Dephosphorylation of cGAS by PPP6C impairs its substrate binding activity and innate antiviral response
Received date: 29 Mar 2020
Accepted date: 24 Apr 2020
Published date: 15 Aug 2020
Copyright
The cyclic GMP-AMP (cGAMP) synthase (cGAS) plays a critical role in host defense by sensing cytosolic DNA derived from microbial pathogens or mis-located cellular DNA. Upon DNA binding, cGAS utilizes GTP and ATP as substrates to synthesize cGAMP, leading to MITA-mediated innate immune response. In this study, we identified the phosphatase PPP6C as a negative regulator of cGASmediated innate immune response. PPP6C is constitutively associated with cGAS in un-stimulated cells. DNA virus infection causes rapid disassociation of PPP6C from cGAS, resulting in phosphorylation of human cGAS S435 or mouse cGAS S420 in its catalytic pocket. Mutation of this serine residue of cGAS impairs its ability to synthesize cGAMP upon DNA virus infection. In vitro experiments indicate that S420-phosphorylated mcGAS has higher affinity to GTP and enzymatic activity. PPP6Cdeficiency promotes innate immune response to DNA virus in various cells. Our findings suggest that PPP6Cmediated dephosphorylation of a catalytic pocket serine residue of cGAS impairs its substrate binding activity and innate immune response, which provides a mechanism for keeping the DNA sensor cGAS inactive in the absence of infection to avoid autoimmune response.
Key words: DNA virus; PPP6C; cGAS; innate immune response; phosphorylation; substrate binding
Mi Li , Hong-Bing Shu . Dephosphorylation of cGAS by PPP6C impairs its substrate binding activity and innate antiviral response[J]. Protein & Cell, 2020 , 11(8) : 584 -599 . DOI: 10.1007/s13238-020-00729-3
1 |
Akira S, Uematsu S, Takeuchi O (2006) Pathogen recognition and innate immunity. Cell 124:783–801
|
2 |
An J, Durcan L, Karr RM, Briggs TA, Rice GI, Teal TH, Woodward JJ, Elkon KB (2017) Expression of cyclic GMP-AMP synthase in patients with systemic lupus erythematosus. Arthritis Rheumatol 69:800–807
|
3 |
Bonilla X, Parmentier L, King B, Bezrukov F, Kaya G, Zoete V, Seplyarskiy VB, Sharpe HJ, McKee T, Letourneau A
|
4 |
Brautigan DL, Shenolikar S (2018) Protein serine/threonine phosphatases: keys to unlocking regulators and substrates. Annu Rev Biochem 87:921–964
|
5 |
Dai J, Huang YJ, He X, Zhao M, Wang X, Liu ZS, Xue W, Cai H, Zhan XY, Huang SY
|
6 |
Fang C, Wei X, Wei Y (2016) Mitochondrial DNA in the regulation of innate immune responses. Protein Cell 7:11–16
|
7 |
Gao P, Ascano M, Wu Y, Barchet W, Gaffney BL, Zillinger T, Serganov AA, Liu Y, Jones RA, Hartmann G
|
8 |
Gray EE, Treuting PM, Woodward JJ, Stetson DB (2015) Cutting edge: cGAS is required for lethal autoimmune disease in the Trex1-deficient mouse model of aicardi-goutieres syndrome.J Immunol 195:1939–1943
|
9 |
Harding SM, Benci JL, Irianto J, Discher DE, Minn AJ, Greenberg RA (2017) Mitotic progression following DNA damage enables pattern recognition within micronuclei. Nature 548:466–470
|
10 |
Hiratsuka T (1975) 2’ (or 3’)-O-(2, 4, 6-trinitrophenyl)adenosine 5’-triphosphate as a probe for the binding site of heavy meromyosin ATPase. J Biochem 78:1135–1147
|
11 |
Hodis E, Watson IR, Kryukov GV, Arold ST, Imielinski M, Theurillat JP, Nickerson E, Auclair D, Li L, Place C
|
12 |
Hooy R, Sohn J (2019) A pyrophosphatase-coupled assay to monitor the NTase activity of cGAS. Methods Enzymol 625:77–86
|
13 |
Hosing AS, Valerie NC, Dziegielewski J, Brautigan DL, Larner JM (2012) PP6 regulatory subunit R1 is bidentate anchor for targeting protein phosphatase-6 to DNA-dependent protein kinase. J Biol Chem 287:9230–9239
|
14 |
Hu MM, Shu HB (2018) Cytoplasmic Mechanisms of Recognition and Defense of Microbial Nucleic Acids. Annu Rev Cell Dev Biol 34:357–379
|
15 |
Hu MM, Shu HB (2019) Innate immune response to cytoplasmic DNA: mechanisms and diseases. Annu Rev Immunol
|
16 |
Hu MM, Yang Q, Xie XQ, Liao CY, Lin H, Liu TT, Yin L, Shu HB (2016) Sumoylation promotes the stability of the DNA sensor cGAS and the adaptor STING to regulate the kinetics of response to DNA virus. Immunity 45:555–569
|
17 |
