The pnd gene in E. coli plasmid R16: nucleotide sequence and gene expression leading to cell Mg2+ release and stable...

The pnd gene promotes the degradation of stable RNA in the presence of rifampicin at 42 degrees C, but is repressed during normal growth (Ohnishi, Y. and Akimoto, S. (1980) J. Bacteriol. 144, 833-835). We have determined the sequence of a third srnB-pnd-type gene, and have analyzed the effects of its expression from an inducible promoter. The nucleotide sequence of the pnd gene of the R16 plasmid exhibits an open reading frame for a polypeptide with 50 amino-acid residues, with high sequence homology to the pnd gene of a plasmid (R483) of a different incompatibility group. A possible base-paired stem and loop structure, which may participate in the regulation of gene expression, was detected between the promoter and the initiation codon, analogous to that in two comparable genes, srnB in the F and pnd in the R483 plasmid. When bacterial cells containing a lac-pnd fusion plasmid were incubated with a lac inducer at 30 degrees C, magnesium was released from the cells in bulk, and spheroplasts of the cells lysed even in hypertonic solution. Furthermore, when Mg2+ efflux was inhibited in the medium containing 5 mM Mg2+ or in Tris-HCl buffer, the degradation of stable RNA at 42 degrees C was inhibited. These results suggest that expression of the pnd gene effects a release of cellular magnesium by a membrane alterations, resulting in the stable RNA degradation at a higher temperature.