Interactions of the Type III Secretion Pathway Proteins LcrV and LcrG from Yersinia pestis Are Mediated by Coiled-Coil Domains

Abstract

The type III secretion system is used by pathogenic Yersinia to translocate virulence factors into the host cell. A key component is the multifunctional LcrV protein, which is present on the bacterial surface prior to host cell contact and up-regulates translocation by blocking the repressive action of the LcrG protein on the cytosolic side of the secretion apparatus. The functions of LcrV are proposed to involve self-interactions (multimerization) and interactions with other proteins including LcrG. Coiled-coil motifs predicted to be present are thought to play a role in mediating these proteinprotein interactions. We have purified recombinant LcrV, LcrG, and site-directed mutants of LcrV and demonstrated the structural integrity of these proteins using circular dichroism spectroscopy. We show that LcrV interacts both with itself and with LcrG and have obtained micromolar and nanomolar affinities for these interactions, respectively. The effects of LcrV mutations upon LcrG binding suggest that coiled-coil interactions indeed play a significant role in complex formation. In addition, comparisons of secretion patterns of effector proteins in Yersinia, arising from wild type and mutants of LcrV, support the proposed role of LcrG in titration of LcrV in vivo but also suggest that other factors may be involved.

Diminished LcrV Secretion Attenuates Yersinia pseudotuberculosis Virulence

“…As a result, LcrG could be considered a potential chaperone for LcrV (23). Although the region of amino acids 108 to 125 is not known to bind LcrG (22), it might still aid in generating a secondary LcrG chaperone-dependent secretion signal for LcrV secretion.…”

Oral Vaccination withSalmonellaSimultaneously ExpressingYersinia pestisF1 and V Antigens Protects against Bubonic and Pneumonic Plague

“…V-Ag is a membrane protein in Y. pestis (13,22), but where it is expressed within Salmonella is unknown. This can be further complicated when both F1-and V-Ags are coexpressed in Salmonella-(F1ϩV)Ags.…”

“…V-Ag is a known secreted protein that is present on Y. pestis's cell surface before host cell contact (22), and it is present in the outer membrane (13). To determine which cell fraction within Salmonella V-Ag localizes, Salmonella-(F1ϩV)Ags cells cultured from LB broth were harvested and separated into the following three subcellular components: spheroplastic fraction, periplasmic fraction, and outer membrane (including cell walls) fraction by conventional methods.…”

Interaction between Yersinia pestis and the Host Immune System

“…LcrV is a multifunctional protein that is involved in the formation of a tip complex at the tip of the T3SS needle. It activates the secretion and translocation of effectors by binding to the negative regulator LcrG (42) and works together with Yops B and D for delivering Yops into eukaryotic cells (24). It is also exposed on the bacterial cell surface prior to contact with mammalian cells and may play a role in cell-cell adhesion (67).…”