Poster abstracts
Poster number 21 submitted by Wei-Hsiang Weng
Molecular Mechanisms Underlying CIB-mediated TMC Activation in Inner-ear Mechanotransduction
Wei-Hsiang Weng (Biophysics Graduate Program, Ohio State University, Columbus, OH, USA), Jonathan Montgomery (The Ohio State Biochemistry Program OSBP, Ohio State University, Columbus, OH, USA), Jeffrey M. Lotthammer (Department of Chemistry and Biochemistry, Ohio State University, Columbus, OH, USA), Sanket Walujkar (Chemical Physics Graduate Program, Ohio State University, Columbus, OH, USA), Mark P. Foster (Department of Chemistry and Biochemistry, Ohio State University, Columbus, OH, USA), Marcos Sotomayor (Department of Chemistry and Biochemistry, Ohio State University, Columbus, OH, USA)
Abstract:
Inner-ear mechanotransduction is initiated when mechanical stimuli from sound and head movements stretch tip-link protein filaments to open ion channels that trigger sensory perception. This allows for cation influx which causes hair-cell depolarization to initiate sensory perception. Transmembrane channel-like proteins (TMCs) 1 and 2 have been established as the pore-forming components of hair-cell transduction channels with calcium- and integrin-binding (CIB) proteins 2 and 3 functioning as essential auxiliary subunits. Separate studies have reported that CIB proteins bind to either the N-terminal (NT) or to the intracellular loop-1 (IL1) domains of TMC1. Additionally, CIB proteins may be myristoylated (addition of a 14-carbon unsaturated fatty acid) at their N-terminal glycine following removal of the first methionine to mediate association with biological membranes. However, structural details regarding how CIB2/3 interact with TMC1/2 intracellularly to modulate mechanotransduction remain unresolved. It is also unclear how force is transmitted to the channel for mechanotransduction. Here, we present AlphaFold 2 models of TMC:CIB complexes that suggest ‘clamp-like’ models in which CIB is interacting with both TMC NT and IL1 cytosolic domains. We also present data from nuclear magnetic resonance (NMR) experiments testing the interaction between full-length [15N]-labeled CIB proteins with TMC-IL1 and TMC-NT peptides, which suggest that both peptides bind [15N]-CIB non-competitively. Molecular dynamics simulations reveal a trend in which TMC1 exhibits higher conductance with CIB2 bound than without it at microsecond timescales under near physiological voltages. Simulations show that the CIB-myristoylation facilitates association of the complex with the lipid bilayer. Our models also feature an amphipathic helix inserted in the membrane bilayer in a near-parallel fashion. Association of TMC:CIB with the lipid bilayer suggest that membrane tension might play a role in the channel activation in the inner ear. These data and models consolidate experimental results about the interactions between TMC and CIB proteins and suggest how the complex may function in inner-ear mechanotransduction.
Keywords: Mechanotransduction, Molecular dynamics simulation, Nuclear magnetic resonance