State Key Laboratory of Crop Stress Biology for Arid Areas, China-Australia Joint Research Centre for Abiotic and Biotic Stress Management, College of Plant Protection, Northwest A&F University, Shaanxi, Yangling 712100, China.
Institut für Phytomedizin, Universität Hohenheim, 70599 Stuttgart, Germany.
Mol Plant Pathol. 2018 Jul;19(7):1639-1651. doi: 10.1111/mpp.12645. Epub 2018 Feb 13.
Velvet protein family members are important fungal-specific regulators which are involved in conidial development, secondary metabolism and virulence. To gain a broader insight into the physiological functions of the velvet protein family of Valsa mali, which causes a highly destructive canker disease on apple, we conducted a functional analysis of two velvet protein family members (VmVeA and VmVelB) via a gene replacement strategy. Deletion mutants of VmVeA and VmVelB showed increased melanin production, conidiation and sensitivity to abiotic stresses, but exhibited reduced virulence on detached apple leaves and twigs. Further studies demonstrated that the regulation of conidiation by VmVeA and VmVelB was positively correlated with the melanin synthesis transcription factor VmCmr1. More importantly, transcript levels of pectinase genes were shown to be decreased in deletion mutants compared with those of the wild-type during infection. However, the expression of other cell wall-degrading enzyme genes, including cellulase, hemi-cellulase and ligninase genes, was not affected in the deletion mutants. Furthermore, the determination of pectinase activity and immunogold labelling of pectin demonstrated that the capacity for pectin degradation was attenuated as a result of deletions of VmVeA and VmVelB. Finally, the interaction of VmVeA with VmVelB was identified through co-immunoprecipitation assays. VmVeA and VmVelB play critical roles in conidiation and virulence, probably via the regulation of the melanin synthesis transcription factor VmCmr1 and their effect on pectinase gene expression in V. mali, respectively.
天鹅绒蛋白家族成员是真菌特异性调控因子，参与分生孢子发育、次生代谢和毒力。为了更深入地了解引起苹果毁灭性溃疡病的苹果腐烂病菌天鹅绒蛋白家族的生理功能，我们通过基因替换策略对两个天鹅绒蛋白家族成员（VmVeA 和 VmVelB）进行了功能分析。VmVeA 和 VmVelB 的缺失突变体表现出黑色素产量增加、分生孢子形成增加和对非生物胁迫的敏感性增加，但在离体苹果叶片和小枝上的毒力降低。进一步的研究表明，VmVeA 和 VmVelB 对分生孢子形成的调控与黑色素合成转录因子 VmCmr1 呈正相关。更重要的是，与野生型相比，缺失突变体在感染过程中果胶酶基因的转录水平降低。然而，细胞壁降解酶基因，包括纤维素酶、半纤维素酶和木质素酶基因的表达在缺失突变体中不受影响。此外，果胶酶活性的测定和果胶的免疫金标记表明，由于 VmVeA 和 VmVelB 的缺失，果胶的降解能力减弱。最后，通过共免疫沉淀实验鉴定了 VmVeA 与 VmVelB 的相互作用。VmVeA 和 VmVelB 在分生孢子形成和毒力中发挥关键作用，可能分别通过调节黑色素合成转录因子 VmCmr1 和影响 V. mali 中的果胶酶基因表达来实现。
