Isolation And Characterization Of MawuAP2 And Its Interacting Genes From Magnolia Wufengensis

The discovery of functions and regulatory mechanisms of flower organ trait genes can not only provide important genetic resources for the cultivation of garden ornamental plants,but also provide important theoretical guidance for the improvement of their genetic breeding.Magnolia wufengensis is peculiar to China as a rare ornamental species,and its ornamental and economic values are mainly due to the abundant diversity in floral organ traits.However,the regulatory mechanisms of floral organ morphogenesis and the variation in tepal number have not yet been discovered.AP2(APETALA2)has long been considered as an A-class floral organ identity gene to specify the tepal morphogenesis,and it is closely related to the formation of additional tepals.Previous functional studies of AP2-like genes have mainly focused on core eudicot and monocot taxa.However,the functions of AP2-like genes of the more ancient angiosperm taxa were not well characterized until now.Considering the primitive evolutionary position in angiosperms,and the abundant diversity in tepal number of M wufengensis,therefore,it will be of great scientific significance that characterization of the AP2 orthologous gene of M wufengensis to understand the function of AP2-like genes in the ancient angiosperms and the functional evolution of AP2-like genes in the angiosperms.To this end,we isolated and charaterized the M wufengensis AP2 ortholog MawuAP2 on the analysis of phylogeny,sequence organization,expression pattern and function;we obtained and characterized the genes that directly interacted with MawuAP2 using yeast two hybrid screening technology to further elucidate the function of the MawuAP2 and the relationship among them.The main research results are as follows:(1)MawuAP2 and AP2 of Arabidopsis thaliana were grouped together in the euAP2 subgroup lineage,indicating that MawuAP2 cloned in this study was the ortholog of AP2(2)Although the AP2 and ANT group lineage genes share highly similar AP2 domain,their exon-intron organization is greatly different;in addition,exon fusion frequently appeared in the ANT group lineage genes.(3)MawuAP2 was expressed in roots,stems,leaves,fruits,seeds and all the whorls of floral organs in M wufengensis,rather than being limited to tepals,this broad expression pattern suggests that MawuAP2 is likely to have the broader range of functions.(4)Although the AP2 gene of model plant Arabidopsis thaliana has always been considered as an A class functional gene that determines the morphological formation of perianth,functional analysis showed that the MawuAP2 has the function of inhibiting flowering,rather than regulating the morphologensis of tepals.(5)Based on the analysis of phylogeny,expression and functions of the whole AP2/EREBP family genes,it is further clarified that the function that inhibiting flowering transition may be the real mainstream function of the AP2 group genes,and even the common function of all the AP2/EREBP family genes.(6)The FT/TFL1 family genes of M wufengensis possessed conserved regulatory functions in flowering transition,but they have produced unique expression patterns different from that of herbs,and the regulation of their expression level may be the key to realize the transition of juvenile phase and the seasonal vegetative to reproductive growth.(7)MawuAP2 can established interaction with MawuFT specifically,which may be to regulate their functions at the protein level.In summary,although the molecular mechanism of multi-petal formation in M wufengensis has still not been discovered,we found that the real mainstream function of AP2 and its homologous genes is to inhibit flowering transition rather than to specify the morphogenesis of tepals,which has broken the traditional awareness of the mainstream function of AP2 and its homologous genes.Besides,we discussed that the functional regulation mechanism between MawuFT and MawuAP2 at the protein-protein interaction level were also identified,and the molecular mechanism that how FT/TFL1 family genes regulate the flowering transition of M.wufengensis.Based on the expression patterns and functions of M wufengensis FT/TFL1 family genes,the molecular regulatory mechanism of FT/TFL1 family genes involved in the transition from juvenile to adult phase and the seasonal transition from vegetative to reproductive growth in M wufengensis was proposed for the first time.The biological function of the specific protein interaction between MawuAP2 and MawuFT was also explored.This study not only systematically analyzed the functions and regulatory mechanisms of MawuAP2 and FT/TFL1 family genes of M wufengensis in flower development,but also provided important gene resources and theoretical guidance for the cultivation and the improvement of ornamental value of the new varieties of M wufengensis and other woody plants.