RESUMEN
In plants, flowering as a crucial developmental event is highly regulated by both genetic programs and environmental signals. Genetic analysis of flowering time mutants is instrumental in dissecting the regulatory pathways of flowering induction. In this study, we isolated the OsLF gene by its association with the T-DNA insertion in the rice late flowering mutant named A654. The OsLF gene encodes an atypical HLH protein composed of 419 amino acids (aa). Overexpression of the OsLF gene in wild type rice recapitulated the late flowering phenotype of A654, indicating that the OsLF gene negatively regulates flowering. Flowering genes downstream of OsPRR1 such as OsGI and Hd1 were down regulated in the A654 mutant. Yeast two hybrid and colocalization assays revealed that OsLF interacts strongly with OsPIL13 and OsPIL15 that are potentially involved in light signaling. In addition, OsPIL13 and OsPIL15 colocalize with OsPRR1, an ortholog of the Arabidopsis APRR1 gene that controls photoperiodic flowering response through clock function. Together, these results suggest that overexpression of OsLF might repress expression of OsGI and Hd1 by competing with OsPRR1 in interacting with OsPIL13 and OsPIL15 and thus induce late flowering.
Asunto(s)
Flores/metabolismo , Genes de Plantas/fisiología , Oryza/metabolismo , Fotoperiodo , Proteínas de Plantas/metabolismo , Factores de Transcripción/metabolismo , Flores/genética , Regulación de la Expresión Génica de las Plantas/fisiología , Secuencias Hélice-Giro-Hélice , Oryza/genética , Proteínas de Plantas/genética , Factores de Transcripción/genéticaRESUMEN
Flowering time is regulated by genetic programs and environment signals in plants. Genetic analysis of flowering time mutants is instrumental in dissecting the regulatory pathways of flower induction. Genotype W378 is a rice (Oryza sativa) late-flowering mutant selected from our collections of T-DNA insertion line. The T-DNA flanking gene in mutant W378 codes OsLFL1 (O. sativa LEC2 and FUSCA3 Like 1), a putative B3 DNA-binding domain-containing transcription factor. In wild-type rice OsLFL1 is expressed exclusively in spikes and young embryos, while in mutant W378 it is ectopically expressed. Introduction of OsLFL1-RNAi into mutant W378 successfully down-regulated OsLFL1 expression and restored flowering to almost normal time, indicating that overexpression of OsLFL1 confers late flowering for mutant W378. The flowering-promoting gene Ehd1 and its downstream genes are all down-regulated in W378. Thus, overexpression of OsLFL1 might delay the flowering of W378 by repressing the expression of Ehd1.
Asunto(s)
Flores/fisiología , Oryza/fisiología , Proteínas de Plantas/metabolismo , Factores de Transcripción/metabolismo , Regiones no Traducidas 5'/genética , Secuencia de Bases , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Regulación hacia Abajo/genética , Flores/genética , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Datos de Secuencia Molecular , Mutación/genética , Oryza/genética , Fenotipo , Proteínas de Plantas/genética , Transporte de Proteínas , Proteínas Recombinantes de Fusión/metabolismo , Fracciones Subcelulares/metabolismo , Factores de Tiempo , Factores de Transcripción/genética , Activación TranscripcionalRESUMEN
B3 domain was identified as a novel DNA-binding motif specific to higher plant species. The B3 proteins play important roles in plant development. In the mutant W378, the mutant gene coding OsLFL1, a putative B3 transcription factor gene, was ectopically expressed. In this study, it was found that the flowering promoting gene Ehd1 and its putative downstream genes were all repressed by OsLFL1. Electrophoretic mobility shift assays (EMSA) and chromatin immunoprecipitation (ChIP) analyses suggest that OsLFL1 binds to the RY cis-elements (CATGCATG) in the promoter of the Ehd1 gene. Thus, ectopically expressed OsLFL1 might repress Ehd1 via binding directly to the RY cis-elements in its promoter.
Asunto(s)
Regulación de la Expresión Génica de las Plantas/genética , Oryza/genética , Proteínas de Plantas/genética , Regiones Promotoras Genéticas/genética , Secuencias Reguladoras de Ácidos Nucleicos/genética , Factores de Transcripción/genética , Sitios de Unión , Unión ProteicaRESUMEN
Gama-glutamylcysteine synthetase (GCS) is a rate-limiting enzyme in GSH biosynthesis. The GCS gene has been cloned in Arabidopsis thaliana and other plants, but has still not been reported in rice. From rice mutant population generated from T-DNA insertion, we cloned the rice GCS gene from mutant L395 by T-DNA tag cloning method, and named it OsGCS (Genbank accession No. AJ508915). Full length OsGCS cDNA clones were obtained from a rice cDNA library by the PCR method. A comparison of the genome and cDNA sequence (Genbank accession No. AJ508916) shows that OsGCS gene is composed of 15 exons and 14 introns and coding a 493-amino acid protein. The OsGCS gene is highly homologous with the AtGCS gene in the coding region but completely different in the promoter region. The putative transcription start site (TSS) confirmed by RT-PCR was located 211 bp upstream of the translation start codon "ATG". In mutant L395, a single T-DNA copy was integrated between the second intron and second exon of OsGCS gene, causing one nucleotide deletion in the second exon and two nucleotide deletions in the second intron. No significant differences were found in Cd(2+) stress tolerance, rice GCS gene expression level and GSH content between mutant L395 and Zhonghua 11. It is possible that another GCS gene on chromosome 7 might complement function of OsGCS gene on chromosome 5.
