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Characterization of tomato PHYB1 and identification of molecular defects in four mutant alleles
Authors:Galina I. Lazarova  Tamayo Kubota  Shannon Frances  Janny L. Peters  Martin J.G. Hughes  Jörg Brandstädter  Márta Széll  Minami Matsui  Richard E. Kendrick  Marie-Michèle Cordonnier-Pratt  Lee H. Pratt
Affiliation:(1) Laboratory for Photoperception and Signal Transduction, Frontier Research Program, The Institute of Physical and Chemical Research (RIKEN), Hirosawa 2-1, Wako-shi, Saitama, 351-01, Japan;(2) Department of Botany, University of Georgia, Athens, GA, 30602, USA ( authors for correspondence)
Abstract:The structure of the gene encoding the apoprotein of phytochrome B1 (PHYB1) in tomato has been determined from genomic and cDNA sequences. In contrast to PHYA, PHYB1 lacks an intron upstream of the first ATG. A single transcription start site was found by 5prime RACE at –116. Tomato PHYB1 spans 7 kb starting from the first ATG. The coding region is organized into four exons as for other angiosperm PHY. The deduced apoprotein consists of 1131 amino acids, with a molecular mass of 125.4 kDa. Tomato phytochrome B1 shares 78% and 74% identity with Arabidopsis phytochromes B and D, respectively. Along with the normally spliced full-length transcripts, sequences of reverse transcriptase-PCR clones revealed five types of alternative transcripts. Each type of alternative transcript was missing a considerable part of the coding region, including the chromophore-binding site.The four putative PHYB1 mutants in tomato, which are temporarily red-light insensitive (tri), were each confirmed to have a mutation in PHYB1. Each mutation arose from a different, single-base substitution. Allele tri1 is presumably a null because the mutation introduces a stop at codon 92. In tri3, val-238 is replaced by Phe. The importance of this valine residue is evidenced by the fact that the tri3 phenotype is as strong as that of tri1. Alleles tri2 and tri4 encode proteins truncated at their C-termini. The former lacks either 170 or 438 amino acids, depending upon which of two types of splicing occurs during transcript maturation, while the latter lacks 225.
Keywords:photomorphogenic mutants  phytochrome  pre-mRNA splicing  alternative splicing  tomato
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