Characterization of cGAS homologs in innate and adaptive mucosal immunities in zebrafish gives evolutionary insights into cGAS‐STING pathway

Abstract

Cyclic GMP‐AMP synthase (cGAS) is one of the most‐characterized cytoplasmic DNA sensors in humans and other mammals. However, knowledge about cGAS homologs in nonmammalian species remains limited. In this study, we report the molecular and functional identification of two cGAS homologs, namely, DrcGASa and DrcGASb, from a zebrafish (Danio rerio) model. DrcGASa and DrcGASb share the same overall conservative structural architectures and functional domains/residues to mammalian cGASs. Both homologs synthesized a 2′3′‐cGAMP isomer but not a 3′3′‐cGAMP isomer via oligomerization in response to DNA stimulation. Overexpression of DrcGASa/b in HEK293T cells and zebrafish embryos significantly activated NF‐κB and IFN‐I signaling pathways in a STING‐dependent manner. Knockdown of DrcGASa or DrSTING impaired such activations, thereby reducing the host innate immunity against bacterial and viral infections. DrcGASa, but not DrcGASb, was involved in immunoglobulin Z‐mediated mucosal immunity in gill‐associated lymphoid tissue, suggesting differential functions between the two DrcGASs. This reaction was associated with the DrcGAS‐DrSTING‐IFNφ1 signaling axis in GALT’s γδ T cells. Our findings provide experimental evidence that a modern cGAS‐STING pathway that mainly participates in IFN‐mediated immunity originated from teleost fish based on the functional constraint of cGAS and STING proteins during vertebrate evolution.

Animal Models for the Study of Nucleic Acid Immunity: Novel Tools and New Perspectives

“…implying that other reported sensors may be poorly relevant in vivo. Activation of cGAS zebrafish isoforms also triggers synthesis of cGAMP and activation of STING-mediated IFN responses in vitro and in vivo [40]. These recent data contrast with previously reported DNA sensing mechanisms in zebrafish larvae.…”

“…Characterized zebrafish orthologues of mammalian cGAS were dispensable in this context [41], although STING was essential as in humans and mice [34,39]. These data are contrasted with the ability of zebrafish cGAS orthologues to synthesize cGAMP and activate STING-mediated IFN signalling in response to DNA stimulation [40].…”

Salmonid Antibacterial Immunity: An Aquaculture Perspective

“…To date, there are six classes of PRRs which are categorized based on shared structural elements: toll-like receptors (TLRs), C-type lectin receptors (CLRs), NOD-like receptors (NLRs), AIM2-like receptors (ALRs), RIG-I-like receptors (RLRs) and cGAS/STING receptors (reviewed in [ 41 ]). Though not as well understood as their mammalian counterparts, receptors from all but the ALRs have been identified in teleosts [ 43 , 44 , 45 , 46 , 47 ]. Although there are some similarities between mammalian and teleostean PRRs, some discrepancies do exist.…”

Cytosolic Sensors for Pathogenic Viral and Bacterial Nucleic Acids in Fish

“…cGAS is conserved in various species spanning from fish to mammals. In teleosts, the N-terminal region of the cGAS protein is markedly conserved, while the amino acids involved in the DNA-binding surface and cGAMP synthesis in the mab21 domain are also highly conserved [ 105 , 160 ].…”

“…Although, the zebrafish cGAS homologue has low amino acid identity to human cGAS (less than 35%), their similar activities were determined [ 179 ]. Liu and co-workers [ 160 ] reported that overexpression of zebrafish cGASa/b in HEK293T cells and zebrafish embryos significantly activated NF-κB and type I IFN signaling pathways in a STING-dependent manner, and that cGASa, but not cGASb, was involved in immunoglobulin Z-mediated mucosal immunity in gill-associated lymphoid tissue, suggesting differential functions between the two DrcGASs [ 160 ]. Interestingly, it was revealed that the ortholog of cGAS in grass carp, cGAS-like (cGASL) by interacting with STING down-regulates transcription of type I IFN gene and functions as a negative regulator of IFN response.…”