title: Unraveling the regulatory network of the MADS box transcription factor RIN in fruit ripening.
abstract: The MADS box transcription factor RIN is a global regulator of fruit ripening. However, the direct targets modulated by RIN and the mechanisms underlying the transcriptional regulation remain largely unknown. Here we identified 41 protein spots representing 35 individual genes as potential targets of RIN by comparative proteomic analysis of a rin mutant in tomato fruits. Gene expression analysis showed that the mRNA level of 26 genes correlated well with the protein level. After examining the promoter regions of the candidate genes, a variable number of RIN binding sites were found. Five genes (E8, TomloxC, PNAE, PGK and ADH2) were identified as novel direct targets of RIN by chromatin immunoprecipitation. The results of a gel mobility shift assay confirmed the direct binding of RIN to the promoters of these genes. Of the direct target genes, TomloxC and ADH2, which encode lipoxygenase (LOX) and alcohol dehydrogenase, respectively, are critical for the production of characteristic tomato aromas derived from LOX pathway. Further study indicated that RIN also directly regulates the expression of HPL, which encodes hydroperoxide lyase, another rate-limiting enzyme in the LOX pathway. Loss of function of RIN causes de-regulation of the LOX pathway, leading to a specific defect in the generation of aroma compounds derived from this pathway. These results indicate that RIN modulates aroma formation by direct and rigorous regulation of expression of genes in the LOX pathway. Taken together, our findings suggest that the regulatory effect of RIN on fruit ripening is achieved by targeting specific molecular pathways.
authors.affiliation: Key Laboratory of Plant Resources, Institute of Botany, Chinese Academy of Sciences, No. 20 Nanxincun, Xiangshan, Haidian District, Beijing 100093, China. gzqin@ibcas.ac.cn
authors.forename: Guozheng
authors.initials: G
authors.lastname: Qin
authors.forename: Yuying
authors.initials: Y
authors.lastname: Wang
authors.forename: Baohua
authors.initials: B
authors.lastname: Cao
authors.forename: Weihao
authors.initials: W
authors.lastname: Wang
authors.forename: Shiping
authors.initials: S
authors.lastname: Tian
mesh_terms: D000426:Alcohol Dehydrogenase / Q000235:genetics / Q000378:metabolism; D000446:Aldehyde-Lyases / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D015153:Blotting, Western; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D005638:Fruit / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D053263:Gene Regulatory Networks; D008084:Lipoxygenase / Q000235:genetics / Q000378:metabolism; D018551:Solanum lycopersicum / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D026161:MADS Domain Proteins / Q000235:genetics / Q000378:metabolism*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D020543:Proteome / Q000235:genetics / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction
chemical_list: D026161:MADS Domain Proteins; D010940:Plant Proteins; D020543:Proteome; D003577:Cytochrome P-450 Enzyme System; D000426:Alcohol Dehydrogenase; D008084:Lipoxygenase; D000446:Aldehyde-Lyases; C054540:hydroperoxide lyase