Hu MM, He WR, Gao P, Yang Q, He K, Cao LB, Li S, Feng YQ, Shu HB (2019) Virus-induced accumulation of intracellular bile acids activates the TGR5-beta-arrestin-SRC axis to enable innate antiviral immunity. Cell Res 29:193–205
|
18 |
Ishikawa H, Barber GN (2008) STING is an endoplasmic reticulum adaptor that facilitates innate immune signalling. Nature 455:674–678
|
19 |
Janeway CA Jr, Medzhitov R (2002) Innate immune recognition. Annu Rev Immunol 20:197–216
|
20 |
Kajino T, Ren H, Iemura S, Natsume T, Stefansson B, Brautigan DL, Matsumoto K, Ninomiya-Tsuji J (2006) Protein phosphatase 6 down-regulates TAK1 kinase activation in the IL-1 signaling pathway. J Biol Chem 281:39891–39896
|
21 |
Li X, Shu C, Yi G, Chaton CT, Shelton CL, Diao J, Zuo X, Kao CC, Herr AB, Li P (2013) Cyclic GMP-AMP synthase is activated by double-stranded DNA-induced oligomerization. Immunity 39:1019–1031
|
22 |
Liu Y, Jesus AA, Marrero B, Yang D, Ramsey SE, Sanchez GAM, Tenbrock K, Wittkowski H, Jones OY, Kuehn HS
|
23 |
Liu S, Cai X, Wu J, Cong Q, Chen X, Li T, Du F, Ren J. Wu YT, Grishin NV
|
24 |
Liu H, Zhang H, Wu X, Ma D, Wu J, Wang L, Jiang Y, Fei Y, Zhu C, Tan R
|
25 |
Long L, Deng Y, Yao F, Guan D, Feng Y, Jiang H, Li X, Hu P, Lu X, Wang H
|
26 |
Luo WW, Shu HB (2018) Delicate regulation of the cGAS-MITAmediated innate immune response. Cell Mol Immunol 15:666–675
|
27 |
Mackenzie KJ, Carroll P, Martin CA, Murina O, Fluteau A, Simpson DJ, Olova N, Sutcliffe H, Rainger JK, Leitch A
|
28 |
Ogoh H, Tanuma N, Matsui Y, Hayakawa N, Inagaki A, Sumiyoshi M, Momoi Y, Kishimoto A, Suzuki M, Sasaki N
|
29 |
Pirman NL, Barber KW, Aerni HR, Ma NJ, Haimovich AD, Rogulina S, Isaacs FJ, Rinehart J (2015) A flexible codon in genomically recoded Escherichia coli permits programmable protein phosphorylation. Nat Commun 6:8130
|
30 |
Shang J. Xia T, Han QQ, Zhao X, Hu MM, Shu HB, Guo L (2018) Quantitative proteomics identified TTC4 as a TBK1 interactor and a positive regulator of SeV-induced innate immunity. Proteomics 18:1
|
31 |
Sun L, Wu J, Du F, Chen X, Chen ZJ (2013) Cyclic GMP-AMP synthase is a cytosolic DNA sensor that activates the type I interferon pathway. Science 339:786–791
|
32 |
Sun W, Li Y, Chen L, Chen H, You F, Zhou X, Zhou Y, Zhai Z, Chen D, Jiang Z (2009) ERIS, an endoplasmic reticulum IFN stimulator,activates innate immune signaling through dimerization. ProcNatl Acad Sci USA 106:8653–8658
|
33 |
West AP, Khoury-Hanold W, Staron M, Tal MC, Pineda CM, Lang SM, Bestwick M, Duguay BA, Raimundo N, MacDuff DA
|
34 |
Wies E, Wang MK, Maharaj NP, Chen K, Zhou S, Finberg RW, Gack MU (2013) Dephosphorylation of the RNA sensors RIG-I and MDA5 by the phosphatase PP1 is essential for innate immune signaling. Immunity 38:437–449
|
35 |
Willard FS, Kimple AJ, Johnston CA, Siderovski DP (2005) A direct fluorescence-based assay for RGS domain GTPase accelerating activity. Anal Biochem 340:341–351
|
36 |
Wu J, Sun L, Chen X, Du F, Shi H, Chen C, Chen ZJ (2013) Cyclic GMP-AMP is an endogenous second messenger in innate immune signaling by cytosolic DNA. Science 339:826–830
|
37 |
Xia P, Wang S, Gao P, Gao G, Fan Z (2016) DNA sensor cGASmediated immune recognition. Protein Cell 7:777–791
|
38 |
Xia T, Yi XM, Wu X, Shang J, Shu HB (2019) PTPN1/2-mediated dephosphorylation of MITA/STING promotes its 20S proteasomal degradation and attenuates innate antiviral response. Proc Natl Acad Sci USA 116:20063–20069
|
39 |
Xiong M, Wang S, Wang YY, Ran Y (2018) The regulation of cGAS. Virol Sin 33:117–124
|
40 |
Yan BR, Zhou L, Hu MM, Li M, Lin H, Yang Y, Wang YY, Shu HB (2017) PKACs attenuate innate antiviral response by phosphorylating VISA and priming it for MARCH5-mediated degradation. PLoS Pathog 13:e1006648
|
41 |
Zhan Z, Cao H, Xie X, Yang L, Zhang P, Chen Y, Fan H, Liu Z, Liu X (2015) Phosphatase PP4 negatively regulates type I IFN production and antiviral innate immunity by dephosphorylating and deactivating TBK1. J Immunol 195:3849–3857
|
42 |
Zhong B, Yang Y, Li S, Wang YY, Li Y, Diao F, Lei C, He X, Zhang L, Tien P
|
43 |
Zhong J, Liao J, Liu X, Wang P, Liu J, Hou W, Zhu B, Yao L, Wang J, Li J
|
/
〈 | 〉 |