TY - JOUR
T1 - Ubiquitin-like conjugation by bacterial cGAS enhances anti-phage defence
AU - Jenson, Justin M.
AU - Li, Tuo
AU - Du, Fenghe
AU - Ea, Chee Kwee
AU - Chen, Zhijian J.
N1 - Funding Information:
We thank the UTSW Structural Biology Laboratory at UT Southwestern Medical Center for support with X-ray crystallographic studies, especially D. Tomchick, Z. Chen and H. Aronovich. Results shown in this report are derived from work performed at Argonne National Laboratory, Structural Biology Center (SBC) at the Advanced Photon Source. SBC is operated by the US Department of Energy, Office of Biological and Environmental Research under contract DE-AC02-06CH11357. We thank C. Brautigam and S. Tso from the UTSW Macromolecular Biophysics Resource for assistance with ITC and AUC. We thank members of the Chen laboratory, especially B. Ramirez, I. Dehghan and X. Chen for assistance maintaining the mass spectrometry facility. We thank J. Cabrera for assisting with graphics. Work in the Chen laboratory is supported by grants from the National Cancer Institute (U54CA244719), Welch Foundation (I-1389) and Cancer Prevention and Research Institute of Texas (RP180725). J.M.J. was supported by a postdoctoral fellowship from the Cancer Research Institute and is supported by the Dermatology Research Training Program T32 Grant T32AR065969. Z.J.C. is an investigator of the Howard Hughes Medical Institute.
Funding Information:
We thank the UTSW Structural Biology Laboratory at UT Southwestern Medical Center for support with X-ray crystallographic studies, especially D. Tomchick, Z. Chen and H. Aronovich. Results shown in this report are derived from work performed at Argonne National Laboratory, Structural Biology Center (SBC) at the Advanced Photon Source. SBC is operated by the US Department of Energy, Office of Biological and Environmental Research under contract DE-AC02-06CH11357. We thank C. Brautigam and S. Tso from the UTSW Macromolecular Biophysics Resource for assistance with ITC and AUC. We thank members of the Chen laboratory, especially B. Ramirez, I. Dehghan and X. Chen for assistance maintaining the mass spectrometry facility. We thank J. Cabrera for assisting with graphics. Work in the Chen laboratory is supported by grants from the National Cancer Institute (U54CA244719), Welch Foundation (I-1389) and Cancer Prevention and Research Institute of Texas (RP180725). J.M.J. was supported by a postdoctoral fellowship from the Cancer Research Institute and is supported by the Dermatology Research Training Program T32 Grant T32AR065969. Z.J.C. is an investigator of the Howard Hughes Medical Institute.
Publisher Copyright:
© 2023, The Author(s).
PY - 2023/4/13
Y1 - 2023/4/13
N2 - cGAS is an evolutionarily conserved enzyme that has a pivotal role in immune defence against infection1–3. In vertebrate animals, cGAS is activated by DNA to produce cyclic GMP–AMP (cGAMP)4,5, which leads to the expression of antimicrobial genes6,7. In bacteria, cyclic dinucleotide (CDN)-based anti-phage signalling systems (CBASS) have been discovered8–11. These systems are composed of cGAS-like enzymes and various effector proteins that kill bacteria on phage infection, thereby stopping phage spread. Of the CBASS systems reported, approximately 39% contain Cap2 and Cap3, which encode proteins with homology to ubiquitin conjugating (E1/E2) and deconjugating enzymes, respectively8,12. Although these proteins are required to prevent infection of some bacteriophages8, the mechanism by which the enzymatic activities exert an anti-phage effect is unknown. Here we show that Cap2 forms a thioester bond with the C-terminal glycine of cGAS and promotes conjugation of cGAS to target proteins in a process that resembles ubiquitin conjugation. The covalent conjugation of cGAS increases the production of cGAMP. Using a genetic screen, we found that the phage protein Vs.4 antagonized cGAS signalling by binding tightly to cGAMP (dissociation constant of approximately 30 nM) and sequestering it. A crystal structure of Vs.4 bound to cGAMP showed that Vs.4 formed a hexamer that was bound to three molecules of cGAMP. These results reveal a ubiquitin-like conjugation mechanism that regulates cGAS activity in bacteria and illustrates an arms race between bacteria and viruses through controlling CDN levels.
AB - cGAS is an evolutionarily conserved enzyme that has a pivotal role in immune defence against infection1–3. In vertebrate animals, cGAS is activated by DNA to produce cyclic GMP–AMP (cGAMP)4,5, which leads to the expression of antimicrobial genes6,7. In bacteria, cyclic dinucleotide (CDN)-based anti-phage signalling systems (CBASS) have been discovered8–11. These systems are composed of cGAS-like enzymes and various effector proteins that kill bacteria on phage infection, thereby stopping phage spread. Of the CBASS systems reported, approximately 39% contain Cap2 and Cap3, which encode proteins with homology to ubiquitin conjugating (E1/E2) and deconjugating enzymes, respectively8,12. Although these proteins are required to prevent infection of some bacteriophages8, the mechanism by which the enzymatic activities exert an anti-phage effect is unknown. Here we show that Cap2 forms a thioester bond with the C-terminal glycine of cGAS and promotes conjugation of cGAS to target proteins in a process that resembles ubiquitin conjugation. The covalent conjugation of cGAS increases the production of cGAMP. Using a genetic screen, we found that the phage protein Vs.4 antagonized cGAS signalling by binding tightly to cGAMP (dissociation constant of approximately 30 nM) and sequestering it. A crystal structure of Vs.4 bound to cGAMP showed that Vs.4 formed a hexamer that was bound to three molecules of cGAMP. These results reveal a ubiquitin-like conjugation mechanism that regulates cGAS activity in bacteria and illustrates an arms race between bacteria and viruses through controlling CDN levels.
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U2 - 10.1038/s41586-023-05862-7
DO - 10.1038/s41586-023-05862-7
M3 - Article
C2 - 36848932
AN - SCOPUS:85151366999
SN - 0028-0836
VL - 616
SP - 326
EP - 331
JO - Nature
JF - Nature
IS - 7956
ER -