Final Report Summary - AIM2 INFLAMMASOME (Cytosolic recognition of foreign nucleic acids: Molecular and functional characterization of AIM2, a central player in DNA-triggered inflammasome activation)
In this grant project, we have studied the role of AIM2-dependent and –independent (see extension of the initial grant proposal) cytosolic DNA recognition pathways of the mammalian innate immune system.
AIM2 is a member of the so-called PYHIN family of proteins that harbors an N-terminal Pyrin domain as well as a C-terminal, ligand-binding HIN domain. AIM2 is expressed in the cytoplasm of myeloid cells and triggers the formation of an inflammasome complex upon cytosolic DNA recognition. Given the fact that AIM2 recognizes double stranded DNA independent of its sequence composition, this receptor cannot discriminate self from non-self. In fact, DNA of both exogenous (e.g. bacterial or virus) as well as endogenous sources can result in AIM2 activation upon cytosolic delivery. In this grant proposal, we set out to study the mechanisms of AIM2 activation and the physiological context and role of its activation in both microbial infection as well as sterile inflammation. In the course of these studies we could decipher the minimal ligand requirements for AIM2 activation to be in the range of 80 bp of double stranded DNA, with AIM2 binding showing positive cooperativity. In conjunction with the crystal structure of the HIN domain of AIM2 being solved, this provided a working model of AIM2 being activated by double stranded DNA. Moreover, we and others could show that AIM2 detects microbial DNA in the context of bacterial or viral infection. For example in the case of Listeria monocytogenes infection, bacterial DNA gains access to the cytoplasm where it triggers AIM2 inflammasome activation and subsequent cytokine maturation. Given the fact that AIM2 is a bona fide DNA receptor, this study provided indirect proof for the long-discussed hypothesis that microbial DNA can be sensed within the cytoplasm. To explore the role of AIM2 in the context of sterile inflammation, we turned to a mouse model, in which we could investigate the role of aberrant, cytosolic DNA recognition without exogenous manipulation. To do so, we studied the model of DNase II deficiency, in which the failure to degrade DNA in phagocytic cells results in progressive joint inflammation and subsequent polyarthritis. Studying this mouse model in the context of AIM2 deficiency, we observed a largely reduced activation of the inflammasome pathway that was associated with a strong decrease in polyarthritis. Given the fact that this mouse model results in the translocation of endogenous DNA species into the cytoplasm of AIM2 expressing cells, a direct link of AIM2 recognizing endogenous DNA could be established. In follow up studies could substantiate the role of AIM2 as a sensor for endogenous
While AIM2 is non-redundantly required to convey inflammasome activation in response to cytosolic DNA, (an) additional DNA sensing pathway(s) exist(s) in the cytosol that regulates the expression of pro-inflammatory and antiviral factors. A crucial component of this pathway was recently identified to be cGAS, a cytosolic receptor that exerts nucleotidyltransferase activity upon the recognition of DNA. Studying this receptor, we were able to identify the nature of the secondary messenger molecule that is produced upon cytosolic DNA recognition by cGAS. To this end, we could show that cGAS produces the mixed cyclic dinucleotide cGAMP with an unorthodox combination of two different phosphodiester bonds (2’-5’ and 3’-5’). At the same time we could contribute to the elucidation of the structural mechanisms of cGAS sensing DNA and of its conformational switch leading to cGAMP production. Upon synthesis, cGAMP binds to and activates the ER-resident receptor STING, which results in the activation of transcription factors that lead to the expression of antiviral genes, including the important family of type I IFN cytokines. Interestingly, we found that this 2nd messenger-dependent signaling cascade also exerts functional consequences beyond cell intrinsic immunity. To this effect, we could show that cGAMP is horizontally transferred from one cell to another cell via gap junctions. Of note, this in trans signaling cascade initiates antiviral protection a lot quicker than cytokine-dependent induction of antiviral effector mechanisms.
