Distinct Cytosolic Complexes Containing the Type III Secretion System ATPase Resolved by Three-Dimensional Single-Molecule Tracking in Live Yersinia enterocolitica

Abstract

Injectisomes are membrane-embedded, multiprotein assemblies used by bacterial pathogens to inject virulent effector proteins into eukaryotic host cells. Protein secretion is regulated by cytosolic proteins that dynamically bind and unbind at injectisomes.

Ultrafast Spectroscopy Reveals Slow Water Dynamics in Biocondensates

“…Similar analyses are typically applied to measured protein diffusion rate histograms in cells to quantify populations of bound protein. 60 This population of water with slow water diffusion agrees quite well with observed protein diffusion rates in polymer coacervates and agrees quite well with a measured slowdown of diffusive properties of proteins within the postsynaptic density condensate. 61,62 Our analysis shows that the dynamics within the condensate are slower overall than those within the bulk as a result of bound water, which explains the longer single H-bond lifetimes computed from the trajectories.…”

Super‐resolution fluorescence microscopy for investigating bacterial cell biology

“…Thereby, subcellular distributions and rough numbers of needle complexes, sorting platform components, tip complex, and an effector could be determined. Needle complexes including export apparatus were almost exclusively located at the bacterial plasma membrane, whereas a considerable fraction of sorting platform components was also in the cytoplasm, suggesting that sorting platforms are transiently and dynamically associated with the needle complexes (Figure 2e; Diepold et al, 2017;Prindle et al, 2022).…”

“…Blue trajectories: diffusion coefficient D* < 0.5 μm 2 /s. Orange trajectories: diffusion coefficients D * > 0.5 μm 2 /s (Prindle et al., 2022). (f) Trajectories of 2D‐PALM SMT of the PAmCherry‐labeled beta' subunit of RNA polymerases (RNAPs) in E. coli .…”

“…SMT and SMLM in live Yersinia enterocolitica revealed distinct diffusive states of the eYFP, eGFP, and PAmCherry‐labeled sorting platform components YscQ, YscL, and YscN (unified nomenclature: SctQ, SctL, and SctN) and suggested that they form distinct cytosolic complexes before binding to the needle complex (Figure 2e; Diepold et al., 2015; Prindle et al., 2022; Rocha et al., 2018). SMT of eGFP‐labeled YscD (unified nomenclature: SctD) showed partial disassembly of the T3SS basal body component at low external pH (Wimmi et al., 2021).…”

The sorting platform in the type III secretion pathway: From assembly to function

“…A fascinating question that arises then is how the chaperone/effector complexes are brought to the ATPase for unfolding and secretion? Complexes of some SP proteins can be detected freely diffusing in the bacterial cytosol, [ 58 ] it is possible that these complexes usher chaperone/effector complexes to the SP for secretion. Consistent with this hypothesis, it has been proposed that the dynamic interchange between the injectisome‐bound SctQ and the cytosolic pool could potentially reflect a shuttle‐like mechanism, wherein SctQ facilitates the handover of cytosolic substrates to the injectisome.…”