Functional Analysis Of Cotton Genes,GhWRKY6-like And GhMYB108-like Under Abiotic Stresses

With the climate changes,drought and salt stresses are the serious issues related to crops to be solved.Transcriptional factors,i.e.,WRKY and MYB are major transcriptional regulators of plant signalling pathways involved in response to biotic and abiotic stresses such as drought,salt and osmotic stresses.GhWRKY6-like and GhMYB108-like genes were functionally characterized against salt and drought stresses in cotton and Arabidopsis.GhWRKY6-like consists of 5 exons and 4 introns,the open reading frame(ORF)contained 1653 base pairs and encoded a polypeptide of 550 amino acids with an isoelectric point(p I)of 6.95 and a molecular weight(Mw)of 59.6 k Da,respectively.While,GhMYB108-like consists of 1 exon and 807 base pairs ORF encoded a protein of268 amino acids with an p I of 5.51 and Mw of 30.3 k Da respectively.Both,GhWRKY6-like and GhMYB108-like proteins are nuclear-localized.Expression of GhWRKY6-like was remarkably induced by salt,poly ethylene glycol(PEG)and abscisic acid(ABA)treatments,while GhMYB108-like was significantly induced by PEG and salt treatments.To further investigate the functions of GhWRKY6-like and GhMYB108-like,overexpression construct was developed and transformed into Arabidopsis thaliana,while the GhMYB108-like was also transformed into cotton(Gossypium hirsutum cv.Jin668).In addition,GhWRKY6-like and GhMYB108-like genes were silenced by using virus-induced gene silencing(VIGS)technology.Our findings showed that the overexpressing GhWRKY6-like significantly improved the germination rate and root length of transgenic lines than wild type(WT)plants under salt,mannitol and ABA treatments.Additionally,the overexpressing lines showed greater salt tolerance than WT plants in soil.Moreover,overexpressing plants accumulated less H2O2 and malondialdehyde(MDA),while higher proline content,superoxide dismutase(SOD)and peroxidase(POD)activities were detected under salt stress.In contrast,GhWRKY6-like VIGS plants showed enhanced sensitivity compared to WT plants during salt and drought stresses.Additionally,expression analysis of stress responsive genes in overexpressing GhWRKY6-like Arabidopsis revealed that there was increased expression of genes involved in the ABA signalling pathway(At ABF4,At ABI5,At MYC2)and osmotic stress(At SOS2,At RD29 a,At RD29b).On the other side,root growth was significantly repressed in overexpressing GhMYB108-like transgenic plants compared to WT under mannitol and salt treatments.In addition,transgenic lines were found sensitive compared to WT plants under salt and drought stresses.Moreover,H2O2 and MDA content were accumulated higher in transgenic plants,while proline content,SOD and POD activities were detected lower in transgenic plants compared to WT.The expression of salt-and/or drought-induced marker genes(GhNHX1,GhRD26,GhSOS3,GhMYB5 and GhWRKY34)were remarkably suppressed in GhMYB108-like-overexpressing plants than WT under salt or drought stress.Our results revealed that overexpressing GhWRKY6-like enhanced salt tolerance in Arabidopsis by scavenging reactive oxygen species(ROS)and regulating the ABA signalling pathway.While,overexpressing GhMYB108-like enhanced salt and drought sensitivity in transgenic cotton plants by modulating ROS.Overall,our findings suggests that GhWRKY6-like plays a positive role in salt tolerance,while GhMYB108-like plays a negative role in response to salt and drought stresses.