Small, Dynamic Clusters of Tir-Intimin Seed Actin Polymerization

Area of Science:

• Microbiology
• Cell Biology
• Biophysics

Background:

• Actin pedestal formation by enteropathogenic Escherichia coli (EPEC) is crucial for bacterial attachment.
• Current understanding relies on static data, lacking insights into dynamic processes.

Purpose of the Study:

• To investigate the dynamic nature of signaling components during actin pedestal formation.
• To characterize the formation and regulation of intimin receptor (Tir) clusters.

Main Methods:

• Utilized live-cell imaging and single-molecule tracking.
• Employed supported lipid bilayers and transfected cells expressing Tir.

Main Results:

• Demonstrated dynamic signaling clusters on the subsecond timescale, resembling phase condensates.
• Observed Tir-intimin clusters (200 nm) regulated in size and spacing.
• Tir diffusion within clusters and exchange with the plasma membrane were shown.
• Nck and N-WASP binding to clusters and exchange with cytoplasmic pools were identified.
• Actin polymerization required N-WASP binding and Arp2/3 activation, stabilizing N-WASP clusters.

Conclusions:

• Tir-intimin clusters are dynamically regulated in size by molecular exchange.
• These clusters enhance signaling for actin polymerization, enabling stable bacterial attachment.
• The dynamic properties differ significantly from in vitro systems and are cell-regulated.