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An alternatively spliced STING isoform localizes in the cytoplasmic membrane and directly senses extracellular cGAMP
Xiaobo Li, … , Shengnan Luo, Tongsen Zheng
Xiaobo Li, … , Shengnan Luo, Tongsen Zheng
Published December 14, 2021
Citation Information: J Clin Invest. 2022;132(3):e144339. https://doi.org/10.1172/JCI144339.
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Research Article Immunology Oncology

An alternatively spliced STING isoform localizes in the cytoplasmic membrane and directly senses extracellular cGAMP

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Abstract

It has been revealed that 2′3′-cyclic-GMP-AMP (cGAMP), a second messenger that activates the antiviral stimulator of IFN genes (STING), elicits an antitumoral immune response. Since cGAMP cannot cross the cell membrane, it is not clear how intracellular STING has been activated by extracellular cGAMP until SLC19A1 was identified as an importer to transport extracellular cGAMP into the cytosol. However, SLC19A1-deficient cells also sense extracellular cGAMP, suggesting the presence of mechanisms other than the facilitating transporters for STING sensing extracellular cGAMP. Here, using immunoprecipitation, immunofluorescence, and flow cytometry, we identified an alternatively spliced STING isoform, plasmatic membrane STING (pmSTING), that localized in the plasma membrane with its C-terminus outside the cell, due to a lack of 1 transmembrane domain in its N-terminus compared with canonical STING. Further studies showed that extracellular cGAMP not only promoted the dimerization of pmSTING and interaction of pmSTING with TANK-binding kinase 1 (TBK1) and IFN regulatory factor 3 (IRF3), but also enhanced the phosphorylation of TBK1 and IRF3 and the production of IFN in pmSTING-transfected cells. Additionally, we also identified similar pmSTING isoforms in other species including human. This study suggests a conserved role for pmSTING in sensing extracellular cGAMP and provides insight into the role of cGAMP as an immunotransmitter.

Authors

Xiaobo Li, Yuanyuan Zhu, Xiao Zhang, Xiang An, Mingjiao Weng, Jiaqi Shi, Song Wang, Caiqi Liu, Shengnan Luo, Tongsen Zheng

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Figure 2

Identification of a cell surface STING projecting its C-terminus outside of splenocytes.

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Identification of a cell surface STING projecting its C-terminus outside...
(A) A cell surface STING with its C-terminus outside of mouse splenocytes was detected with 3 antibodies against the STING C-terminal epitope using flow cytometry. (B) WT splenocytes were incubated with the indicated antibodies and then washed and lysed. Immunoblotting was performed to detect the existence of IgG in the cell lysate using a second antibody against rabbit IgG. (C) Colocalization of cell surface STING with surface proteins of T cells (CD3), B cells (CD19), and myeloid cells (CD11b) from C57BL/6 mice was detected using confocal microscopy. Scale bars: 5 μm. (D) Expression of cell surface STING and surface proteins of T cells (CD3), B cells (CD19), and myeloid cells (CD11b) from STING-deficient mice (Tmem173gt) using confocal microscopy. Scale bars: 5 μm. (E) Extracellular cGAMP–induced production of IFN-β was detected by ELISA in WT splenocytes preincubated with the indicated antibodies (n = 3). **P < 0.01, by 1-way ANOVA followed by Dunnett’s test for comparison with the isotype antibody and the cGAMP treatment group.

Copyright © 2025 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)

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