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dc.contributor.advisorMacknight, Richard
dc.contributor.advisorVarkonyi-Gasic, Erika
dc.contributor.authorWu, Rongmei
dc.date.available2018-11-02T00:37:27Z
dc.date.copyright2018
dc.identifier.citationWu, R. (2018). The role of the SVP gene family in controlling bud dormancy (Thesis, Doctor of Philosophy). University of Otago. Retrieved from http://hdl.handle.net/10523/8518en
dc.identifier.urihttp://hdl.handle.net/10523/8518
dc.description.abstractSHORT VEGETATIVE PHASE (SVP)-like genes in the StMADS11 gene family have roles in the regulation of flowering time in annual plants and there is evidence that they also function in the regulation of dormancy in perennials. Expression patterns of SVP-like DORMANCY-ASSOCIATED MADS-BOX (DAM) genes in some Prunus species indicate their role in dormancy and their deletion is likely responsible for the non-dormant, evergrowing peach phenotype. As these Prunus species are recalcitrant to transformation, the hypothesis that SVP genes regulate dormancy could not be tested by transgenic overexpression. Kiwifruit (Actinidia spp.) SVP-like genes are predominantly expressed in axillary buds at the dormant stage and have been described as potential homologs of DAM genes, indicating they might be involved in controlling dormancy in kiwifruit. This study examined the function of the kiwifruit AcSVP2 gene by overexpressing it in two kiwifruit species with different chilling requirements for spring budbreak and in annual tobacco. The results suggest that in the high-chill kiwifruit A. chinensis var. deliciosa AcSVP2 has no significant effect on establishing dormancy, but delays budbreak. This delay in budbreak can be overcome by sufficient chilling. Overexpression of AcSVP2 in the low-chill kiwifruit species A. eriantha, had minimal effect and in transgenic tobacco it reduced plant height, delayed root growth and seed germination, but did not affect flowering time. Transcriptomic analyses (RNA-seq) of AcSVP2 transgenic lines during dormancy revealed that many of AcSVP2 regulated genes are involved in stress responses, mostly osmotic and cold response genes, with a subset also identified as ABA-responsive genes. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) of AcSVP2 transgenic lines showed that AcSVP2 directly binds to many target genes that function in a range of biological processes, especially those involved in repressing meristem activity and ABA-mediated dehydration pathways. AcSVP2 targeted genes also largely overlap with Arabidopsis SVP targeted genes expressed during vegetative growth and flower development. This suggests a conserved mechanism of action exists between SVP genes of taxonomically distant plants. Two regulatory systems associated with the control of AcSVP2 expression were studied: epigenetic regulation by histone modification was examined using chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) and regulation mediated by transcription factors (TFs) was investigated using in vivo Nicotiana benthamiana transient assays. Results from the ChIP-qPCR experiments showed that accumulation and reduction of trimethylated H3K4 (H3K4me3) and acetylated H3 (H3ac) correlated with the expression of AcSVP2 during dormancy in kiwifruit (A. chinensis). A 2.3kb AcSVP2 promoter region was used to screen 101 kiwifruit transcription factors and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1)-like genes, and potential candidate genes of bZIP, NAC, and MYB in the osmotic and abscisic acid signalling pathway were identified as possible regulators of AcSVP2 expression. Cis-regulatory elements (CREs) were identified in a 2.3kb AcSVP2 promoter region, including MADS-box protein binding elements, CArG-boxes, ABA response elements (ABRE), and drought response TFs binding motifs (MYCATERD1), and are likely to represent the binding sites of the TFs investigated. Kiwifruit AcSVP family genes, AcSVP1 and AcSVP4, showed elevated expression over the period of winter dormancy. Similar to AcSVP2, overexpression of AcSVP1 and AcSVP4 in the high-chill kiwifruit A. chinensis var. deliciosa, also delayed budbreak in spring, but had no effect on budbreak and flowering in the low-chill A. eriantha. Overexpression of AcSVP1 in tobacco ‘Maryland Mammoth’ significantly delayed flowering and seed germination, while AcSVP4 only had a minor effect on flowering and seed germination. Analysis of cis-regulatory motifs in the promoter regions of AcSVP1 and AcSVP4 and promoter transactivation assay revealed that expression of AcSVP1 and AcSVP4 may be controlled by transcription factors, similar to those identified as possible regulators of AcSVP2. These results suggests a conserved regulatory mechanism and a synergistic role for kiwifruit AcSVP1, AcSVP2 and AcSVP4 in preventing premature kiwifruit budbreak during winter. Interrogation of SVP-like genes in apple (Malus × domestica ‘Royal Gala’) genome confirmed that there are three DAM-like and two SVP-like genes. Expression analysis in apical buds of ‘Sciros’ apple trees over the seasons revealed that transcripts of MdDAMa and MdDAMc were elevated in the early autumn, and mostly absent over the winter and spring months. MdDAMb, MdSVPa and MdSVPb were expressed in the winter. Ectopic expression of MdDAMb and MdSVPa in ‘Royal Gala’ apple plants resulted in delayed budbreak in spring. Overall, the findings of this study have advanced our understanding of the roles the SVP/DAM-like genes play in the dormancy process of woody perennials.
dc.language.isoen
dc.publisherUniversity of Otago
dc.rightsAll items in OUR Archive are provided for private study and research purposes and are protected by copyright with all rights reserved unless otherwise indicated.
dc.subjectabscisic acid (ABA)
dc.subjectapple (Malus spp.)
dc.subjectbudbreak
dc.subjectdormancy
dc.subjectChromatin-immunoprecipitation followed by sequencing (ChIP-seq)
dc.subjectDormancy associated MADS-box gene (DAM)
dc.subjecthistone modification
dc.subjectkiwifruit (Actinidia spp.)
dc.subjectPromoter fusions
dc.subjectRNA-seq
dc.subjectShort Vegetative Phase gene (SVP)
dc.subjecttranscription factors
dc.titleThe role of the SVP gene family in controlling bud dormancy
dc.typeThesis
dc.date.updated2018-11-01T22:50:43Z
dc.language.rfc3066en
thesis.degree.disciplineBiochemistry
thesis.degree.nameDoctor of Philosophy
thesis.degree.grantorUniversity of Otago
thesis.degree.levelDoctoral
otago.interloanyes
otago.openaccessAbstract Only
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