Characterization of cDNAs encoding adhesin proteins involved in trichomonas vaginalis cytoadherence

Abstract

Trichomonas vaginalis cytoadherence is mediated by four adhesins (AP65, AP51, AP33 and AP23). Adhesin gene expression was previously shown to be up-regulated by iron. Therefore, a cDNA library was constructed from mRNA of T. vaginalis grown in a high-iron medium. Ten cDNA clones (three for AP65, one for AP51, and six for AP33) were recognized with polyclonal antiserum or monoclonal antibody raised against these adhesins. No cross-hybridization among cDNAs of the three adhesins was observed in Southern analysis, confirming restriction mapping analysis of representative cDNAs. Southern analysis probed with representative cDNAs of the adhesins indicated multiple copies for each of the three adhesin genes. Northern analysis showed transcripts of 1.8 kilobases (kb), 1.4 kb, and 0.9 kb for AP65, AP51 and AP33, respectively. Consistent with adhesin expression, mRNAs for these adhesins were detected only when parasites were grown in high-iron conditions. Specific antibodies eluted from E. coli expressing recombinant proteins reacted only with the respective parasite adhesins. Recombinant proteins bind to fixed HeLa cells and competed with radiolabeled trichomonad adhesins for binding. Although proteinase activity is required for cytoadherence, recombinant proteins show no detectable proteinase activity. These data show that recombinant proteins from these clones exhibited characteristics of the trichomonad adhesins.