|Porch, Timothy - Tim|
Submitted to: Plant Journal
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/20/2005
Publication Date: 1/1/2006
Citation: Suzuki, M., Settles, M.A., Tseung, C., Li, Q., Latshaw, S., Wu, S., Porch, T., Schmelz, E.A., James, M.G., McCarty, D.R. 2006. The Maize viviparous 15 locus encodes the molybdopterin synthase small subunit. The Plant Journal 44(2):264-274. Interpretive Summary: An important mechanism for the response of plants to the environment is through the synthesis or perception of plant hormones. Abscisic acid (ABA) is a key plant hormone involved in the response of plants to stress and in the regulation of seed development. Corn seed lacking the ability to perceive or synthesize ABA germinates on the cob before the cob reaches maturity, a condition that is termed vivipary. In this study, the viviparous15 (vp15) corn gene, which is involved in the synthesis of ABA, was cloned. Thus, the identity, general function and location of Vp15 has been determined. Vp15 was cloned using the corn Mutator transposable element system. Transposable elements are mobile DNA fragments able to move to different positions within the genome. Using the Mutator transposon mutagenesis system and a novel PCR-based method, the vp15 gene was identified. The approach to cloning vp15 presented in this study is robust and has broad application for the cloning of other mutants in corn.
Technical Abstract: A new Zea mays viviparous seed mutant, viviparous15 (vp15), was isolated from the UniformMu transposon tagging population. In addition to precocious germination vp15 has an early seedling lethal phenotype. Biochemical analysis showed reduced activities of several enzymes that require molybdenum cofactor (MoCo) in vp15 mutant seedlings. Because MoCo is required for abscisic acid (ABA) biosynthesis, the viviparous phenotype is most likely due to ABA deficiency. We cloned the vp15 mutant using a novel high-throughput strategy for analysis of high-copy Mu lines: We used MUTAIL PCR to extract genomic sequences flanking the Mu transposons in the vp15 line. The Mu insertions specific to the vp15 line were identified by in silico subtraction using a database of MuTAIL sequences from 90 UniformMu lines. Annotation of the vp15 specific sequences revealed a Mu insertion in a gene homologous to human MOCS2A, the small subunit of molydopterin (MPT) synthase. Molecular analysis of two allelic mutations confirmed that Vp15 encodes a plant MPT synthase small subunit (ZmCNX7). Our results and a related paper reporting the cloning of maize viviparous13, demonstrate robust cloning strategies based on MuTAIL-PCR. Comparisons of Vp15 orthologs in genomes of three cereals and Arabidopsis thaliana identified a conserved sequence element in the 5’ untranslated region that resembles the A box regulatory motif in tRNA genes. In addition, the cereal orthologs contain a serine tRNA gene in the 5’ flanking sequence. These features suggest a possible involvement of RNA polymerase III factors in regulation of CNX7 in plants.