Type 3 Inositol 1,4,5-Trisphosphate Receptor is a Crucial Regulator of Calcium Dynamics Mediated by Endoplasmic Reticulum in HEK Cells

Abstract

Being the largest the Ca2+ store in mammalian cells, endoplasmic reticulum (ER)-mediated Ca2+ signalling often involves both Ca2+ release via inositol 1, 4, 5-trisphosphate receptors (IP3R) and store operated Ca2+ entries (SOCE) through Ca2+ release activated Ca2+ (CRAC) channels on plasma membrane (PM). IP3Rs are functionally coupled with CRAC channels and other Ca2+ handling proteins. However, it still remains less well defined as to whether IP3Rs could regulate ER-mediated Ca2+ signals independent of their Ca2+ releasing ability. To address this, we generated IP3Rs triple and double knockout human embryonic kidney (HEK) cell lines (IP3Rs-TKO, IP3Rs-DKO), and systemically examined ER Ca2+ dynamics and CRAC channel activity in these cells. The results showed that the rate of ER Ca2+ leakage and refilling, as well as SOCE were all significantly reduced in IP3Rs-TKO cells. And these TKO effects could be rescued by over-expression of IP3R3. Further, results showed that the diminished SOCE was caused by NEDD4L-mediated ubiquitination of Orai1 protein. Together, our findings indicate that IP3R3 is one crucial player in coordinating ER-mediated Ca2+ signalling.

Metabolic adaptation to the chronic loss of Ca2+ signaling induced by KO of IP3 receptors or the mitochondrial Ca2+ uniporter

“…Some of these observations are in agreement with previous studies. A lower rate of proliferation and migration in IP 3 R TKO HEK293 cells has been reported recently ( 31 ). A reduced baseline and FCCP-stimulated oxygen consumption was observed in Seahorse assays ( 85 ).…”

“…They concluded that, unlike normal cells, the stimulation of autophagy in cancer cells is not sufficient to compensate for the increased energy demand of uncontrolled growth, with cell death resulting from a “bioenergetic crisis” during mitosis. The advent of new methods of gene disruption has enabled all three IP 3 R isoforms to be deleted in human embryonic kidney 293 (HEK293) ( 30 , 31 ) and HeLa ( 32 ) human cancer cell lines. While these KO cells have primarily been used as an expression system to study the functional properties of IP 3 R mutants, they also offer the opportunity to investigate the metabolic and bioenergetic consequences of the chronic effects of a total loss of Ca 2+ signaling.…”

CEMIP promotes extracellular matrix‐detached prostate cancer cell survival by inhibiting ferroptosis

“… 32 ITPR3 is a Ca 2+ release channel that resides in the membranes of the ER, which is extremely important for intracellular Ca 2+ homeostasis. 54 The present study revealed that the interaction of CEMIP and ITPR3 led to ER Ca 2+ leakage and increased cytosolic Ca 2+ levels in ECM‐detached PCa cells, subsequently activating CaMKII. Furthermore, functional studies revealed that CaMKII facilitates NRF2 phosphorylation at S40 and nuclear localization, leading to elevated SLC7A11 transcription in PCa cells.…”

Inhibiting endoplasmic reticulum stress by activation of G‐protein‐coupled estrogen receptor to protect retinal astrocytes under hyperoxia

“…Inositol‐1,4,5‐triphosphate (IP3) and its receptor IP3R located at the surface of the ER are responsible for regulating intracellular Ca 2+ release. [ 18 ] In principle, Mag‐Fluo‐AM, a type of fluorescent dye with low Ca 2+ affinity, is specifically captured by ER to detect the changes of Ca 2+ concentration and evaluate the transfer activity of IP3R. First, the IP3R activity was detected by an IP3R functional fluorescence detection kit (Shanghai Yuduo Biotech Ltd, China).…”