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Inverse PCR was used to amplify major cold shock protein (MCSP) gene families from a diverse range of bacteria, including the psychrotolerantYersinia enterocolitica , which was found to have two almost identical MCSP coding regions (cspA1 andcspA2 ) located approximately 300 bp apart. This tandem gene duplication was also found inY. pestis ,Y. pseudotuberculosis , andY. ruckeri but not in other bacteria. Analysis of the transcriptional regulation of this MCSP gene inY. enterocolitica , performed by using both reverse transcriptase-PCR and Northern blot assays, showed there to be two cold-inducible mRNA templates arising from this locus: a monocistronic template of approximately 450 bp (cspA1 ) and a bicistronic template of approximately 900 bp (cspA1/A2 ). The former may be due to a secondary structure betweencspA1 andcspA2 causing either 3′ degradation protection ofcspA1 or, more probably, partial termination aftercspA1 . Primer extension experiments identified a putative transcriptional start site (+1) which is flanked by a cold-box motif and promoter elements (−10 and −35) similar to those found inEscherichia coli cold-inducible MCSP genes. At 30°C, the level of both mRNA molecules was negligible; however, upon a temperature downshift to 10°C, transcription of the bicistronic mRNA was both substantial (300-fold increase) and immediate, with transcription of the monocistronic mRNA being approximately 10-fold less (30-fold increase) and significantly slower. The ratio of bicistronic to monocistronic mRNA changed with time after cold shock and was higher when cells were shocked to a lower temperature. High-resolution, two-dimensional protein gel electrophoresis showed that synthesis of the corresponding proteins, both CspA1 and CspA2, was apparent after only 10 min of cold shock from 30°C to 10°C. The data demonstrate an extraordinary capacity of the psychrotolerantY. enterocolitica to produce major cold shock proteins upon cold shock.

journal of bacteriology

Pathogenic <i>Yersinia</i> Species Carry a Novel, Cold-Inducible Major Cold Shock Protein Tandem Gene Duplication Producing both Bicistronic and Monocistronic mRNA

Pathogenic Yersinia Species Carry a Novel, Cold-Inducible Major Cold Shock Protein Tandem Gene Duplication Producing both Bicistronic and Monocistronic mRNA