Characterization And Function Analysis Of Genes Encoding Calcineurin In Setosphaeria Turcica

Northern Corn Leaf Blight, caused by Exserohilum turcicum, is one of the most important diseases in corn planting area. In prevalent year heavy loss in corn plant field is often caused by Setosphaeria turcica. The growth, developoment and virulence of plant pathogenic fungi is regulated by extracellular signal transduction pathways, which inclute the Ca2+signal transduction pathway, the cAMP signal transduction pathway and the MAPK signal transduction pathway. Ca2+ pathway is majorly involve of calmodulin(CaM), phospholipase C(PLC), calcineurin(CaN). Calcineurin with one catalytic subunit CnA and one regulating subunit CnB, also called PP2B, is only the Ser/Thr Protein phosphatase regulated by Ca2+ and CaM complex in the Ca2+ signal transduction pathway. CnB with 4 Ca2+ binding sites, is stuctrurally similar to CaM. When CnB binding with 4 Ca2+ and Ca2+ CaM complex are bound to different sites on CnA, Calcineurin startes catalytic activity. CaN, a multifunctional signalase, not only mediates Ca2+ signal transduction pathway but also couple Ca2+ signals with many other signal transduction pathway by dephosphorylation, which co-ordinates cellular functions. CaN plays an important role in the regulation of ion balance, hyphal morphogenesis, cell wall integrity and pathogenicity of plant pathogenic fungi.In this paper, we obtained 26 strains CnA gene RNAi transforments and 42 strains CnB gene RNAi transforments which had been identified by the screening hygromycin B-resistance and PCR, Southern blot and semi-quantitative RT-PCR analysis of all transformants, by taking advantage of pSilent-1 plasmid, CnA and CnB RNAi vector were constructed, and transformed into protoplasts of Setosphaeria turcica. The properties of mutant strains cultured in media were described as follow: Observing from the front of colony, mutant strain A-2: gray, dense, thin,a tidy edge of colony, a little amount of aerial hyphae, green-brown in back colony; Mutant B-1: light brown in front colony, delicate, thin, a little amount of aerial hyphae, untidy edge of colony, brown in back colony; Under microscope slender, slightly transparent hyphae of mutant A-2 and slender, transparent hyphae of mutant B-1were observed; The melanin content of mutant hyphae can be decreased speculatively. Mutant A-2 and wild-type had similar growth rate of hyphae in media, while mutant B-1 had slower growth rate of hyphae than wild-type in media. The wild-type strain cultured in media produced conidia; while no conidia were not produced in mutant B-1 and A-2. Virulence of Ht-toxins and pathogenicity of mutant B-1 and A-2 strains on maize levels were weakened. Gene expression analysis showed that the expression of PKA and STK1 in mutant A-2 and the expression of STK2 in mutant B-1 were declined; and that while the expression of CAM was not changed significantly in mutant B-1 and A-2.