Activation of Zoosporogenesis-Specific Genes in Phytophthora infestans Involves a 7-Nucleotide Promoter Motif and Cold-Induced Membrane Rigidity

Abstract

Infections of plants by the oomycete Phytophthora infestans typically result from zoospores, which develop from sporangia at cold temperatures. To help understand the relevant cold-induced signaling pathway, factors regulating the transcription of the zoosporogenesis-specific NIF (nuclear LIM-interactor-interacting factor) gene family were examined. Sequences required for inducing PinifC3 were identified by analyzing truncated and mutated promoters using the beta-glucuronidase reporter in stable transformants. A 7-nucleotide (nt) sequence located 139 bases upstream of the major transcription start point (GGACGAG) proved essential for the induction of PinifC3 when sporangia were shifted from ambient to cold temperatures. The motif, named the cold box, also conferred cold inducibility to a promoter normally activated only during sexual development. An identical motif was detected in the two other zoosporogenesis-specific NIF genes from P. infestans and three Phytophthora sojae orthologues, and a closely related sequence was found in Phytophthora ramorum orthologues. The 7-nt motif was also found in the promoters of other zoosporogenesis-induced genes. The presence of a cold box-interacting protein in nuclear extracts of P. infestans sporangia was demonstrated using electrophoretic mobility shift assays. Furthermore, zoospore release and cold box-regulated transcription were stimulated by the membrane rigidizer dimethyl sulfoxide and inhibited by the membrane fluidizer benzyl alcohol. The data therefore delineate a pathway in which sporangia perceive cold temperatures through membrane rigidity, which activates signals that drive both zoosporogenesis and cold-box-mediated transcription.