Transposition of the endogenous insertion sequence element IS1126 modulates gingipain expression in Porphyrmonas gingivalis

We have previously reported on a Tn4351-generated mutant of Porphyromonas g ingivalis (MSM-3) which expresses enhanced arginine-specific proteinase act ivity and does not utilize hemin or hemoglobin far growth (C. A Genco ct al ., Infect. Immun. 63:2459-2466, 1995). In the process of characterizing the genetic lesion in P. gingivalis MSM-3, we have determined that the endogen ous P. gingivalis insertion sequence element IS1126 is capable of transposi tion within P. gingivalis. We have also determined that IS1126 transpositio n modulates the transcription of the genes encoding the lysine-specific pro teinase, gingipain K (kgp) and the arginine-specific proteinase, gingipain R2 (rgpB). Sequence analysis of P. gingivalis MSM-3 revealed that Tn4351 ha d inserted 60 bp upstream of the P. gingivalis endogenous IS element IS1126 . Furthermore, P. gingivalis MSM-3 exhibited mo additional copies of IS1126 compared to the parental strain A7436. Examination of the first additional IS1126 element, IS1126(1), indicated that it has inserted into the putativ e promoter region of the P. gingivalis kgp gene. Analysis of total RNA extr acted from P. gingivalis MSM-3 demonstrated no detectable kgp transcript; l ikewise, P. gingivalis MSM-3 was devoid of lysine-specific proteinase activ ity The increased arginine-specific proteinase activity exhibited by P. gin givalis MSM-3 was demonstrated to correlate with an increase in the rgpA. a nd rgpB transcripts. The second additional IS1126 element, IS1126(2), was f ound to have inserted upstream of a newly identified gene, hmuR, which exhi bits homolog to a number of TonB-dependent genes involved in hemin and iron acquisition. Analysis of total RNA from P. gingivalis MSM-3 demonstrated t hat hmuR is transcribed, indicating that the insertion of IS1126 had not pr oduced a polar effect on hmuR transcription. The hemin-hemoglobin defect in P. gingivalis MSM-3 is proposed to result from the inactivation of Kgp, wh ich has recently been demonstrated to function in hemoglobin binding. Taken together, the results presented here demonstrate that the introduction of Tn4351 into the P. gingivalis chromosome has resulted in two previously und ocumented phenomena in P. gingivalis: (i) the transposition of the endogeno us insertion sequence element IS1126 and (ii) the modulation of gingipain t ranscription and translation as a result of IS1126 transposition.