Development of cGAMP-Luc, a sensitive and precise coupled enzyme assay to measure cGAMP in complex biological samples.

The Journal of Biological Chemistry
Rachel E. MardjukiLingyin Li

Abstract

2',5'/3',5'-cGMP-AMP (cGAMP) is a second messenger produced in response to cytosolic dsDNA that activates the stimulator of interferon genes (STING) pathway. We recently discovered that cGAMP is exported by cancer cells and that this extracellular signal is an immunotransmitter key to tumor detection and elimination by the innate immune system. The enhancement of extracellular cGAMP levels therefore holds great promise for managing cancer. However, there is still much more to understand about the basic biology of cGAMP before its full therapeutic potential can be realized. To answer these questions, we must be able to detect and quantitate cGAMP with an assay that is high-throughput, sensitive, and precise. Existing assays fall short of these needs. Here, we describe the development of cGAMP-Luc, a coupled enzyme assay that relies on the degradation of cGAMP to AMP by ectonucleotide pyrophosphatase phosphodiesterase 1 (ENPP1) and an optimized assay for the detection of AMP by luciferase. We also developed STING-CAP, a STING-mediated method to concentrate and purify cGAMP from any type of biological sample. We conclude that cGAMP-Luc is an economical high-throughput assay that matches the accuracy of and surpasses the detection ...Continue Reading

References

Oct 20, 2005·Journal of Bioscience and Bioengineering·A KamedaT Noguchi
May 13, 2009·Proceedings of the National Academy of Sciences of the United States of America·Wenxiang SunZhengfan Jiang
Sep 29, 2011·Nature·Dara L BurdetteRussell E Vance
Jul 4, 2012·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Kazuki KatoOsamu Nureki
Oct 3, 2012·Proceedings of the National Academy of Sciences of the United States of America·Kazuki KatoOsamu Nureki
Dec 12, 2012·Molecular and Cellular Biology·Amir ApelbaumGideon Schreiber
Oct 27, 2014·Nature Chemical Biology·Lingyin LiTimothy J Mitchison
Jul 25, 2017·Nature·Karen J MackenzieAndrew P Jackson
Jan 18, 2018·Nature·Samuel F BakhoumLewis C Cantley
May 28, 2019·Molecular Cell·Christopher RitchieLingyin Li
Sep 13, 2019·Nature·Rutger D LuteijnDavid H Raulet

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Citations

Nov 4, 2020·Chembiochem : a European Journal of Chemical Biology·Shivam A ZaverJoshua J Woodward
Jan 7, 2021·SLAS Discovery·Meera Kumar, Robert G Lowery
Jan 12, 2021·Human Vaccines & Immunotherapeutics·Chen ChenBinbin Cui
Nov 11, 2020·Molecular Cell·Lauren J LaheyLingyin Li
Nov 7, 2020·EMBO Reports·Jonny Hertzog, Jan Rehwinkel

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