Origin, Evolution, And Functional Characterization Of CDPK Genes And PPCK Genes From KFC Gene Family In Vitis Vinifera

Grapevine is a fruit crop with more than 8000 years of history as the human food,and it becomes the first and third important fruit crop in cultivated area and production respectively, which has valuable economic and cultural importance.Although the being non-model plant, the representative grapevine species(Vitis vinifera L. 'Pinot noir'),had finished the genome sequencing and marked the research focuses have moved into post-genomic level. In this era, two sorts of researches are favored, one from top to bottom, i.e mapping the genes regulating the phenotypes, and the other from bottom to top, i.e. identifying the functions of genes.In this dissertation, based on various tools including transcriptome sequencings and comparative genomic analysis, qPCR validations, and the combination of bioinformatics mining and statistical analyses, I studied the evolutionary connection of the grapevine CDPK genes and the PPCK genes in the view of evolution-development. The main results are as follows:1. Based on analyses of genomes and transcriptomes from representative plants, I identified that CDPK genes and PPCK genes are decendants from a single fused gene ancestor. Along evolution, decendants of this ancestor evolved into a multi-gene monophyly. I found this monophyly at least evolved in the ancestor of red algae.Considering it has no name available now, I named it as kinase fused calmodulin,KFC gene family. The CDPKs evolved at least in ancestor of red algae, whereas PPCKs date back to ancestor of gymnosperm. Based on transcriptional,structural,and expansion analysis, I found KFCs have played important roles in each of 6 major transitions in plant adaptation evolution.2. I compared the KFC gene family (including CDPKs) with CIPK-CBL family from grapevine and orchids, in terms of copy number evolution, subcellular locations,functional roles, and working mechanisms. I found that the KFC serves as the major calcium signal converter system in all plants including the grapevine.3. Relying on the haploid genome of Vitis vinifera, the bioinformatics mining of genome and transcriptome data, and molecular experimental validation, I found 17 CDPK genes in the grapevine, and found that all of which are at least as old as 100 million-year-old without gene duplications. With long history of negative selection, 4 CDPKs were found to transit functions from housekeeping expressional pattern to pollen-specific expressional pattern. This expressional pattern could be used for breeding male-sterile plants and seedless grapevine. I also found several grapevine CDPKs respond to drought, cold, and/or salt stresses, with potential for studying the crosstalk of these three abiotic stress signaling pathways.4. I have sequenced 6 transcriptome samples from day and night samplings from three CAM photosynthetic orchids in order to study the functions of PPCKs. I compared transcriptomes of a total of 13 orchids, all of which have well coverage of genes and good sequencing quality. Based on the transcriptome data,I build the tree of orchid life that divides the orchids into 5 subfamilies. The open accessed transcriptome data will also fasciliate future researches.5. Compared to eudicots including the Vitis vinifera, I identified PPCKs from different kinds of photosynthesis orchids. I found two PPCK clusters within the Orchidaceae. One of the clusters displayed the most significant diurnal rhythmed expressional pattern among all the 13 gene families enrolled in carbon fixation in CAM, and 4 sites of specific amino acids were also identified in the kinase domain.6. Based on the expressional pattern of PPCK, ME, PPDK, and PEPC genes, and their biochemical properties, I proposed a new carbon fixation pathway mediated by ME-PPDK, but not by PPCK-PEPC genes.