Istituto di Virologia Vegetale, CNR, Torino, Italy.
Phytopathology. 2010 Oct;100(10):1100-10. doi: 10.1094/PHYTO-02-10-0044.
ABSTRACT The role of Cpkk1, a mitogen-activated protein kinase from Cryphonectria parasitica, was investigated by generating a number of mutant strains that overexpress, under the control of the cryparin promoter, both the wild-type protein and its allele with an extensive deletion in the catalytic domain. Furthermore, a hairpin construct was built and expressed to cause specific silencing of Cpkk1 mRNA transcripts. Specific mRNA silencing or overexpression was confirmed on both Northern and Western blot analysis. Selected C. parasitica strains with Cpkk1 either silenced or overexpressed were evaluated for their biological characteristics, including virulence on European chestnut, growth on different substrates, conidial sporulation, and resistance to cell-wall-degrading enzymes. Silencing of Cpkk1 and the overexpression of a defective Cpkk1 correlated with a marked reduction in virulence on 3-year-old chestnut trees, with no statistically significant effect on fungal growth in the various conditions tested.
摘要 本研究通过生成多个突变株来调查 Cryphonectria parasitica 丝氨酸/苏氨酸蛋白激酶 Cpkk1 的作用，这些突变株在 cryparin 启动子的控制下过表达野生型蛋白及其催化结构域广泛缺失的等位基因。此外，构建了发夹结构并进行表达，以特异性沉默 Cpkk1 mRNA 转录本。通过 Northern 和 Western blot 分析证实了特定的 mRNA 沉默或过表达。对 Cpkk1 沉默或过表达的选定 C. parasitica 菌株进行了生物学特性评估，包括对欧洲栗的毒力、在不同基质上的生长、分生孢子产孢和对细胞壁降解酶的抗性。Cpkk1 的沉默和缺陷型 Cpkk1 的过表达与在 3 年生栗树上毒力的显著降低相关，但对各种测试条件下真菌生长的影响没有统计学意义。
