title: The Arabidopsis 14-3-3 protein, GF14omega, binds to the Schizosaccharomyces pombe Cdc25 phosphatase and rescues checkpoint defects in the rad24- mutant.
abstract: The fission yeast (S. pombe) mitotic inducer gene, Spcdc25, interacts with the plant cell cycle to establish a small cell size phenotype compared with wild-type cells. We have investigated the nature of this interaction by yeast two-hybrid screening using Spcdc25 as bait in a cDNA library prepared from root tips of Arabidopsis thaliana (L.) Heynh. Three 14-3-3 proteins were detected: G-box Factor-like (GF)14kappa, lambda and omega; binding with Spcdc25 was confirmed by an independent immunoprecipitation assay. To test for cell cycle checkpoint function, GF14kappa, lambda and omega were transformed independently, using the strong nmt1+ promoter, into rad24-, a fission yeast mutant deficient in a 14-3-3 checkpoint protein. When exposed to UV irradiation or in the presence of 10 mM hydroxyurea, only cells transformed with GF14omega could fully rescue the defects in the DNA-damage and DNA-replication checkpoints of this mutant. Supporting evidence for a GF14omega cell cycle function was provided by semi-quantitative reverse transcription-polymerase chain reaction indicating that expression of this gene was elevated in regions of the plant that comprise dividing cells whereas GF14kappa and lambda expression was more evenly detected in all tissues examined. The data are consistent with the hypothesis that interaction between Spcdc25 and the plant cell cycle occurs at the level of a 14-3-3 protein with distinct checkpoint properties.
authors.affiliation: Hannah Research Institute, KA6 5HL, Ayr, UK.
authors.forename: David A
authors.initials: DA
authors.lastname: Sorrell
authors.forename: Angela M
authors.initials: AM
authors.lastname: Marchbank
authors.forename: David A
authors.initials: DA
authors.lastname: Chrimes
authors.forename: J Richard
authors.initials: JR
authors.lastname: Dickinson
authors.forename: Hilary J
authors.initials: HJ
authors.lastname: Rogers
authors.forename: Dennis
authors.initials: D
authors.lastname: Francis
authors.forename: Claire S
authors.initials: CS
authors.lastname: Grierson
authors.forename: Nigel G
authors.initials: NG
authors.lastname: Halford
mesh_terms: D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004249:DNA Damage; D017931:DNA Primers; D018744:DNA, Plant / Q000528:radiation effects; D016366:Open Reading Frames; D016133:Polymerase Chain Reaction; D012568:Schizosaccharomyces / Q000201:enzymology*; D014466:Ultraviolet Rays; D020687:cdc25 Phosphatases / Q000378:metabolism*
chemical_list: D029681:Arabidopsis Proteins; D002135:Calcium-Binding Proteins; D017931:DNA Primers; D018744:DNA, Plant; C087620:GF14 protein, Arabidopsis; D020687:cdc25 Phosphatases