The Many Ways to Deal with STING

Abstract

The stimulator of interferon genes (STING) is an adaptor protein involved in the activation of IFN-β and many other genes associated with the immune response activation in vertebrates. STING induction has gained attention from different angles such as the potential to trigger an early immune response against different signs of infection and cell damage, or to be used as an adjuvant in cancer immune treatments. Pharmacological control of aberrant STING activation can be used to mitigate the pathology of some autoimmune diseases. The STING structure has a well-defined ligand binding site that can harbor natural ligands such as specific purine cyclic di-nucleotides (CDN). In addition to a canonical stimulation by CDNs, other non-canonical stimuli have also been described, whose exact mechanism has not been well defined. Understanding the molecular insights underlying the activation of STING is important to realize the different angles that need to be considered when designing new STING-binding molecules as therapeutic drugs since STING acts as a versatile platform for immune modulators. This review analyzes the different determinants of STING regulation from the structural, molecular, and cell biology points of view.

Keywords: IFN; STING; antiviral response.

Figure 1

cGAS-STING canonical pathway: Schematic representation of STING canonical activation including post-translational modifications, modulator proteins, and main effectors involved in the production of Type I IFN and proinflammatory cytokines.

Figure 2

Non-canonical activations of STING. 1. Release of nuclear or mitochondrial DNA into the cytoplasm during viral infection or after cell damage. 2. Activation of the STING–TBK1–IRF3 axis by ER-stress inducers such as ethanol. 3. cGAS-independent activation of STING triggered by membrane fusion. 4. Activation of STING by RIG-I-like Receptors (RLRs) after detection of viral RNA in a MAVS-independent process. 5. Ca²⁺ depletion in the ER triggers activation of STIM1 sensor, an inhibitor of STING, and induces its migration to join the calcium channel ORAI1, increasing STING activity. 6. IFI16 pathway of nuclear-damaged DNA detection in the absence of cGAS activation. Question marks (?) represent “unknown activation mechanism”.

Figure 3

(A) Amino acid sequence of human STING. Rainbow color coding indicates the direction from the N-terminal (blue) to the C-terminal (red) ends. (B) Schematic representation of the tertiary structure of a human STING monomer with the same rainbow color coding. Amino acid numbering of the secondary structure has been taken from the human STING structure under PDB code 7SII. (C) Left. PyMOL cartoon representation of human STING as found in PDB code 7SII with rainbow color coding and the 2′3′-cGAMP colored as red sticks. The transmembrane domain is highlighted in yellow. Right. PyMOL representation of the closed conformation of the human STING dimer extracted from PDB code 7SII in which one of the monomers is shown as cartoons with the rainbow color coding and the other is shown as a surface to account for the intertwining of the transmembrane α-helixes. For the sake of clarity, the 2′3′-cGAMP is not shown. (D) PyMOL representation of the superimposition of two human STING monomers in the open (PDB code 6NT5) and closed (PDB code 7SII) conformations colored in light pink and blue, respectively, depicting the 180° rotation of the cytosolic domain over the transmembrane domain of each of the monomers as seen from the side (left) and top (right) after rotation of the superimposed monomers 90° in the Z-axis. (E) PyMOL representation of the open (left) and closed (right) conformation of human STING as found in PDB codes 6NT7 and 7SII, respectively. In both conformations, each monomer is colored green (chain A) and cyan (chain B). For the sake of clarity, in the closed conformation, only the 2′3′-cGAMP molecule is shown as red sticks, whereas ligand 9IM is not shown in this image.

Figure 4

The amino acid sequence of the original human STING and the corresponding mutations that confer gain (yellow) or loss (blue) of the protein function.

Figure 5

CDN finding to STING: (A) PyMOL representation of the binding mode of 2′3′-cGAMP (yellow) to human STING as found in PDB code 7SII. The two side chains of R232 and R238 of both monomers establish ionic interactions with the negatively charged phosphates in the phosphodiester bond. Additionally, and given the position of the side chain of R230 in both monomers, the guanidinium group establishes a π-cation interaction with the nitrogenated base of the 2′3′-cGAMP of the opposite monomer. For the sake of clarity, none of the hydrogen atoms nor the water molecules present in the PDB are shown, and hydrogen bonds are shown as dashed lines. (B) PyMOL representation of the binding mode of cdiGMP to the Myroides sp. ZB35 bacterial STING as found in PDB code 7EBL. The side chain of R230 of both monomers establishes very distinct hydrogen bonding interactions that allow the recognition of the guanidine of the cdiGMP, while the side chain of T254 stabilizes the positions of the phosphates in the phosphodiester bond by establishing hydrogen bonds.

Figure 6

2D structure of the modified CDNs. Compounds that can be found deposited in the PDB in complex with hSTING are identified by the ligand name of the PDB structure and those that are not found crystalized with human STING are identified with the name given in the original article.

Figure 7

2D structure of the non-CDNs. Compounds that can be found deposited in the PDB in complex with human STING are identified by the ligand name of the PDB structure, those that are not found crystalized with human STING are identified with the name given in the original article.

Figure 8

2D structure of the palmitoylation inhibitors and the STING-directed PROTACs. Compounds are identified by the name given in the original article.