The plant disease resistance gene Asc‐1 prevents disruption of sphingolipid metabolism during AAL‐toxin‐induced programmed cell death

Abstract

These authors contributed equally to this work. SummaryThe nectrotrophic fungus Alternaria alternata f.sp. lycopersici infects tomato plants of the genotype asc/asc by utilizing a host-selective toxin, AAL-toxin, that kills the host cells by inducing programmed cell death. Asc-1 is homologous to genes found in most eukaryotes from yeast to humans, suggesting a conserved function. A yeast strain with deletions in the homologous genes LAG1 and LAC1 was functionally complemented by Asc-1, indicating that Asc-1 functions in an analogous manner to the yeast homologues. Examination of the yeast sphingolipids, which are almost absent in the lag1Dlac1D mutant, showed that Asc-1 was able to restore the synthesis of sphingolipids. We therefore examined the biosynthesis of sphingolipids in tomato by labeling leaf discs with L-[3-3 H]serine. In the absence of AAL-toxin, there was no detectable difference in sphingolipid labeling between leaf discs from Asc/Asc or asc/asc leaves. In the presence of pathologically significant concentrations of AAL-toxin however, asc/asc leaf discs showed severely reduced labeling of sphingolipids and increased label in dihydrosphingosine (DHS) and 3-ketodihydrosphingosine (3-KDHS). Leaf discs from Asc/Asc leaves responded to AAL-toxin treatment by incorporating label into different sphingolipid species. The effects of AAL-toxin on asc/asc leaflets could be partially blocked by the simultaneous application of AAL-toxin and myriocin. Leaf discs simultaneously treated with AAL-toxin and myriocin showed no incorporation of label into sphingolipids or long-chain bases as expected. These results indicate that the presence of Asc-1 is able to relieve an AAL-toxin-induced block on sphingolipid synthesis that would otherwise lead to programmed cell death.

An extensive microarray analysis of AAL-toxin-induced cell death in Arabidopsis thaliana brings new insights into the complexity of programmed cell death in plants

“…Results after the AAL toxin treatment are very reproducible with respect to timing of gene expression, morphological effects and cell death. The AAL toxin triggers PCD through interference with sphingolipid metabolism, although the exact mechanism of activating the cell death machinery remains elusive [3,6]. The role of sphingolipids in promoting cell proliferation or death is well recognized in animals [19].…”

“…The labelling, hybridization and data extraction were done at ServiceXS (The Netherlands) according to the instructions of Agilent Technologies. Northerns were performed with 6 mg of total RNA as described elsewhere [6]. The quality and quantity of the RNA was verified by staining the membranes with methylene blue.…”

“…Although the exact mechanisms of cell death activation and execution remain elusive, the presence of a disease resistance gene called Asc is known to prevent the perturbations in sphingolipid metabolism and cell death [6]. Asc is homologous to the yeast longevity assurance gene LAG1 [5].…”

“…sphingolipid biosynthesis, called Asc [5], as Asc can to a large extent prevent the disruptions in sphingolipid metabolism and the subsequent cell death [6]. To use the power of Arabidopsis thaliana genomics, our group has identified a knockout of the A. thaliana homologue of Asc that rendered the mutant plants sensitive to growth inhibition by the AAL toxin [J. E. Markham and J. Hille, unpublished results].…”

Mining Effector Proteins in Phytopathogenic Fungi

“…Oomycete effectors play dual role in disease and plant defense, fungal effector proteins were no exception. For example, victorin of Cochliobolus victoriae, NIP1 of R.secalis, and AAL toxin of Alternaria alternate played the role in toxin (Lorang et al, 2007;Navarre and Wolpert, 1999;Rohe et al, 1995;Spassieva et al, 2002;van't Slot and Knogge, 2002;Wang et al, 1996, Wolpert et al, 2002, but some studies showed NIP1 and ToxA also interacted with corresponding host-resistance or toxinsensitivity genes, which resulted in NIP1 and ToxA acted in the same way as the Avr genes (Schürch et al, 2004;Stukenbrock and McDonald, 2007). So, it is necessary to clarify fungal effector proteins incompatibly or compatibly interact with host receptors when they were transported into host cells.…”