PROMOTER STRENGTH INFLUENCES PHASE VARIATION OF NEISSERIAL OPA GENES

The opa multigene family of Neisseria gonorrhoeae encodes 11 related o uter-membrane proteins which phase vary in vitro and in vivo. Illegiti mate recombination within direct pentameric DNA repeats, encoding the signal-peptide region of pre-Opas, leads to switches in expression sta tes. Despite the conserved nature of the variation mechanism, specific genes are expressed at high frequencies in the transition from Opa- t o Opa+. The genes which are expressed at elevated frequencies differ f rom the rest of the family with respect to promoter structure, based o n sequence comparisons between the cpa genes of strain MS11mk. We have analysed transcription of the cpa gene family of N. gonorrhoeae MS11m k, focussing on the different promoters found among the 11 genes to de termine whether increased levels of expression are associated with inc reased phase-variation rates. Primer extension and Northern blotting w as used to assess the levels of transcription of three representative opa genes (opaA, B and C) in 'on' and 'off' states. Full-length opa mR NA was detected primarily in strains expressing the homologous gene. T runcated opa mRNA was constitutively expressed from all cpa genes rega rdless of their expression state. Quantitative comparisons in N. gonor rhoeae were complicated by the simultaneous expression of all 11 genes and the cross-reactivity of mRNA probes. Expression levels from the i ndividual promoters were therefore assessed by creating transcriptiona l and translational lacZ fusions to each of the representative cpa pro moters which lacked the DNA repeats responsible for variation. The exp ression levels were compared to the phase-variation rates of translati onal opa::phoA fusions containing the same promoters in addition to th e corresponding coding repeat regions. A strong correlation was found between expression levels from the different promoters and the variati on rates at which 'on' variants appeared from an 'off' population (i.e . opaA > opaB > opaC). These results provide an explanation for the fa voured expression of specific Opa proteins and indicate that expressio n of cpa genes may be regulated at the level of transcription.