Editors' ChoiceCalcium signaling

Partners for IP3 Receptors

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Science's STKE  08 Feb 2005:
Vol. 2005, Issue 270, pp. tw50
DOI: 10.1126/stke.2702005tw50

Two articles from the Snyder lab this week describe regulation of calcium release from the endoplasmic reticulum by proteins that interact with the inositol trisphosphate receptor (IP3R). Patterson et al. describe a mechanism for the local production of nicotinamide adenine dinucleotide NADH in the vicinity of the IP3R and show that glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which generated NADH, binds directly to the IP3R. Yeast two-hybrid assays, followed by binding of purified proteins and coimmunoprecipitation assays, indicated that the IP3R directly bound GAPDH. Furthermore, the interaction involved an active site cysteine of GAPDH and two cysteines in the 981 to 1000 amino acid region of the IP3R, suggesting a possible disulfide bridge. IP3Rs reconstituted into lipid vesicles exhibited an increase in Ca2+ flux in response to IP3 when GAPDH and NAD+ were both present, indicating that the catalytic activity of GAPDH may convert NAD+ to NADH, which is known to augment IP3R activity. In microsomal preparations of cells expressing a mutated form of the IP3R that could not bind GAPDH, Ca2+ flux was not increased by the addition of GAPDH and NAD+. This interaction may allow changes in cellular energy status to be tightly coupled to IP3R function and calcium signaling. Indeed, this may contribute to the pathological increase in intracellular calcium concentration ([Ca2+]i) associated with cell death during hypoxic conditions or periods of inhibition of mitochondrial respiration. Boehning et al. followed the mechanism by which this pathological increase in [Ca2+]i occurs in response to apoptotic stimuli and describe a peptide inhibitor that blocks the interaction of cytochrome c with the IP3R, thereby inhibiting cell death. The current work follows previous studies that indicated that cytochrome c blocked calcium-mediated inhibition of the IP3R, a negative feedback pathway, and thus contributed to potentially toxic elevations in [Ca2+]i. Boehning et al. determined that the binding site on the type 1 IP3R for cytochrome c involved two glutamic acid residues, and the interaction was presumably mediated by electrostatic interactions that are characteristic of the interaction of cytochrome c with many other proteins. They generated a cell-permeant peptide that blocked the interaction between cytochrome c and the IP3R and found that this peptide reduced apoptosis induced by Fas ligand application to Jurkat T lymphoma cells or apoptosis induced by staurosporine in HeLa cells. Fas ligand also triggered a relocation of cytochrome c from the mitochondrial fraction to the light membrane fraction (that includes the endoplasmic reticulum) and promoted the interaction between cytochrome c and IP3R, which was detected by coimmunoprecipitation. Surprisingly, the cell-permeant peptide that inhibited cell death did not decrease calcium mobilization in either the Fas ligand-treated Jurkat cells or the staurosporine-treated HeLa cells. Thus, the mechanism by which this peptide exhibits its cell survival effects remains unclear.

R. L. Patterson, D. B. van Rossum, A. I. Kaplin, R. K. Barrow, S. H. Snyder, Inositol 1,4,5-trisphosphate receptor/GAPDH complex augments Ca2+ release via locally derived NADH. Proc. Natl. Acad. Sci. U.S.A. 102, 1357-1359 (2005). [Abstract] [Full Text]

R. Boehning, D. B. van Rossum. R. L. Patterson, S. H. Snyder, A peptide inhibitor of cytochrome c/inositol 1,4,5 trisphosphate receptor binding blocks intrinsic and extrinsic cell death pathways. Proc. Natl. Acad. Sci. U.S.A. 102, 1466-1471 (2005). [Abstract] [Full Text]