Structural mechanism of human TRPC3 and TRPC6 channel regulation by their intracellular calcium-binding sites

Area of Science:

• Molecular biology
• Structural biology
• Biophysics

Background:

• Transient Receptor Potential Canonical (TRPC) channels, including TRPC3 and TRPC6, are calcium-permeable cation channels vital for physiological functions.
• Gain-of-function (GOF) mutations in TRPC6 are linked to familial focal segmental glomerulosclerosis (FSGS), a progressive kidney disease, but the underlying mechanisms are unclear.

Purpose of the Study:

• To elucidate the structural basis of TRPC3 and TRPC6 channel regulation by intracellular calcium.
• To understand the pathogenic mechanisms of TRPC6 GOF mutations in FSGS.
• To provide structural templates for developing TRPC6-specific inhibitors.

Main Methods:

• Cryo-electron microscopy (cryo-EM) to determine high-resolution structures of human TRPC3 and TRPC6.
• Electrophysiological studies to assess channel activity and calcium sensitivity.
• Structural analysis of TRPC6 in complex with inhibitors.

Main Results:

• Identified distinct inhibitory and activating calcium-binding sites in TRPC3 that modulate basal channel activity.
• Demonstrated structural and functional conservation of these calcium sensors in TRPC6.
• Revealed that TRPC6 GOF mutations activate the channel by disrupting inhibitory calcium binding.
• Obtained structures of TRPC6 with inhibitors, showcasing distinct transmembrane domain conformations.

Conclusions:

• Intracellular calcium directly regulates TRPC3 and TRPC6 channel gating through conserved calcium-binding sites.
• TRPC6 GOF mutations in FSGS result from the allosteric disruption of inhibitory calcium sensing.
• The determined structures provide a foundation for structure-based drug design against TRPC6-related diseases like FSGS.