Dataset Information

VvPL15 Is the Core Member of the Pectate Lyase Gene Family Involved in Grape Berries Ripening and Softening.

ABSTRACT: The process of ripening and softening in grape begins at veraison and is closely related to the depolymerization of pectin components. A variety of enzymes are involved in pectin metabolism and one class of enzyme, pectin lyases (PLs), have been reported to play an important role in softening in many fruits; however, little information is available on the VvPL gene family in grape. In this study, 16 VvPL genes were identified in the grape genome using bioinformatics methods. Among them, VvPL5, VvPL9, and VvPL15 had the highest expression levels during grape ripening, which suggests that these genes are involved in grape ripening and softening. Furthermore, overexpression of VvPL15 affects the contents of water-soluble pectin (WSP) and acid-soluble pectin (ASP) in the leaves of Arabidopsis and significantly changes the growth of Arabidopsis plants. The relationship between VvPL15 and pectin content was further determined by antisense expression of VvPL15. In addition, we also studied the effect of VvPL15 on fruit in transgenic tomato plants, which showed that VvPL15 accelerated fruit ripening and softening. Our results indicate that VvPL15 plays an important role in grape berry softening during ripening by depolymerizing pectin.

SUBMITTER: Ma Y

PROVIDER: S-EPMC10253464 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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VvPL15 Is the Core Member of the Pectate Lyase Gene Family Involved in Grape Berries Ripening and Softening.

Ma Yuying Y Wang Chukun C Gao Zhen Z Yao Yuxin Y Kang Hui H Du Yuanpeng Y

International journal of molecular sciences 20230526 11

The process of ripening and softening in grape begins at veraison and is closely related to the depolymerization of pectin components. A variety of enzymes are involved in pectin metabolism and one class of enzyme, pectin lyases (PLs), have been reported to play an important role in softening in many fruits; however, little information is available on the VvPL gene family in grape. In this study, 16 VvPL genes were identified in the grape genome using bioinformatics methods. Among ...[more]

PMID: 37298267

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Project description:BackgroundPectin is one of the major and most complex plant cell wall components that needs to be overcome by microorganisms as part of their strategies for plant invasion or nutrition. Microbial pectinolytic enzymes therefore play a significant role for plant-associated microorganisms and for the decomposition and recycling of plant organic matter. Recently, comparative studies revealed significant gene copy number expansion of the polysaccharide lyase 1 (PL1) pectin/pectate lyase gene family in the Clonostachys rosea genome, while only low numbers were found in Trichoderma species. Both of these fungal genera are widely known for their ability to parasitize and kill other fungi (mycoparasitism) and certain species are thus used for biocontrol of plant pathogenic fungi.ResultsIn order to understand the role of the high number of pectin degrading enzymes in Clonostachys, we studied diversity and evolution of the PL1 gene family in C. rosea compared with other Sordariomycetes with varying nutritional life styles. Out of 17 members of C. rosea PL1, we could only detect two to be secreted at acidic pH. One of them, the pectate lyase pel12 gene was found to be strongly induced by pectin and, to a lower degree, by polygalacturonic acid. Heterologous expression of the PEL12 in a PL1-free background of T. reesei revealed direct enzymatic involvement of this protein in utilization of pectin at pH 5 without a requirement for Ca2+. The mutants showed increased utilization of pectin compounds, but did not increase biocontrol ability in detached leaf assay against the plant pathogen Botrytis cinerea compared to the wild type.ConclusionsIn this study, we aimed to gain insight into diversity and evolution of the PL1 gene family in C. rosea and other Sordariomycete species in relation to their nutritional modes. We show that C. rosea PL1 expansion does not correlate with its mycoparasitic nutritional mode and resembles those of strong plant pathogenic fungi. We further investigated regulation, specificity and function of the C. rosea PEL12 and show that this enzyme is directly involved in degradation of pectin and pectin-related compounds, but not in C. rosea biocontrol.

Project description:Background and aimsThe programmed softening occurring during fruit development requires scission of cell wall polysaccharides, especially pectin. Proposed mechanisms include the action of wall enzymes or hydroxyl radicals. Enzyme activities found in fruit extracts include pectate lyase (PL) and endo-polygalacturonase (EPG), which, in vitro, cleave de-esterified homogalacturonan in mid-chain by β-elimination and hydrolysis, respectively. However, the important biological question of whether PL exhibits action in vivo had not been tested.MethodsWe developed a method for specifically and sensitively detecting in-vivo PL products, based on Driselase digestion of cell wall polysaccharides and detection of the characteristic unsaturated product of PL action.Key resultsIn model in-vitro experiments, pectic homogalacturonan that had been partially cleaved by commercial PL was digested to completion with Driselase, releasing an unsaturated disaccharide ('ΔUA-GalA'), taken as diagnostic of PL action. ΔUA-GalA was separated from saturated oligogalacturonides (EPG products) by electrophoresis, then subjected to thin-layer chromatography (TLC), resolving ΔUA-GalA from higher homologues. The ΔUA-GalA was confirmed as 4-deoxy-β-l-threo-hex-4-enopyranuronosyl-(1→4)-d-galacturonic acid by NMR spectroscopy. Driselase digestion of cell walls from ripe fruits of date (Phoenix dactylifera), pear (Pyrus communis), rowan (Sorbus aucuparia) and apple (Malus pumila) yielded ΔUA-GalA, demonstrating that PL had been acting in vivo in these fruits prior to harvest. Date-derived ΔUA-GalA was verified by negative-mode mass spectrometry, including collision-induced dissociation (CID) fragmentation. The ΔUA-GalA:GalA ratio from ripe dates was roughly 1:20 (mol mol-1), indicating that approx. 5 % of the bonds in endogenous homogalacturonan had been cleaved by in-vivo PL action.ConclusionsThe results provide the first demonstration that PL, previously known from studies of fruit gene expression, proteomic studies and in-vitro enzyme activity, exhibits enzyme action in the walls of soft fruits and may thus be proposed to contribute to fruit softening.

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VvPL15 Is the Core Member of the Pectate Lyase Gene Family Involved in Grape Berries Ripening and Softening.

Publications

<i>VvPL15</i> Is the Core Member of the Pectate Lyase Gene Family Involved in Grape Berries Ripening and Softening.

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