Isolation And Characterization Of Genes In Cotton Responsive To Multiple Stresses

Under natural conditions,plants have to deal with multiple abiotic and biotic stresses,including drought,salinity,cold,pathogens and herbivores.To survive,plants have evolved a multitude of mechanisms for adaptation to the changing environmental conditions.These adaptive responses at the cost of growth and reproduction,trade-offs often exist among them.Current studies reported so far have defined few stress-related genes to regulate the multiple stress responses and the development.So the issue about isolation and characterization of regulators to control plant stress response and development is essential.In this project,we identified 38 cotton genes putatively responsive to multiple abiotic stresses by data-mining and expression pattern analysis.And three cotton genes among them were isolated and characterized.The main results are as follows:1.A total of 826 Arabidopsis genes responsive to salt or cold stress were identified from STIFDB and GENEVESTIGATOR V3.According to the annotations corresponding to the 826 Arabidopsis genes,38 homologous EST from cotton were selected and their expression patterns responsive to ABA,salt and cold stress were confirmed using qRT-PCR.The results showed that 37 genes were regulated by ABA or salt or cold stress.Therefore,a successful method was provided for the selection of cotton multiple stress responsive genes.We isolated and characterized three genes among these 37 genes,which were regulated by ABA and salt and cold stress.2.We isolated a cotton NAC transcription factor gene,GhATAF1,which was not only up-regulated by ABA,cold and salt stresses,but also be highly induced by MeJA,SA and Verticillium dahliae inoculation.GhATAF1 was localized in the nucleus and possessed transactivation activity.Over-expression of GhATAF1 enhanced cotton plant tolerance to salt stress by enhancing the expression of various stress-related genes,including the ABA response gene GhABI4;the transporter gene GhHKT1,involved in Na+/K+ homeostasis;and several stress-response genes(GhAVP1,GhRD22,GhDREB2 A,GhLEA3 and GhLEA6).Additionally,over-expressing GhATAF1 increased cotton plant susceptibility to the fungal pathogens V.dahliae and Botrytis cinerea,coupled with the suppression of JA-mediated signaling and the activation of SA-mediated signaling.Our results suggested that GhATAF1 positively regulates the salinity tolerance and negatively regulates the resistance to V.dahliae by coordinating the photyhormone signaling networks in cotton.3.A cotton jasmoante ZIM-domain protein,GhJAZ2,was isolated.This gene was up-regulated by ABA,MeJA,SA,cold,salt stresses and V.dahliae inoculation.GhJAZ2 was localized in the nucleus and interacted with GhCOI1,and MeJA treatment induces its proteasome degradation.Gh JAZ2 plays as a repressor in the JA signaling in cotton,negatively regulates the JA-inducced anthocyanin accumulation and the JA inhibition of growth.GhJAZ2 interacted with GhJAZ12 to form a heteromeric complex,directly represses the transcriptional activity of GhMYC2-like,and negatively regulates the resistance to pathogen fungi V.dahliae and herbivory insect in cotton.And GhJAZ2 promotes flowering and seed germination in cotton,it might be mediated through the interaction of GhJAZ2-GhDELLAs.We also found that GhJAZ2 negatively regulates the size of cotton seeds and the angel of sympodial branch by unkown mechanisms.4.We isolated and characterized a cotton HD-ZIP protein,GhHB12,which was a pleiotropic gene regulating the abiotic and biotic stress responses and the growth and development progresses.GhHB12 predominantly expresses in roots and axillary buds,was up-regulated by ABA,MeJA,SA,cold,salt stresses and V.dahliae inoculation.GhHB12 decreases the drought and slat tolerance by negatively regulating ABA signaling;GhHB12 increases the susceptibility to fungal pathogens and herbivory insect through supressing JA signaling.GhHB12 interacts with GhTCP1,negatively regulates the IAA biosynthesis and signaling,reduces the apical dominance and plant height,and promotes the vegetative branch outgrowth in cotton.GhHB12 suppresses the expression of GhFT,GhSOC1,and GhFUL,inhibits the cotton fruit branch development and delays flowering time by mediating the MiR157-SPLs pathway.We also found that GhHB12 was regulated by circadian clock and photoperiodic,and the GhHB12-mediated development were rescued under the short-day condition(the winter in SanYa,Hai Nan).In summary,these reulsts suggested that it is effective in large-scale indentification of cotton genes involved in multiple stresses by data-mining and expression pattern analysis,and our reulsts serve as a foundation for analyses of the physiological mechanisms underlying cotton responsive to multiple stresses.