PP2Ac/STRN4 negatively regulates STING-type I IFN signaling in tumor-associated macrophages
Winson S. Ho, Isha Mondal, Beisi Xu, Oishika Das, Raymond Sun, Pochin Chiou, Xiaomin Cai, Foozhan Tahmasebinia, Elizabeth McFadden, Caren Yu-Ju Wu, Zhihao Wu, William Matsui, Michael Lim, Zhipeng Meng, Rongze Olivia Lu
Winson S. Ho, Isha Mondal, Beisi Xu, Oishika Das, Raymond Sun, Pochin Chiou, Xiaomin Cai, Foozhan Tahmasebinia, Elizabeth McFadden, Caren Yu-Ju Wu, Zhihao Wu, William Matsui, Michael Lim, Zhipeng Meng, Rongze Olivia Lu
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Research Article Immunology Oncology

PP2Ac/STRN4 negatively regulates STING-type I IFN signaling in tumor-associated macrophages

Abstract

Stimulator of IFN genes type I (STING-Type I) IFN signaling in myeloid cells plays a critical role in effective antitumor immune responses, but STING agonists as monotherapy have shown limited efficacy in clinical trials. The mechanisms that downregulate STING signaling are not fully understood. Here, we report that protein phosphatase 2A (PP2A), with its specific B regulatory subunit Striatin 4 (STRN4), negatively regulated STING-Type I IFN in macrophages. Mice with macrophage PP2A deficiency exhibited reduced tumor progression. The tumor microenvironment showed decreased immunosuppressive and increased IFN-activated macrophages and CD8+ T cells. Mechanistically, we demonstrated that Hippo kinase MST1/2 was required for STING activation. STING agonists induced dissociation of PP2A from MST1/2 in normal macrophages, but not in tumor conditioned macrophages. Furthermore, our data showed that STRN4 mediated PP2A binding to and dephosphorylation of Hippo kinase MST1/2, resulting in stabilization of YAP/TAZ to antagonize STING activation. In human patients with glioblastoma (GBM), YAP/TAZ was highly expressed in tumor-associated macrophages but not in nontumor macrophages. We also demonstrated that PP2A/STRN4 deficiency in macrophages reduced YAP/TAZ expression and sensitized tumor-conditioned macrophages to STING stimulation. In summary, we demonstrated that PP2A/STRN4-YAP/TAZ has, in our opinion, been an unappreciated mechanism that mediates immunosuppression in tumor-associated macrophages, and targeting the PP2A/STRN4-YAP/TAZ axis can sensitize tumors to immunotherapy.

Authors

Winson S. Ho, Isha Mondal, Beisi Xu, Oishika Das, Raymond Sun, Pochin Chiou, Xiaomin Cai, Foozhan Tahmasebinia, Elizabeth McFadden, Caren Yu-Ju Wu, Zhihao Wu, William Matsui, Michael Lim, Zhipeng Meng, Rongze Olivia Lu

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Figure 6

STRN4, a regulatory B subunit of PP2A, negatively regulates STING-Type I IFN by modulation of Hippo kinase MST1/2 and YAP/TAZ.

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STRN4, a regulatory B subunit of PP2A, negatively regulates STING-Type I...
(A) siRNA screen identified PP2A subunits involved in cGAS-STING response. RAW cells were transfected with siRNA of each of 14 regulatory and 2 scaffold subunits of PP2A. 48 hours after transfection, cells were treated with cGAMP (10 μg/mL) for 4 hours. CXCL10 expression was measured via real time PCR. Fold change is relative to nontargeting siRNA. (B) CTL and STRN4KO RAW cells were treated with cGAMP (10 μg/mL), protein expression was analyzed by immunoblotting at different time points after stimulation. (C) RAW cells were transfected with overexpression plasmids for STRN4 or PP2Ac. 48 hours after transfection, cells were treated with cGAMP (10 μg/mL) for 4 hours. Expression of IFNβ and CXCL10 was measured via real time PCR. (D) CTL and STRN4KO THP-1 differentiated macrophages were treated with cGAMP (10 μg/mL) for 4 hours. Expression of IFNβ and CXCL10 was measured via quantitative PCR. (E) CTL and STRN4KO THP-1 differentiated macrophages were treated with cGAMP (10 μg/mL). Protein expression was analyzed by immunoblotting at different time points after stimulation. (F) THP-1 differentiated macrophages were treated with MST-1 inhibitor, XMU-MP-1 (1 μM), for 2 hours, before stimulation with cGAMP (10 μg/mL). 4 hours later, expression of IFNβ and CXCL10 was measured via real time PCR. (G) THP-1 differentiated macrophages were treated with or without cGAMP (10 μg/mL) and 1.5 hours later protein was collected. MST1/2 antibody was used for co-IP and blotted for PP2Ac and MST1/2. Data are from 1 experiment representative of 3 (for B–E) and 2 (for A and G) independent experiments with similar results. Lanes (E and F) separated by black vertical line were run on the same gel but were noncontiguous. Error bars depict SEM. P values were calculated by 2-tailed unpaired t test. ***P < 0.001, ****P < 0.0001.