PM-Toxin A

Southern Corn Leaf Blight, one of the worst plant disease epidemics in modern history, was caused by Cochliobolus heterostrophus race T, which produces T-toxin, a determinant of high virulence to maize carrying Texas male sterile cytoplasm. The genetics of T-toxin production is complex and the evolutionary origin of associated genes is uncertain. It is known that ability to produce T-toxin requires three genes encoded at two unlinked loci, Tox1A and Tox1B, which map to the breakpoints of a reciprocal translocation. DNA associated with Tox1A and Tox1B sums to about 1.2 Mb of A+T rich, repeated DNA that is not found in less virulent race O or other Cochliobolus species. Here, we describe identification and targeted deletion of six additional genes, three mapping to Tox1A and three to Tox1B. Mutant screens indicate that all six genes are involved in T-toxin production and high virulence to maize. The nine known Tox1 genes encode two polyketide synthases (PKS), one decarboxylase, five dehydrogenases, and one unknown protein. Only two have a similar phylogenetic profile. To trace evolutionary history of one of the core PKS, DNA from more than 100 Dothideomycete species were screened for homologs. An ortholog (60% identity) was confirmed in Didymella zeae-maydis, which produces PM-toxin, a polyketide of similar structure and biological specificity as T-toxin. Only one additional Dothideomycete species, the dung ascomycete Delitschia winteri harbored a paralog. The unresolved evolutionary history and distinctive gene signature of the PKS (fast-evolving, discontinuous taxonomic distribution) leaves open the question of lateral or vertical transmission.

alpha-Latrotoxin (alpha-LT) potently enhances both "spontaneous" and "depolarization-evoked" quantal secretion from neurons. Here we have used the patch-clamped rat adrenal chromaffin cell to examine simultaneously the effects of alpha-LT on membrane current or voltage, cytosolic Ca, and membrane capacitance, the latter used as an assay for exocytosis. In chromaffin cells exposed to toxin concentrations of >100 pM, the development of large conductance, Ca-permeable ion channels, accompanied by a rise in cytosolic Ca to levels near 1 microM, precedes the initiation of spontaneous exocytosis. These channels appear to be induced de novo, because they occur concurrently with massive reduction or pharmacological block of voltage-dependent Na and Ca currents. However, enhancement of depolarization-evoked release, seen in many cells at <50 pM toxin, often occurs in the absence of a rise in background cytosolic Ca or de novo channel activity. These results favor Ca entry through toxin-induced channels underlying initiation of spontaneous release and direct modulation of the secretory machinery by the toxin-bound receptor contributing to enhancement of depolarization-evoked secretion as well as spontaneous release.

The Southern corn leaf blight (SCLB) epidemic of 1970 devastated fields of T-cytoplasm corn planted in monoculture throughout the eastern United States. The epidemic was driven by race T, a previously unseen race of Cochliobolus heterostrophus. A second fungus, Phyllosticta zeae-maydis, with the same biological specificity, appeared coincidentally. Race T produces T-toxin, while Phyllosticta zeae-maydis produces PM-toxin, both host-selective polyketide toxins necessary for supervirulence. The present abundance of genome sequences offers an opportunity to tackle the evolutionary origins of T- and PM- toxin biosynthetic genes, previously thought unique to these species. Using the C. heterostrophus genes as probes, we identified orthologs in six additional Dothideomycete and three Eurotiomycete species. In stark contrast to the genetically fragmented race T Tox1 locus that encodes these genes, all newly found Tox1-like genes in other species reside at a single collinear locus. This compact arrangement, phylogenetic analyses, comparisons of Tox1 protein tree topology to a species tree, and Tox1 gene characteristics suggest that the locus is ancient and that some species, including C. heterostrophus, gained Tox1 by horizontal gene transfer. C. heterostrophus and Phyllosticta zeae-maydis did not exchange Tox1 DNA at the time of the SCLB epidemic, but how they acquired Tox1 remains uncertain. The presence of additional genes in Tox1-like clusters of other species, although not in C. heterostrophus and Phyllosticta zeae-maydis, suggests that the metabolites produced differ from T- and PM-toxin.