Role of TRPC3 in the formation of receptor- and store-operated calcium channels in A431 carcinoma cells

Activation of PLC-linked intracellular signaling cascades in the non-excitable cells evokes the release of calcium ions from the inositol 1,4,5-trisphosphate (InsP3)-sensitive intracellular Ca2+ stores and Ca2+ entry in the cytosol via Ca2+-channels of plasma membrane. The properties and molecular identity of these channels are now under intense investigation. It is speculated that mammalian proteins belonging to the TRP-related family take part in either receptor- and store- dependent entry, though data linking the specific TRP proteins and any endogenous Ca 2+- channel are very scarce. Thus we aimed to study the role of TRPC3 in the formation of receptor- and store- operated calcium entry pathways in A431 cells. Both whole-cell current recordings and fluorescent measurements of intracellular Ca2+ concentration have shown that partial inhibition of TRPC3 expression with small interfering RNAs (siRNA) suppresses the store-dependent Ca2+ entry, but does not affect the receptor-mediated Ca2+ entry. The investigations on the single-channel level revealed that TRPC3 suppression leads to the disappearance of one of the types of store operated Ca2+- channels in the plasma membrane and appearance of a new type of a store-independent channel in it. It might indicate that TRPC3 protein is needed for the functioning of store-dependent channel in A431 cells

Role of TRPC3 in the formation of receptor- and store-operated calcium channels in A431 carcinoma cells

Activation of PLC-linked intracellular signaling cascades in the non-excitable cells evokes the release of calcium ions from the inositol 1,4,5-trisphosphate (InsP3)-sensitive intracellular Ca2+ stores and Ca2+ entry in the cytosol via Ca2+-channels of plasma membrane. The properties and molecular identity of these channels are now under intense investigation. It is speculated that mammalian proteins belonging to the TRP-related family take part in either receptor- and store- dependent entry, though data linking the specific TRP proteins and any endogenous Ca 2+- channel are very scarce. Thus we aimed to study the role of TRPC3 in the formation of receptor- and store- operated calcium entry pathways in A431 cells. Both whole-cell current recordings and fluorescent measurements of intracellular Ca2+ concentration have shown that partial inhibition of TRPC3 expression with small interfering RNAs (siRNA) suppresses the store-dependent Ca2+ entry, but does not affect the receptor-mediated Ca2+ entry. The investigations on the single-channel level revealed that TRPC3 suppression leads to the disappearance of one of the types of store operated Ca2+- channels in the plasma membrane and appearance of a new type of a store-independent channel in it. It might indicate that TRPC3 protein is needed for the functioning of store-dependent channel in A431 cells

Suppression of TRPC3 leads to disappearance of store-operated channels and formation of a new type of store-independent channels in A431 cells.

In most non-excitable cells, calcium (Ca(2+)) release from the inositol 1,4,5-trisphosphate (InsP(3))-sensitive intracellular Ca(2+) stores is coupled to Ca(2+) influx through the plasma membrane Ca(2+) channels whose molecular composition is poorly understood. Several members of mammalian TRP-related protein family have been implicated to both receptor- and store-operated Ca(2+) influx. Here we investigated the role of the native transient receptor potential 3 (TRPC3) homologue in mediating the store- and receptor-operated calcium entry in A431 cells. We show that suppression of TRPC3 protein levels by small interfering RNA (siRNA) leads to a significant reduction in store-operated calcium influx without affecting the receptor-operated calcium influx. With single-channel analysis, we further demonstrate that reduction of TRPC3 levels results in suppression of specific subtype of store-operated calcium channels and activation of store-independent channels. Our data suggest that TRPC3 is required for the formation of functional store-operated channels in A431 cells