Yi Zhang,Kedong Xu,Deshui Yu,Zhihui Liu,Chunfeng Peng,Xiaoli Li,Ju Zhang,Yinghui Dong,Yazhen Zhang,Pan Tian,Tiancai Guo,Chengwei Li
Effector proteins secreted by plant pathogens play important roles in promoting colonization.Blumeriaeffector candidate (BEC) 1019, a highly conserved metalloprotease ofBlumeria graminisf. sp.hordei(Bgh), is essential for fungal haustorium formation, and silencingBEC1019significantly reducesBghvirulence. In this study, we found thatBEC1019homologs inB. graminisf. sp.tritici(Bgt) andGaeumannomyces graminisvar.tritici(Ggt) have complete sequence identity with those inBgh, prompting us to investigate their functions. Transcript levels ofBEC1019were abundantly induced concomitant with haustorium formation inBgtand necrosis development inGgt-infected plants.BEC1019overexpression considerably increased wheat susceptibility toBgtandGgt, whereas silencing this gene using host-induced gene silencing significantly enhanced wheat resistance toBgtandGgt, which was associated with hydrogen peroxide accumulation, cell death, and pathogenesis-related gene expression. Additionally, we found that the full and partial sequences ofBEC1019can trigger cell death inNicotiana benthamianaleaves. These results indicate thatBgtandGgtcan utilizeBEC1019as a virulence effector to promote plant colonization, and thus these genes represent promising new targets in breeding wheat cultivars with broad-spectrum resistance.Keywords:effector protein;Blumeria graminisf. sp.tritici;Gaeumannomyces graminisvar.tritici;haustorium formation;necrosis development;cell death
Carabrone, a botanical bicyclic sesquiterpenic lactone, has broad-spectrum antifungal activity and is particularly efficient against the devastating phytopathogen Gaeumannomyces graminis var. tritici (Ggt). The antifungal mechanism of carabrone against Ggt, however, remains unclear. The main objective of this study was to investigate the subcellular localization of carabrone in Ggt to gain a better understanding of its mechanism of action. When Ggt was exposed to carabrone (EC50 value of 28.45 μg/mL) for 7 days, a decline in mitochondrial concentration together with some obvious alternations in mitochondrial structure, including hazy outlines, medullary transitions, excess accumulation of unclear settlings, and vacuolar degeneration, were observed, indicating that carbrone may act on the mitochondria directly. A fluorescent conjugate (TTY) was thus designed and synthesized as a surrogate of carabrone that possessed comparable antifungal activity against Ggt (EC50 of 33.68 μg/mL). Additionally, a polyclonal antibody specific to carabrone and with a high titer (256 000) was also prepared by immunizing mice. Subsequently, two imaging techniques, the use of the fluorescent conjugate (FC) and immunofluorescence (IF), were applied to determine the subcellular localization of carabrone. Both FC and IF fluorescent signals demonstrated its mitochondrial localization with a Pearson’s coefficient of 0.83 for FC and 0.86 for IF. These results imply that carabrone exerts its antifungal activity against Ggt by interfering with mitochondrial function.
危险品化学品经营许可证（带存储）