Mycobacterium tuberculosisRv3194c efficiently facilitates mycobacterial-lung epithelial interaction through its HA-binding site

Mycobacterium tuberculosis adhesins are surface-exposed molecules that mediate pathogen-host interaction, a fundamental step towards host infection. Here we show that serine protease (Rv3194c) promotes mycobacterial infection to lung epithelial through its hyaluronic acid (HA)-binding site. Both enzyme-linked immunosorbent assay and surface plasmon resonance analysis revealed that Rv3194c bound to HA. Utilizing synthetic peptides, we next defined HA-binding site of 20 amino acids from 91 to 110 of Rv3194c (P91-110). Immunofluorescence assay and an FACScan showed that Rv3194c was interacted with A549 cells (human lung epithelial cells), and its interaction was abolished by the addition of hyaluronidase or P91-110. Experimental infection in Vitro revealed that Rv3194c participates in attachment of recombinant Mycobacterium smegmatis (Rv3194c/MS) to A549 cells, and P91-110 treatment of A549 cells almost inhibited Rv3194c/MS-A549 cells interaction. To provide in vivo evidence, we constructed a reporter strain of M. smegmatis expressed a derivative of the firefly luciferase that is shifted to red (FFlucRT) in combination with Rv3194c (Rv3194c+FFlucRT/MS) to infect the rodents and monitor the progression of the disease. Using bioluminescence imaging and bacterial counts in lung tissue confirmed that Rv3194c dramatically enhanced the persistence of M. smegmatis . In addition, treatment of intratracheal Rv3194c+FFlucRT/MS-infected mice with P91-110 significantly suppressed the growth of Rv3194c+FFlucRT/MS in vivo . Taken together, these results demonstrate that Rv3194c was identified as a HA-binding adhesin, and P91-110 as anti-adhesion agents has potential for therapeutic and prophylactic interventions in mycobacterial infection.