Figure 1.

Characterization of HEK IP₃Rs-TKO and IP₃Rs-DKO cells. (A–C) Typical carbachol (CCh, 100 μM)-induced Ca²⁺ responses from ER in HEK IP₃Rs-TKO and IP₃Rs-DKO cells. (A) CCh-induced Ca²⁺ changes as shown with transiently expressed cytoplasmic Ca²⁺ indicator GCaMP6m (Green) or stably expressed ER Ca²⁺ indicator R-CEPIA1er (Red). Left panel, responses of WT HEK cell stabling expressing R-CEPIA1er; right panel, R-CEPIA1er stable cells with IP₃Rs-TKO (RIPK). (B) Representative CCh-induced responses in HEK GCaMP6m stable cells without (WT, black trace) or with IP₃Rs-TKO (GIPK, green trace). (C) Typical CCh-induced Ca²⁺ releases in HEK GCaMP6m WT cells or those with IP₃Rs-DKO. 1 mM GdCl₃ was included in bath solution to block Ca²⁺ movements across PM in B) and C) (n = 3). (D) Statistics showing relative sizes of mean CCh-induced Ca²⁺ releases in RIPK or GIPK cells transiently expressing IP₃Rs (see Figure S1A,B for typical curves). (E) Relative mRNA levels of three types of IP₃Rs in HEK cells and IP₃Rs-DKO. mRNA levels were first normalized against corresponding GAPDH levels, then normalized against corresponding IP₃R1 levels of WT cells. Expression levels of IP₃R1 in WT cells were set as 1 (mean ± SEM, *** P < 0.0001, Student’s t-test). (F) Typical proliferation (left panel) and migration (right panel) curves of RIPK and WT cells.