title: A point mutation at the Miniature1 seed locus reduces levels of the encoded protein, but not its mRNA, in maize.
abstract: We report here on the molecular nature of an EMS-induced mutant, mn1-89, a leaky semidominant allele of the Miniature1 (Mn1) seed locus that encodes a seed-specific cell wall invertase, INCW2. The mn1-89 locus specifies normal levels of the Incw2 transcript but extremely low levels (about 6% of normal) of the protein and enzyme activity are expressed. Sequence analysis of Incw2 clones derived from the parental Mn1 and the mutant genotypes shows a C to T transition in the mn1-89 allele, leading to a single amino acid alteration (proline to leucine) near the C-terminus of the mutant INCW2 protein. Although this change is not in the catalytic domain, putative N-glycosylation sites, or the beta-fructosidase motif, it does lie in a motif that is well conserved among all plant invertases and related fructosyltransferases. On the basis of these genetic in planta data, we believe we have identified a proline residue in a hitherto unknown GPFG motif as critical for the stability of such proteins. The single base change (C to T) also leads to the elimination of a BglI restriction site in the mutant allele. Indeed, BglI restriction digests of genomic DNAs from mn1-89 and Mn1 genotypes show one and two fragments, respectively. Sequence analysis of RT-PCR-derived endosperm Incw clones from mn1-1 (the reference allele) seeds predict five amino acid substitutions relative to Mn1. Whether or not these sequences are encoded by the mn1-1 locus or another non-allelic Incw gene in the maize genome remains to be elucidated.
authors.affiliation: USDA-ARS, Department of Plant Pathology, University of Florida, Gainesville 32611-0680, USA.
authors.forename: S J
authors.initials: SJ
authors.lastname: Carlson
authors.forename: S
authors.initials: S
authors.lastname: Shanker
authors.forename: P S
authors.initials: PS
authors.lastname: Chourey
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015723:Gene Library; D008969:Molecular Sequence Data; D017354:Point Mutation; D011110:Polymorphism, Genetic; D012333:RNA, Messenger / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D003313:Zea mays / Q000235:genetics*
chemical_list: D018076:DNA, Complementary; D012333:RNA, Messenger