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      An integrative analysis of the transcriptome and proteome of the pulp of a spontaneous late-ripening sweet orange mutant and its wild type improves our understanding of fruit ripening in citrus

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          Summary:

          ABA, ethylene, sucrose, and their related genes and pathways are involved in fruit ripening of citrus, and ABA may play a central role during the ripening process.

          Abstract

          Fruit ripening is a complex, genetically programmed process that occurs in conjunction with the differentiation of chloroplasts into chromoplasts and involves changes to the organoleptic properties of the fruit. In this study, an integrative analysis of the transcriptome and proteome was performed to identify important regulators and pathways involved in fruit ripening in a spontaneous late-ripening mutant (‘Fengwan’ orange, Citrus sinensis L. Osbeck) and its wild type (‘Fengjie 72-1’). At the transcript level, 628 genes showed a 2-fold or more expression difference between the mutant and wild type as detected by an RNA sequencing approach. At the protein level, 130 proteins differed by 1.5-fold or more in their relative abundance, as indicated by iTRAQ (isobaric tags for relative and absolute quantitation) analysis. A comparison of the transcriptome and proteome data revealed some aspects of the regulation of metabolism during orange fruit ripening. First, a large number of differential genes were found to belong to the plant hormone pathways and cell-wall-related metabolism. Secondly, we noted a correlation between ripening-associated transcripts and sugar metabolites, which suggests the importance of these metabolic pathways during fruit ripening. Thirdly, a number of genes showed inconsistency between the transcript and protein level, which is indicative of post-transcriptional events. These results reveal multiple ripening-associated events during citrus ripening and provide new insights into the molecular mechanisms underlying citrus ripening regulatory networks.

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          Most cited references 47

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          Mapping and quantifying mammalian transcriptomes by RNA-Seq.

          We have mapped and quantified mouse transcriptomes by deeply sequencing them and recording how frequently each gene is represented in the sequence sample (RNA-Seq). This provides a digital measure of the presence and prevalence of transcripts from known and previously unknown genes. We report reference measurements composed of 41-52 million mapped 25-base-pair reads for poly(A)-selected RNA from adult mouse brain, liver and skeletal muscle tissues. We used RNA standards to quantify transcript prevalence and to test the linear range of transcript detection, which spanned five orders of magnitude. Although >90% of uniquely mapped reads fell within known exons, the remaining data suggest new and revised gene models, including changed or additional promoters, exons and 3' untranscribed regions, as well as new candidate microRNA precursors. RNA splice events, which are not readily measured by standard gene expression microarray or serial analysis of gene expression methods, were detected directly by mapping splice-crossing sequence reads. We observed 1.45 x 10(5) distinct splices, and alternative splices were prominent, with 3,500 different genes expressing one or more alternate internal splices.
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            Target-decoy search strategy for increased confidence in large-scale protein identifications by mass spectrometry.

            Liquid chromatography and tandem mass spectrometry (LC-MS/MS) has become the preferred method for conducting large-scale surveys of proteomes. Automated interpretation of tandem mass spectrometry (MS/MS) spectra can be problematic, however, for a variety of reasons. As most sequence search engines return results even for 'unmatchable' spectra, proteome researchers must devise ways to distinguish correct from incorrect peptide identifications. The target-decoy search strategy represents a straightforward and effective way to manage this effort. Despite the apparent simplicity of this method, some controversy surrounds its successful application. Here we clarify our preferred methodology by addressing four issues based on observed decoy hit frequencies: (i) the major assumptions made with this database search strategy are reasonable; (ii) concatenated target-decoy database searches are preferable to separate target and decoy database searches; (iii) the theoretical error associated with target-decoy false positive (FP) rate measurements can be estimated; and (iv) alternate methods for constructing decoy databases are similarly effective once certain considerations are taken into account.
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              CTR1, a negative regulator of the ethylene response pathway in Arabidopsis, encodes a member of the raf family of protein kinases.

              We isolated a recessive Arabidopsis mutant, ctr1, that constitutively exhibits seedling and adult phenotypes observed in plants treated with the plant hormone ethylene. The ctr1 adult morphology can be phenocopied by treatment of wild-type plants with exogenous ethylene and is due, at least in part, to inhibition of cell elongation. Seedlings and adult ctr1 plants show constitutive expression of ethylene-regulated genes. The epistasis of ctr1 and other ethylene response mutants has defined the position of CTR1 in the ethylene signal transduction pathway. The CTR1 gene has been cloned, and the DNA sequences of four mutant alleles were determined. The gene encodes a putative serine/threonine protein kinase that is most closely related to the Raf protein kinase family.
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                Author and article information

                Affiliations
                1Key Laboratory of Horticultural Plant Biology, Ministry of Education, Huazhong Agricultural University , Wuhan 430070, PR China
                2National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University , Wuhan 430070, PR China
                Author notes
                * To whom correspondence should be addressed. E-mail: yihualin@ 123456mail.hzau.edu.cn
                Journal
                J Exp Bot
                J. Exp. Bot
                jexbot
                exbotj
                Journal of Experimental Botany
                Oxford University Press (UK )
                0022-0957
                1460-2431
                April 2014
                5 March 2014
                5 March 2014
                : 65
                : 6
                : 1651-1671
                10.1093/jxb/eru044
                3967095
                24600016
                © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

                Counts
                Pages: 21
                Categories
                Research Paper

                Plant science & Botany

                abscisic acid, ethylene, fruit ripening, itraq, proteome, sucrose, transcriptome.

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