Promoter analysis of the sodium-responsive V-ATPase (ntp) operon in Enterococcus hirae

The Enterococcus hirae ntp operon encodes all subunits of the vacuolar-type ATPase (V-ATPase), which transports Na(+) or Li(+). This operon is expressed preferentially in response to Na(+), but not to Li(+). Deletion analysis of the ntp promoter region in plasmids indicated that the AT-tract between -198 and -132 is required for Na(+)-specific transcriptional regulation. In addition, lithium-tolerant (LTR) mutants were isolated in which functional V-ATPase levels were high even in Na(+)-depleted medium. Western blot and Northern blot experiments revealed an increase in basal Na(+)-independent transcription in one of the mutants (LTR1). The nucleotide sequences of the ntp promoter region of the LTR mutants showed mutational conversion of single base-pairs between positions -23 and +1. Na(+)-independent expression of a reporter gene linked to the ntp promoter in plasmids was elevated by base substitutions at -23 to +1, and promoter activity induced by these base substitutions was lost by deletion of the region between -198 and -132. These results suggest that the AT-tract between -198 and -132 is indispensable for transcription of the ntp operon.