# Regulation of inositol trisphosphate receptors

> **NIH NIH R01** · THOMAS JEFFERSON UNIVERSITY · 2021 · $322,436

## Abstract

Calcium is a universal messenger controlling a multitude of cellular responses including muscle contraction,
exocytosis, memory, fertilization, metabolism, proliferation and cell death. Numerous hormones,
neurotransmitters and growth factors stimulate the formation of inositol 1,4,5-trisphosphate ( IP3 ) which acts
on a family of receptors ( IP3Rs ) located in the endoplasmic reticulum that function as ligand-gated Ca2+
channels. The depletion of intracellular Ca2+ stores also activates Ca2+ influx mechanisms in the plasma
membrane. Thus, the interaction of IP3 with its receptor activates all phases of a Ca2+ signal. IP3Rs are
regulated by Ca2+ and phosphorylation but the molecular basis of this regulation is poorly understood. Ca2+
released from IP3Rs is locally transmitted to the mitochondria and can stimulate metabolism, and in higher
amounts, can also initiate cell death. Cancer cells have been proposed to be highly dependent on IP3R-
mediated mitochondrial Ca2+ transfer for their survival. The advent of CRISPR/Cas-9 technology has allowed
the genetic ablation of all three IP3R isoforms from HEK293 and HeLa cervix carcinoma cells. The proposal is
centered on the use of these cell lines to a) study the adaptive mechanisms allowing the cells to survive in the
complete absence of Ca2+ signaling and b) to take advantage of a null background for structure-function
studies exploring the molecular mechanism by which IP3 opens the channel and the mechanism of feed-back
regulation by Ca2+. The two specific aims are: 1] Characterize adaptive mechanisms in IP3R-3KO cells: The
impact of a lack of Ca2+ regulation of metabolism will be investigated by measuring several bioenergetic
parameters and 13C-tracer metabolism. Proliferation, cell-cycle status, cell death and transcriptional rewiring
will be explored. 2A] How does IP3 open the channel? Chemical crosslinking/mass spectroscopy will be used
to validate cryo-EM structures and monitor conformational changes mediated by IP3 in native IP3Rs.
Mutagenesis of key residues will be used to test proposed allosteric mechanisms of gating. 2B] Identification
of the Ca2+ regulatory sites in IP3Rs: Based on the most recent cryo-EM structures of IP3R1 and IP3R3 we
have identified 3 clusters of negatively charged residues that are candidates for Ca2+ regulatory sites. Their
functional role as stimulatory or inhibitory sites will be assessed by mutagenesis. The role of these sites will
also be tested by mutagenesis of the Ca2+-insensitive IP3R from Capsaspora owczarzaki . The long-term goal
of the proposal is to better understand the role of Ca2+signaling in normal cells, cancer cells and in inherited
disorders inactivating IP3R function. The study should also provide fundamental mechanistic information on
how these important channels work and are regulated.

## Key facts

- **NIH application ID:** 10077856
- **Project number:** 5R01GM132611-02
- **Recipient organization:** THOMAS JEFFERSON UNIVERSITY
- **Principal Investigator:** SURESH K JOSEPH
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $322,436
- **Award type:** 5
- **Project period:** 2020-01-01 → 2023-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10077856

## Citation

> US National Institutes of Health, RePORTER application 10077856, Regulation of inositol trisphosphate receptors (5R01GM132611-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10077856. Licensed CC0.

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