Type IV Fimbrial Biogenesis Is Required for Protease Secretion and Natural Transformation inDichelobacter nodosus

Abstract

The objective of this study was to develop an understanding of the molecular mechanisms by which type IV fimbrial biogenesis, natural transformation, and protease secretion are linked in the ovine foot rot pathogen, Dichelobacter nodosus. We have shown that like the D. nodosus fimbrial subunit FimA, the pilin-like protein PilE and the FimN, FimO, and FimP proteins, which are homologs of PilB, PilC, and PilD from Pseudomonas aeruginosa, are essential for fimbrial biogenesis and natural transformation, indicating that transformation requires an intact type IV fimbrial apparatus. The results also showed that extracellular protease secretion in the fimN, fimO, fimP, and pilE mutants was significantly reduced, which represents the first time that PilB, PilC, and PilE homologs have been shown to be required for the secretion of unrelated extracellular proteins in a type IV fimbriate bacterium. Quantitative real-time PCR analysis of the three extracellular protease genes aprV2, aprV5, and bprV showed that the effects on protease secretion were not mediated at the transcriptional level. Bioinformatic analysis did not identify a classical type II secretion system, and the putative fimbrial biogenesis gene pilQ was the only outer membrane secretin gene identified. Based on these results, it is postulated that in D. nodosus, protease secretion occurs by a type II secretion-related process that directly involves components of the type IV fimbrial biogenesis machinery, which represents the only type II secretion system encoded by the small genome of this highly evolved pathogen.

Type IV Pilin Proteins: Versatile Molecular Modules

“…Current models of T2S function that invoke a pistonlike action pushing substrate proteins through the secretin were informed by models of the extension and retraction of T4P and by studies showing that T2S pseudopilins could form pilus-like fibers under specific conditions (128,190,349,400). Furthermore, there are several examples of proteins that are secreted by T4P systems, including the TcpF colonization factor of V. cholerae (226), proteases of D. nodosus (167), and virulence factors of Francisella tularensis (19,163), showing that the systems are functionally similar.…”

Secretion of TcpF by the Vibrio cholerae Toxin-Coregulated Pilus Biogenesis Apparatus Requires an N-Terminal Determinant

“…Several proteins which have a Sec recognition signal peptide are thought to be secreted across the cytoplasmic membrane in a Sec-dependent manner and then across the outer membrane in a T4P biogenesis apparatusmediated manner: the proteases AprV2, AprV5, and BprV in D. nodosus (29,30) and PepO in F. tularensis (27,28). As seen in Table 2, the N terminus of the mature form of these proteins contains a YXS motif near the Sec cleavage site.…”

“…Thus, it is not surprising that T4P biogenesis apparatuses have been shown to also mediate protein secretion in Myxococcus xanthus (13), Francisella tularensis (27,28), Dichelobacter nodosus (29,30), and V. cholerae (31,32). In these systems the pilin is required for protein secretion, similar to how pseudopilins are required for protein secretion by T2SS (9,19,22), despite the fact that structures of the pilus show no pore for a protein to exit through in either type IVa (11,33) or type IVb (10) pili.…”

Footrot in Clawed and Hoofed Animals: Symptoms, Causes and Treatments

“…Apart from the fimbrial subunit protein, several pilin-like proteins, of which PilE has been described, play a role in the assembling of fimbriae (111). Mutation in the pilE gene leads to a significant decrease in the number of fimbriae, although the protein (Fim A) is produced at almost the same level as in the wild-type strain (49).…”

“…All proteins (FimA, PilE, FimN, FimO and FimP) are required for extracellular protease secretion. Mutation in any one of the genes encoding these proteins leads to a drastic decrease in protease activity (49).…”