The Molecular Mechainsm Of MeCAMTA3 Mediated Cassava Disease Response Against Cassava Bacterial Blight

Cassava（Manihot esculenta Crantz）is one of the tropical crops.However,cassava bacterial blight（CBB）has severely disrupted the yield and quality of cassava production all over the world.Therefore,studying the mechanism of cassava disease resistance and the key genes in response to cassava disease resistance will be conducive to the application of genetic improvement technology in cassava disease resistance,so as to effectively improve the quality of cassava.As a second messenger,calcium(Ca²⁺)is widely involved in the regulation of various physiological processes.Calmodulin（Ca M）can sense the changes of Ca²⁺concentration and interact with Ca M binding protein（Ca MBP）to regulate plant growth,development and response to stresses.And calmodulin-binding transcription activator（CAMTA）is one member of Ca MBP.Herein,we isolated and identified six MeCAMTAs,and analysed their relative transcript levels in response to Xanthomonas axonopodis pv.manihotis（Xam）and subcellular localization.MeCAMTA3 was chosen for further analysed.And MeCAMTA3 was transiently transferred in cassava or silenced via virus-induced gene silencing（VIGS）system to test the function in cassava responds to immune reaction.Based on our experiments,we obtained the following results.1.Six MeCAMTAs of cassava were isolated and identified in this experiment.Phylogenetic tree showed that CAMTAs from cassava were closely related to those from Arabidopsis and rice.And the domains of MeCAMTAs were highly conserved.Subcellular localization results revealed that all six genes were localized in the nucleus.Quantitative real-time PCR showed that all these genes were commonly induced by Xam,especially MeCAMTA3.2.To investigate the transcriptional activation character,p GBKT7-MeCAMTA3was transformed into AH109 yeast cells,the results showed that MeCAMTA3 had strong transcriptional activity.And electrophoretic mobility shift assay（EMSA）evidencedthat MeCAMTA3 can directly bind to the v CGCGb motif.3.At silenced and overexpressed plants,MeCAMTA3 showed negatively regulates the resistanceto CBB.The MeCAMTA3-silenced plants showed higher disease resistance with higher oxygen species（ROS）level,endogenous salicylic acid（SA）level and the callose deposition than those in the control.4.RNA sequencing identified a series of genes regulated by MeCAMTA3,including ubiquitin-conjugating enzyme E2H（UBE2H）,calcium-dependent protein kinase 20（CDPK20）,probable WRKY transcription factor 70（WRKY）,serine/threonine-protein kinase（CTR1）,gibberellin A44 oxidase（GAO44）,auxin response factor 4（ARF4）,probable L-ascorbate peroxidase 6（APX6）and peroxidase A2-like（POD53）.And it was further confirmed that these genes were regulated by MeCAMTA3 by chromatin immunoprecipitation-PCR.In summary,MeCAMTA3 involved in the resistance to CBB through regulated the SA and ROS levels.The results of this experiment provided some help for the development of new cassava disease-resistant varieties.