CHARACTERIZATION OF HAPR, A POSITIVE REGULATOR OF THE VIBRIO-CHOLERAEHA PROTEASE GENE HAP, AND ITS IDENTIFICATION AS A FUNCTIONAL HOMOLOG OF THE VIBRIO-HARVEYI LUXR GENE
Mg. Jobling et Rk. Holmes, CHARACTERIZATION OF HAPR, A POSITIVE REGULATOR OF THE VIBRIO-CHOLERAEHA PROTEASE GENE HAP, AND ITS IDENTIFICATION AS A FUNCTIONAL HOMOLOG OF THE VIBRIO-HARVEYI LUXR GENE, Molecular microbiology, 26(5), 1997, pp. 1023-1034
The Vibrio cholerae HA/protease gene (hap) promoter is inactive in Esc
herichia coli. We cloned and sequenced the 0.7 kb hap promoter fragmen
t from strain 3083-2 and showed that hap is located immediately 3' of
ompW, encoding a minor outer membrane protein, A clone from a genomic
library of strain 3083-2 was isolated, which was required for activati
on of the hap promoter in E. coli, Expression from the hap promoter on
ly occurred late in the growth phase, A single complete open reading f
rame (ORF) designated HapR was identified on a 1.7 kb DNA fragment tha
t was required for activation, Allelic replacements showed that hapR w
as also essential for hap expression in V. cholerae, In El Tor, but no
t in classical biotypes of V. cholerae, hapR mutations also produced a
rugose colonial phenotype, HapR was shown to encode a 203-amino-acid
polypeptide with 71% identity to LuxR of V. harveyi, an essential posi
tive regulator of the lux operon that has no previously identified hom
ologues, The amino-terminal domain (residues 21-68) showed significant
homology to the TetR family of helix-turn-helix DNA-binding domains a
nd was 95% identical to the same domain of LuxR, HapR and LuxR activat
ed both the hap and the lux promoters at near wild-type levels, despit
e only limited homology in the promoter sequences (46% identity with 1
2 gaps over 420 bp). DNA sequences and ORFs 5' (but not 3') of the hap
R and luxR loci were homologous, suggesting a common origin for these
loci, and hapR-hybridizing sequences were found in other vibrios, We c
onclude that HapR is absolutely required for hap expression and that H
apR and LuxR form a new family of transcriptional activator proteins.