Screening And Functional Identification Of Peach Dormancy Regulate Factor PpDAM5 And Its Interaction Proteins

Dormancy is an important process for perennials to withstand extreme weather conditions in winter.Only after safely passing the dormancy period can they grow and develop normally in the following period and start a new cycle.In temperate perennials,the onset and release of environmentally-induced dormancy is coordinated with the cessation and recovery of growth.Recent studies have shown that ABA is a hormone that different species need to regulate dormancy,and it plays an important role in dormancy.The transcription factors of the MADS-box family have the function of regulating plant growth and development.Among them,the SVP(SHORT VEGETATIVE PHASE)-Like gene Pp DAM5(dormancy-associated MADS-box)found in peach plays an important role in the release of endodormancy.However,The molecular regulation mechanism of Pp DAM5 in peach(Prunus persica)has not been clarified yet.SVP genes are usually used as flowering inhibitors.What role Pp DAM5 plays in the process of flower bud germination and flowering is worth exploring.In this experiment,the flower buds of ‘Chunxue’ peach blossoms were used as the test material,and the endodormancy mechanism of peach blossom buds was explained from the physiological and molecular levels.The main findings are as follows:(1)According to the accumulation of Chilling Requirement(CR)and the germination rate of the flower buds of the ‘chunxue’ peach,the flower bud dormancy period was divided.During the entire endodormancy period,the growth and development of flower buds is slow or even stagnant.In the early period of endodormancy,the water content of flower buds continued to decrease,reaching the lowest on November 15 th,and then with the satisfaction of cold requirements,the flower buds entered the endodormancy release period,and the water content of the flower bud begins to increase.During the whole period of endodormancy,the quality of flower buds did not change significantly.After November 15 th,the quality of flower buds increased during the release of endodormancy.(2)The analysis of the expression level of Pp DAM5 during the dormancy period showed that the expression level of Pp DAM5 reached the highest during the endodormancy period,and it was down-regulated after November 15 th,that is,it was down-regulated during the release period of the endodormancy.ABA treatment inhibited the germination of flower buds and up-regulated the expression of Pp DAM5.The overexpression strain Pp DAM5-OE showed a phenotype that inhibited germination.Through yeast two-hybrid screening,we isolated the BTB-TAZ domain protein Pp BT3(BTB AND TAZ DOMAIN PROTEIN 3)that interacts with the Pp DAM5 protein in the peach variety ‘Chunxue’.Analysis of the tissue-specific expression of Pp BT3 gene found that the expression level of Pp BT3 in stems and seeds was significantly higher than other parts,and the expression level of Pp BT3 gene in pistils and sepals was relatively higher than other tissues.Overexpression of Pp BT3 in Arabidopsis thaliana promoted seed germination and reduced At SVP expression.In the Pp DAM5-OE strain,the phenomenon of inhibiting ATBT3 gene expression was also found.On the MS solid mediuum containing 1μM ABA,Pp BT3-OE has obvious resistance to inhibiting ABA to promote germination.In addition,q RT-PCR experiments showed that the transcription level of Pp BT3 gradually increased during the release of endodormancy,which was contrary to the trend of the expression pattern of Pp DAM5.Our results show that Pp BT3 regulates the release of endodormancy of peach buds by interacting with Pp DAM5,thus revealing a new mechanism for regulating the release of endodormancy of flower buds.(3)Through yeast two-hybrid screening,we isolated Pp CMB1(Pp MADS2),which interacts with the Pp DAM5 protein,from MADS-box family in the peach variety "Chunxue".We conducted a tissue-specific analysis of Pp CMB1 and found that Pp CMB1 is mainly expressed in the pistil and carpel.Compared with the Columbia wild-type control,the Pp CMB1 overexpression line showed that Pp CMB1-oe had a significant phenotype that promoted early flowering.Our results indicate that Pp CMB1 promotes the function of early flowering.After the end of ecodormancy in peaches,Pp CMB1 may promote flowering through interacts with Pp DAM5 protein.