Heteromeric TRPC3 with TRPC1 formed via its ankyrin repeats regulates the resting cytosolic Ca²⁺ levels in skeletal muscle

The main tasks of skeletal muscle are muscle contraction and relaxation, which are mediated by changes in cytosolic Ca²⁺ levels. Canonical-type transient receptor potential 3 (TRPC3) contains an ankyrin repeat (AR) region at the N-terminus (38-188 amino acids) and forms extracellular Ca²⁺-entry channels by homo or heteromerization with other TRP subtypes in various cells including skeletal myotubes. However, previous research has not determined which region(s) of TRPC3 is responsible for the heteromerization, whether the AR region participates in the heteromerizations, or what is the role of heteromeric TRPC3s in skeletal muscle. In the present study, the heteromerization of TRPC3 with TRPC1 was first examined by GST pull-down assays of TRPC3 portions with TRPC1. The portion containing the AR region of TRPC3 was bound to the TRPC1, but the binding was inhibited by the very end sub-region of the TRPC3 (1-37 amino acids). In-silico studies have suggested that the very end sub-region possibly induces a structural change in the AR region. Second, the very end sub-region of TRPC3 was expressed in mouse primary skeletal myotubes, resulting in a dominant-negative inhibition of heteromeric TRPC3/1 formation. In addition, the skeletal myotubes expressing the very end sub-region showed a decrease in resting cytosolic Ca²⁺ levels. These results suggest that the AR region of TRPC3 could mediate the heteromeric TRPC3/1 formation, and the heteromeric TRPC3/1 could participate in regulating the resting cytosolic Ca²⁺ levels in skeletal muscle.