title: A defective signal peptide tethers the floury-2 zein to the endoplasmic reticulum membrane.
abstract: The maize (Zea mays L.) floury-2 (fl2) mutation is associated with a general decrease in storage protein synthesis, altered protein body morphology, and the synthesis of a novel 24-kD alpha-zein storage protein. Unlike storage proteins in normal kernels and the majority of storage proteins in fl2 kernels, the 24-kD alpha-zein contains a signal peptide that would normally be removed during protein synthesis and processing. The expected processing site of this alpha-zein reveals a putative mutation alanine-->valine (Ala-->Val) that is not found at other junctions between signal sequences and mature proteins. To investigate the impact of such a mutation on signal peptide cleavage, we have assayed the 24-kD fl2 alpha-zein in a co-translational processing system in vitro. Translation of RNA from fl2 kernels or synthetic RNA encoding the fl2 alpha-zein in the presence of microsomes yielded a 24-kD polypeptide. A normal signal peptide sequence, generated by site-directed mutagenesis, restored the capacity of the RNA to direct synthesis of a properly processed protein in a cell-free system. Both the fl2 alpha-zein and the fl2 alpha-zein (Val-->Ala) were translocated into the lumen of the endoplasmic reticulum. The processed fl2 alpha-zein (Val-->Ala) was localized in the soluble portion of the microsomes, whereas the fl2 alpha-zein co-fractionated with the microsomal membranes. By remaining anchored to protein body membranes during endosperm maturation, the fl2 zein may thus constrain storage protein packing and perturb protein body morphology.
authors.affiliation: Department of Botany, North Carolina State University, Raleigh 27695, USA.
authors.forename: J W
authors.initials: JW
authors.lastname: Gillikin
authors.forename: F
authors.initials: F
authors.lastname: Zhang
authors.forename: C E
authors.initials: CE
authors.lastname: Coleman
authors.forename: H W
authors.initials: HW
authors.lastname: Bass
authors.forename: B A
authors.initials: BA
authors.lastname: Larkins
authors.forename: R S
authors.initials: RS
authors.lastname: Boston
mesh_terms: D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides / Q000235:genetics; D010641:Phenotype; D017354:Point Mutation; D014176:Protein Biosynthesis; D021382:Protein Sorting Signals / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011817:Rabbits; D003313:Zea mays / Q000235:genetics / Q000378:metabolism; D015028:Zein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D009838:Oligodeoxyribonucleotides; D021382:Protein Sorting Signals; D015028:Zein