Altogether, with our studies on AIM2-dependent and –independent cytosolic DNA sensing mechanisms we could contribute to our understanding of innate immune recognition pathways. Understanding these mechanisms is not only of great interest from a basic research point of view, but also provides important information regarding potential signaling nodes that can be targeted for therapeutic intervention.
AIM2 is a member of the so-called PYHIN family of proteins that harbors an N-terminal Pyrin domain as well as a C-terminal, ligand-binding HIN domain. AIM2 is expressed in the cytoplasm of myeloid cells and triggers the formation of an inflammasome complex upon cytosolic DNA recognition. Given the fact that AIM2 recognizes double stranded DNA independent of its sequence composition, this receptor cannot discriminate self from non-self. In fact, DNA of both exogenous (e.g. bacterial or virus) as well as endogenous sources can result in AIM2 activation upon cytosolic delivery. In this grant proposal, we set out to study the mechanisms of AIM2 activation and the physiological context and role of its activation in both microbial infection as well as sterile inflammation. In the course of these studies we could decipher the minimal ligand requirements for AIM2 activation to be in the range of 80 bp of double stranded DNA, with AIM2 binding showing positive cooperativity. In conjunction with the crystal structure of the HIN domain of AIM2 being solved, this provided a working model of AIM2 being activated by double stranded DNA. Moreover, we and others could show that AIM2 detects microbial DNA in the context of bacterial or viral infection. For example in the case of Listeria monocytogenes infection, bacterial DNA gains access to the cytoplasm where it triggers AIM2 inflammasome activation and subsequent cytokine maturation. Given the fact that AIM2 is a bona fide DNA receptor, this study provided indirect proof for the long-discussed hypothesis that microbial DNA can be sensed within the cytoplasm. To explore the role of AIM2 in the context of sterile inflammation, we turned to a mouse model, in which we could investigate the role of aberrant, cytosolic DNA recognition without exogenous manipulation. To do so, we studied the model of DNase II deficiency, in which the failure to degrade DNA in phagocytic cells results in progressive joint inflammation and subsequent polyarthritis. Studying this mouse model in the context of AIM2 deficiency, we observed a largely reduced activation of the inflammasome pathway that was associated with a strong decrease in polyarthritis. Given the fact that this mouse model results in the translocation of endogenous DNA species into the cytoplasm of AIM2 expressing cells, a direct link of AIM2 recognizing endogenous DNA could be established. In follow up studies could substantiate the role of AIM2 as a sensor for endogenous
While AIM2 is non-redundantly required to convey inflammasome activation in response to cytosolic DNA, (an) additional DNA sensing pathway(s) exist(s) in the cytosol that regulates the expression of pro-inflammatory and antiviral factors. A crucial component of this pathway was recently identified to be cGAS, a cytosolic receptor that exerts nucleotidyltransferase activity upon the recognition of DNA. Studying this receptor, we were able to identify the nature of the secondary messenger molecule that is produced upon cytosolic DNA recognition by cGAS. To this end, we could show that cGAS produces the mixed cyclic dinucleotide cGAMP with an unorthodox combination of two different phosphodiester bonds (2’-5’ and 3’-5’). At the same time we could contribute to the elucidation of the structural mechanisms of cGAS sensing DNA and of its conformational switch leading to cGAMP production. Upon synthesis, cGAMP binds to and activates the ER-resident receptor STING, which results in the activation of transcription factors that lead to the expression of antiviral genes, including the important family of type I IFN cytokines. Interestingly, we found that this 2nd messenger-dependent signaling cascade also exerts functional consequences beyond cell intrinsic immunity. To this effect, we could show that cGAMP is horizontally transferred from one cell to another cell via gap junctions. Of note, this in trans signaling cascade initiates antiviral protection a lot quicker than cytokine-dependent induction of antiviral effector mechanisms.
Altogether, with our studies on AIM2-dependent and –independent cytosolic DNA sensing mechanisms we could contribute to our understanding of innate immune recognition pathways. Understanding these mechanisms is not only of great interest from a basic research point of view, but also provides important information regarding potential signaling nodes that can be targeted for therapeutic intervention.