Role of differential expression of streptococcal arginine deiminase in inhibition of fimA expression in Porphyromonas gingivalis

Abstract

Streptococcus cristatus ArcA inhibits production of a major adhesin, FimA, in Porphyromonas gingivalis, a primary periodontal pathogen. In this study, we demonstrate the differential expression of arcA in two streptococcal species. The expression level of arcA in streptococci appears to be controlled by both cis and trans elements.

FIG. 1.

Sequence alignment of promoter region of S. cristatus CC5A arcA gene (GenBank accession no. 884757) with S. gordonii DL1 arcA (GenBank accession no. AF534569). The boldfaced ATG is a potential start codon. TSS A (+1) of CC5A arcA is in the same position as the published TSS of S. gordonii DL1 (17). The transcriptional start sites, the potential −10 and −35 positions, and the potential start codons are in boldface. The potential catabolite response elements are shaded.

FIG. 2.

Western blot analysis of ArcA production with polyclonal antibodies to the CC5A ArcA. Shown are 14-h cultures of streptococcal strains grown in Trypticase peptone broth. Cell extracts (5 μg) from streptococcal cells were used. Lane 1, S. cristatus CC5A; lane 2, S. gordonii DL1; lane 3, S. cristatus ARCE (arcA mutant); lane 4, S. gordonii ARCE (arcA mutant); lane 5, S. cristatus cARCE (S. cristatus ARCE complemented with CC5A arcA gene); lane 6, S. cristatus dARCE (S. cristatus ARCE complemented with DL1 arcA gene); lane 7, S. gordonii dARCE (S. gordonii ARCE complemented with DL1 arcA gene); lane 8, S. gordonii cARCE (S. gordonii ARCE complemented with CC5A arcA gene).

FIG. 3.

Determination of the expression level of fimA in P. gingivalis in response to streptococcal strains. P. gingivalis UPF carrying a fimA promoter-lacZ fusion gene was tested for β-galactosidase activity in the presence or absence of the surface extracts (50 μg) isolated from the S. cristatus or S. gordonii strains. Standard errors are indicated (n = 3). Means with different letters are significantly different (P < 0.05; two-way analysis of variance and Student Newman-Keuls test), and means with the same letter are not significantly different.

FIG. 4.

Expression of arcR and ccpA in S. cristatus CC5A and S. gordonii DL1. Gene expression analysis was examined by using real-time PCR. The transcript level of each gene in S. gordonii DL1 is indicated relative to the expression level of these genes in S. cristatus CC5A as 1 U. The relative abundance of each indicated transcript was also normalized by 23S rRNA. The results presented are the average starting quantities of four independent experiments.