SummaryThe insecticidal toxin complexes (Tcs) are produced by several Enterobacteriaceae associated with insects, such as Photorhabdus luminescens , Serratia entomophila and Xenorhabdus nematophilus. Genome sequences revealed tc -like genes in Yersinia spp., but insecticidal activity of this genus associated with the toxins has not been described. Through the search for genes upregulated at low growth temperatures in Yersinia enterocolitica strain W22703, a genomic island of 19 kb termed tc -PAI Ye with homologues of the toxin genes tcaA , tcaB , tcaC and tccC was identified. Southern blot and polymerase chain reaction (PCR) analysis of 34 strains demonstrated that the tc -PAI Ye is present in biovars 2, 3 and 4, but neither in biovars 1A and 1B, nor in five Yersinia species apathogenic in humans. Using the luxCDABE operon as reporter, the expression of the toxin genes was shown to be completely repressed in cells cultured at 37 °°°° C, and to increase by 4.6 orders of magnitude when the growth temperature was decreased gradually to 10 °°°° C. These data provide the first indication that temperature is a critical parameter for induction or repression of tc gene transcription. Whole-cell extracts of Y. enterocolitica strain W22703 cultivated at 10 °°°° C, but not at 30 °°°° C, led to insect mortality when fed to Manduca sexta larvae, in contrast to an insertional tcaA mutant. Overall the results suggest that the tc -PAI Ye could play an important role in the transmission and survival of pathogenic Y. enterocolitica strains outside mammalian hosts.
Background: Toxin complex (Tc) proteins termed TcaABC, TcdAB, and TccABC with insecticidal activity are present in a variety of bacteria including the yersiniae.
To analyze the transcriptional response of Yersinia enterocolitica cells to prolonged growth at low temperature, a collection of luxCDABE transposon mutants was cultivated in parallel at optimal (30°C) and suboptimal (10°C) temperatures and screened for enhanced promoter activities during growth until entering stationary phase. Among 5,700 Y. enterocolitica mutants, 42 transcriptional units were identified with strongly enhanced or reduced promoter activity at 10°C compared to 30°C, and changes in their transcriptional levels over time were measured. Green fluorescent protein fusions to 10 promoter regions confirmed the data. The temporal order of induction of the temperature-responsive genes of Y. enterocolitica was deduced, starting with the expression of cold shock genes cspA and cspB and the elevated transcription of a glutamate-aspartate symporter. Subsequently, cold-adapted cells drastically up-regulated genes encoding environmental sensors and regulators, such as UhpABC, ArcA, and methyl-accepting chemotaxis protein I (MCPI). Among the most prominent cold-responsive elements that were transcriptionally induced during growth in early and middle exponential phase are the insecticidal toxin genes tcaA and tcaB, as well as genes involved in flagellar synthesis and chemotaxis. The expression pattern of the lateexponential-to early-stationary-growth phase is dominated by factors involved in biodegradative metabolism, namely, a histidine ammonia lyase, three enzymes responsible for uptake and utilization of glycogen, the urease complex, and a subtilisin-like protease. Double-knockout mutants and complementation studies demonstrate inhibitory effects of MCPI and UhpC on the expression of a putative hemolysin transporter. The data partially delineate the spectrum of gene expression of Y. enterocolitica at environmental temperatures, providing evidence that an as-yet-unknown insect phase is part of the life cycle of this human pathogen.
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