in the seeds of transgenic plants

• Published in: Plant molecular biology 1988-01, Vol.11 (6), p.717-729
• Main Authors: Hoffman, L.M, Donaldson, D.D, Herman, E.M
• Format: Article
• Language: English
• Subjects: Agrobacterium tumefaciens; beta -phaseolin; Biological and medical sciences; chemical reactions; endoplasmic reticulum; Fundamental and applied biological sciences. Psychology; Gene expression; gene transfer; genes; genetic transformation; genetically modified organisms; glycosylation; Golgi apparatus; himet phaseolin; immunocytochemistry; insertional mutagenesis; methionine; Molecular and cellular biology; Molecular genetics; Nicotiana tabacum; phaseolin; Phaseolus vulgaris; protein synthesis
• ISSN: 0167-4412; 1573-5028
• DOI: 10.1007/bf00019513

In vitro mutagenesis was used to supplement the sulfur amino acid codon content of a gene encoding beta-phaseolin, a Phaseolus vulgaris storage protein. The number of methionine codons in the phaseolin gene was increased from three to nine by insertion of a 45 base pair (bp) synthetic duplex. Either modified or normal phaseolin genes were integrated into the genome of tobacco plants through Agrobacterium tumefaciens-mediated transformation. Although similar levels of phaseolin RNA are detected in seeds of plants transformed with either the normal or modified (himet) gene, the quantity of himet protein is consistently much lower than normal beta-phaseolin. Himet phaseolin is expressed in a temporal- and organ-specific fashion, and is N-glycosylated and assembled into trimers in the manner of normal phaseolin. After germination, both types of phaseolin are hydrolyzed, but the himet protein is more quickly degraded. Electron microscopic immunocytochemical observations of developing seeds indicate that the himet protein is primarily localized in the endoplasmic reticulum (ER) and in Golgi apparatus secretion vesicles. Himet phaseolin is absent from protein storage vacuoles, termed protein bodies, where normal phaseolin is deposited in transgenic tobacco. We interpret the immunocytochemical data to indicate that himet phaseolin is transported through the ER and Golgi apparatus and is then degraded in Golgi secretion vesicles or the protein bodies.