A temporal gene expression map of Chrysanthemum leaves infected with Alternaria alternata reveals different stages of defense mechanisms

Chrysanthemum(Chrysanthemum morifolium)black spot disease(CBS)poses a major threat to Chrysanthemum cultivation owing to suitable climate conditions and current lack of resistant cultivars for greenhouse cultivation.In this study,we identified a number of genes that respond to Alternaria alternata infection in resistant and susceptible Chrysanthemum cultivars.Based on RNA sequencing technology and a weighted gene coexpression network analysis(WGCNA),we constructed a model to elucidate the response of Chrysanthemum leaves to A.alternata infection at different stages and compared the mapped response of the resistant cultivar‘Jinba’to that of the susceptible cultivar‘Zaoyihong’.In the early stage of infection,when lesions had not yet formed,abscisic acid(ABA),salicylic acid(SA)and EDS1-mediated resistance played important roles in the Chrysanthemum defense system.With the formation of necrotic lesions,ethylene(ET)metabolism and the Ca^(2+)signal transduction pathway strongly responded to A.alternata infection.During the late stage,when necrotic lesions continued to expand,members of the multidrug and toxic compound extrusion(MATE)gene family were highly expressed,and their products may be involved in defense against A.alternata invasion by exporting toxins produced by the pathogen,which plays important roles in the pathogenicity of A.alternata.Furthermore,the function of hub genes was verified by qPCR and transgenic assays.The identification of hub genes at different stages,the comparison of hub genes between the two cultivars and the highly expressed genes in the resistant cultivar‘Jinba’provide a theoretical basis for breeding cultivars resistant to CBS.

CmMLO17 and its partner CmKIC potentially support Alternaria alternata growth in Chrysanthemum morifolium

The Mildew Resistance Locus O(MLO)gene family has been investigated in many species.However,there are few studies on chrysanthemum MLO genes.We report in this study that CmMLO17 in Chrysanthemum morifolium was upregulated after Alternaria alternata infection.Silencing of CmMLO17 by artificial microRNA resulted in reduced susceptibility of chrysanthemum to A.alternata infection.Genes in the abscisic acid(ABA)and Ca 2+signaling pathways were upregulated in the CmMLO17-silenced line R20 compared to the wild-type plants.We speculated that CmMLO17-silenced plants had a faster and stronger defense response that was mediated by the ABA and Ca 2+signaling pathways,resulting in reduced susceptibility of chrysanthemum to A.alternata infection.In addition,a candidate gene,CmKIC,that may interact with CmMLO17 was discovered by the yeast two-hybrid assay.The interaction between CmMLO17 and CmKIC was confirmed using the yeast two-hybrid assay and bimolecular fluorescence complementation(BiFC)analysis.CmMLO17 and CmKIC were both located on the plasma membrane,and CmKIC was also located on the nucleus.CmKIC overexpression increased the susceptibility of chrysanthemum to A.alternata,whereas CmKIC silencing resulted in reduced susceptibility.Therefore,CmMLO17 and CmKIC may work together in C.morifolium to support the growth of A.alternata.The results of this study will provide insight into the potential function of MLO and improve the understanding of plant defense responses to necrotrophic pathogens.