RyR2 regulates store-operated Ca2+ entry, phospholipase C activity, and electrical excitability in the insulinoma cell line INS-1

Abstract

The ER Ca2+ channel ryanodine receptor 2 (RyR2) is required for maintenance of insulin content and glucose-stimulated insulin secretion, in part, via regulation of the protein IRBIT in the insulinoma cell line INS-1. Here, we examined store-operated and depolarization-dependent Ca2+entry using INS-1 cells in which either RyR2 or IRBIT were deleted. Store-operated Ca2+ entry (SOCE) stimulated with thapsigargin was reduced in RyR2KO cells compared to controls, but was unchanged in IRBITKO cells. STIM1 protein levels were not different between the three cell lines. Basal and stimulated (500 μM carbachol) phospholipase C (PLC) activity was also reduced specifically in RyR2KO cells. Insulin secretion stimulated by tolbutamide was reduced in RyR2KO and IRBITKO cells compared to controls, but was potentiated by an EPAC-selective cAMP analog in all three cell lines. Cellular PIP2 levels were increased and cortical f-actin levels were reduced in RyR2KO cells compared to controls. Whole-cell Cav channel current density was increased in RyR2KO cells compared to controls, and barium current was reduced by acute activation of the lipid phosphatase pseudojanin preferentially in RyR2KO cells over control INS-1 cells. Action potentials stimulated by 18 mM glucose were more frequent in RyR2KO cells compared to controls, and insensitive to the SK channel inhibitor apamin. Taken together, these results suggest that RyR2 plays a critical role in regulating PLC activity and PIP2 levels via regulation of SOCE. RyR2 also regulates β-cell electrical activity by controlling Cav current density and SK channel activation.

Dysregulation of calcium homeostasis in cancer and its role in chemoresistance

“…Not defined [92] , PMCA2 [93] , PMCA1 [32,62] , PMCA4 [32,62] Effects wound healing [62] , increased [Ca 2+ ] cyt [62] , increased nuclear NFATc1 accumulation [62] Subjects T-ALL [94,95] , breast adenocarcinoma cell lines [95] Treatment Xestospongin B…”

“…, 2APB [95] IP3R Not defined [94] Effects Reduced mitochondrial respiration [94], cell death [94] , increased NAD + /NADH ratio [94] , inhibition of ER-Ca 2+ release [95] , inhibition of ROS production [95] , inhibition of immunepotent CRP-induced death [95] Subjects NSCLC cells [88] Treatment Afatinib [88] SERCA SERCA2 [88] Effects Autophagy [88] , reduced cell viability [88] Subjects Breast adenocarcinoma cell lines [95] , T-ALL cells [95] , insulinoma cell line (INS-1) [96] Treatment Dantrolene [95] , knockout [96] RyR Not defined [95] , RyR2 [96] Effects…”

“…Inhibition of ER-Ca 2+ release [95] , inhibition of ROS production [95] , inhibition of immunepotent CRP-induced death [95] , impaired SOCE [96] , reduced [Ca 2+ ] cyt [96] , elevated VGCC current density [96] , increased action potential frequency [96] Subjects MCU-knockout transformed fibroblast cells [37] , mice [37] , Pancreatic ductal adenocarcinoma tissue [67] , mice [67] , CRC [70] Treatment Knockout [37,67] , knockdown [70] MCU Not defined [37,67,70] Effects Small tumour size [37] , reduced proliferation [37,70] , cell cycle arrest at S phase [37] , increased glycolysis [37] , inhibited cell migration [67] , inhibited invasion [37,67] , inhibited soft agar colony formation [67] , reduced tumour weight [67] , reduced ROS level [67] , ferroptosis inhibition [67] , Nrf2 upregulation [67] , decreased mitochondrial content [70] , decreased mtDNA copy number [70] , decreased ATP production [70] , decreased expression levels of oxidative phosphorylation-related proteins [70] Subjects Pancreatic ductal adenocarcinoma tissue [67] , colorectal cancer cell lines [97] , knockout mice [97] Treatment CGP37157 [67] , si/shRNA [97] NCLX Not defined…”