Function and transcriptional regulation of CsKCS20 in the elongation of very-long-chain fatty acids and wax biosynthesis in Citrus sinensis flavedo

Abstract

Cuticular wax on plant aerial surfaces plays a vital role in the defense against various stresses, and the genes related to wax metabolism have been well documented in several model plants. However, there is very limited research on the key enzymes and transcription factors (TFs) associated with carbon chain distribution and wax biosynthesis in citrus fruit. In this study, an analysis of wax metabolites indicated that even carbon-chain (C24-C28) metabolites are the dominant wax components in citrus fruit, and a 3-ketoacyl-CoA synthase (KCS) family gene (CsKCS20) plays an important role in the carbon chain distribution during wax biosynthesis in a wax-deficient mutant (MT). Expression of CsKCS20 in yeast indicated that CsKCS20 can catalyze the biosynthesis of C22 and C24 very-long-chain fatty acids (VLCFAs). In addition, transcriptome and sequence analysis indicated that the differential expression of CsKCS20 between the wild-type (WT) and MT fruit can be partly attributed to the regulation of CsMYB96, which was further confirmed by yeast one-hybrid (Y1H) assays, electrophoretic mobility shift assays (EMSAs) and dual luciferase assays. The functions of CsMYB96 and CsKCS20 in wax biosynthesis were further validated by heterologous expression in Arabidopsis. In summary, this study elucidates the important roles of CsKCS20 and CsMYB96 in regulating VLCFA elongation and cuticular wax biosynthesis, which provides new directions for the improvement of citrus fruit wax quality in genetic breeding programs.

Genome-Wide Identification and Expression Profiling of KCS Gene Family in Passion Fruit (Passiflora edulis) Under Fusarium kyushuense and Drought Stress Conditions

“…Due to the important roles of KCS in plant cuticle synthesis and other biological processes, the genome-wide identification of the KCS gene family has been reported in various plant species including Arabidopsis thaliana ( Joubès et al, 2008 ), Zea mays ( Campbell et al, 2019 ), Arachis hypogea ( Huai et al, 2020 ), Brassica napus ( Xue et al, 2020 ), Vitis vinifera ( Guo et al, 2016 ), Gossypium hirsutum , G. arboreum , G. raimondii ( Xiao et al, 2016 ), Hordeum vulgare ( Tong et al, 2021 ), Atalantia buxifolia , Citrus ichangensis , C. medica, C. grandis , C. Sinensis , and C. clementina ( Yang H. et al, 2021 ) ranging from 16 to 58 KCS members. In addition, the KCS family members have been functionally characterized in Arabidopsis ( Lee et al, 2009 ; Kim et al, 2013 ; Hegebarth et al, 2017 ), C. sinensis ( Yang H. et al, 2021 ; Wang et al, 2022 ), O . sativa ( Wang X. et al, 2017 ), H. annuus ( González-Mellado et al, 2019 ), S. tuberosum ( Serra et al, 2009 ), M. domestica ( Lian et al, 2021 ), V. vinifera ( Yang Z. et al, 2020 ), and A. hypogaea ( Lokesh et al, 2019 ).…”

“…In sunflower ( Helianthus annuus ), HaKCS1 and HaKCS2 were expressed in seeds and involved in the elongation of fatty acids from C 18 to C 20 –C 24 respectively ( González-Mellado et al, 2019 ). Recently Wang et al (2022) also reported the involvement of CsKCS20 in VLCFAs elongation and cuticular wax biosynthesis.…”

Overexpression of β-Ketoacyl CoA Synthase 2B.1 from Chenopodium quinoa Promotes Suberin Monomers’ Production and Salt Tolerance in Arabidopsis thaliana

“…The overexpression of KCS2 homologs in Arabidopsis resulted in increased VLCFA production and the significant accumulation of cuticular wax [ 49 ]. Additionally, molecular characterization of Citrus sinensis KCS20 by ectopic expression in Arabidopsis and yeast indicated that it catalyzes the synthesis of C22 and C24 VLCFAs, similar to KCS2 or KCS20 in Arabidopsis [ 50 ]. In our study, overexpression of CqKCS2B.1 in Arabidopsis also led to increased contents of C22 and C24 VLCFAs and their derivatives in suberin monomers, suggesting that the function of KCS2 homologs is evolutionary and functionally conserved among diverse plant species.…”

Genomic and transcriptomic analyses provide insights into valuable fatty acid biosynthesis and environmental adaptation of yellowhorn

“…The latter was catalyzed by 3-ketoacyl-CoA synthase (KCS), 3-ketoacyl-CoA reductase (KCR), 3-hydroxacyl-CoA dehydratase (HCD), and trans -2,3-enoyl-CoA reductase (ECR). Of the four enzymes, KCS is the rate-limiting regulator ( Millar and Kunst, 1997 ; Joubès et al, 2008 ; Haslam and Kunst, 2013 ; Wang Y. et al, 2022 ). KCR, HCD, and ECR are expressed and function in all tissues with fatty acid biosynthesis ( Zheng et al, 2005 ; Paul et al, 2006 ; Bach et al, 2008 ; Beaudoin et al, 2009 ; Haslam and Kunst, 2013 ).…”