What's the point of the type III secretion system needle?

Abstract

Recent work by several groups has significantly expanded our knowledge of the structure, regulation of assembly, and function of components of the extracellular portion of the type III secretion system (T3SS) of Gram-negative bacteria. This perspective presents a structure-informed analysis of functional data and discusses three nonmutually exclusive models of how a key aspect of T3SS biology, the sensing of host cells, may be performed.

Fig. 1.

Structures of needle proteins and their complexes. (A) Cartoons of monomeric needle proteins: the crystal structures of two conformations of MxiH from S. flexneri (form A, red; form B, blue) with the position of Tyr-60 at the bend highlighted in cyan [ref. and Protein Data Bank (PDB) ID code 2CA5], the NMR structures (well ordered regions only) of BsaL from Burkholderia pseudomallei (green) (ref. and PDB ID code 2G0U) and PrgI from Salmonella typhimurium (yellow) (ref. and PDB ID code 2JOW) and an overlay using the head region. (B) The Shigella T3SS needle (120 copies of the MxiH monomer as docked (PDB ID code 2V6l) into the EM reconstruction, EMD-1416) shown protruding from the EM reconstruction of the Shigella basal apparatus [gray surface (8) EMD-1422] with the inner and outer membranes illustrated (planes in sand). (Inset) A magnified region of the needle assembly shows the arrangement of needle subunits. (C) (Left) The crystal structure (ref. and PDB ID code 2UWJ) of a fragment of PscF from Pseudomonas aeruginosa (cyan) bound to the chaperones PscE (orange) and PscG (surface in light gray) overlaid with MxiH (transparent red) via the C terminus (residues 65–80 of MxiH) is shown. (Right) A portion of the Shigella needle showing a MxiH monomer (red) surrounded by adjacent MxiH molecules in the assembly (surface in light orange) is shown.

Fig. 2.

Needle tip-associated adaptor protein structures. (A) Cartoon representation of the four classes of adaptor proteins thus solved: Yersinia family (LcrV; PDB ID code 1R6F) (41); Shigella family (IpaD; PDB ID code 2J0O) (39); Flagellin family (FliC; PDB ID code 1IO1) (22); E. coli family (EspA; PDB ID code 1XOU) (42). The structures demonstrate a common architecture, consisting of a central coiled-coil (green), a needle-distal domain (red), and a needle-proximal domain (purple/blue). The needle-proximal portion of the coiled-coil, which is postulated to be involved in assembly at the needle-tip, is chaperoned (shown in blue). This chaperoning may be via an intramolecular mechanism (IpaD: IpaD N-terminal domain) or separate chaperones [EspA:CesA, PDB ID code 1XOU (42), FliC:FliS, PDB ID codes 2IO1/1ORJ (22, 40), LcrV:LcrG, LcrV; PDB ID code 1R6F; no structure available for LcrG, modeled on IpaD N-terminal domain structure]. (B) Schematic representation of the common architecture. (C) The C-terminal helix of the central coiled-coil is straight in chaperoned adaptor proteins (IpaD chaperoned, PDB ID zj0o and bent in the nonchaperoned structure (IpaD nonchaperoned, PBD ID 2j0n). This bend is comparable to the kink in the C-terminal helix of the Shigella needle protein MxiH (PDB ID code 2CA5) (24).

Fig. 3.

Models for tip-associated protein assembly. (A) Adaptor protein homopentamers as suggested by the LcrV EM data (10) (Upper) and the IpaD crystal structure (39) (Lower) suggest open and closed forms of the tip complex, respectively. The position of the N-terminal chaperoning domain of IpaD is unknown in the pentamer and so is shown in a more extended conformation projecting away from the tip/needle (only two are shown in the side view for clarity). The Shigella needle is shown as cartoons (PDB ID code 2V6L) (24). (B) Ribbon representation of a proposed hetero-pentamer tip complex, consisting of four copies of IpaD (colored as in A) and one copy of IpaB (shown in gray as a blurred surface and modeled on the structure of the pore-forming domain of the Colicin family of pore-forming toxins).

Fig. 4.

Schematic illustrations of possible different states of the T3SS needle complex and sites of action for needle mutants. (A) Changes in three distinct regions of the T3SS may be involved in the regulation of secretion after host-cell sensing: the tip complex (1, blue), the needle itself (2, black), and the basal apparatus (3, red). Secretion substrates are shown in the needle channel in a partially unfolded state (light gray and gray). (B) Mutations of the needle protein can alter secretion patterns by disruption of four different interfaces (red circles): needle subunit to tip complex (a), within the needle subunit, i.e., intramolecular (b), between the needle subunits, i.e., intermolecular (c), and needle subunit to base (d).