Abstract

STIM1, Orai1 and TRPC1 are all reported to be important for store-operated Ca(2+) entry (SOCE) in diverse cells. However, there is no evidence for the functional interaction of the three proteins in SOCE in human liver cells. The objective of this study is to determine whether they are involved in SOCE in normal human liver cells. Liposomal transfection method was used to increase expression levels of the three proteins in HL-7702 cells, a normal human liver cell line. Western blot and single cell RT-PCR were applied to evaluate transfection effectiveness. Changes in store-operated current (I(SOC)) and SOCE were investigated using whole-cell patch-clamp recording and calcium imaging. I(SOC) is detected in HL-7702 cells and it is inhibited either by 2-Aminoethoxydiphenyl borate (2-APB) or La(3+). Overexpression of STIM1 or Orai1 alone did not induce any change in I(SOC). TRPC1-transfection, however, caused approximate 2.5-fold increase in I(SOC). A large increase (>10-fold) in I(SOC) emerged when both STIM1 and Orai1 were co-transfected into HL-7702 cells. Co-overexpression of STIM1 + TRPC1 also caused >10-fold increase in I(SOC), and addition of Orai1 did not cause any further increase. In HL-7702 cells, TRPC1 and Orai1 take part in SOCE independently of each other. Functional interactions of STIM1 and Orai1 or TRPC1 contribute to I(SOC) activation.