The role of DNASE1L3 and its endogenous substrate, chromatin in apoptotic cell microparticles, in human systemic lupus erythematosus and experimental systemic sclerosis

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Antibodies to DNA and chromatin drive autoimmunity in systemic lupus erythematosus (SLE). Major questions of SLE pathogenesis concern the physical form of DNA that can be recognized by autoreactive B cells and the mechanisms that prevent the breakdown of self-tolerance to DNA.Null mutations and hypomorphic variants of the secreted deoxyribonuclease DNASE1L3 are linked to familial and sporadic SLE, respectively. Boris Reizis and colleagues provided a mechanistic explanation for this observation:They identified chromatin from microparticles as latent self-antigen for autoreactive B cells in experimental SLE. Microparticles contain genomic DNA from apoptotic cells. Apparently, circulating DNASE1L3 is uniquely capable of digesting the DNA in microparticles, thereby restricting its antigenicity.Importantly, this model describes a cell-extrinsic mechanism of self-tolerance that involves a secreted enzyme and therefore can be developed for therapeutic purposes.Moreover, the same hypomorphic DNASE1L3 variant is also linked to systemic sclerosis (SSc) and similar types of DNA-driven immune complexes are characteristic for both, SLE and SSc.Thus, the overall goal of this project is to systematically test the involvement of DNASE1L3 and its endogenous DNA substrate in SLE and SSc. This goal will be pursued through two research objectives.Objective 1 explores the role of microparticle DNA as an antigen in human SLE.DNASE1L3 activity, antibody binding to microparticle DNA and microparticle DNA content will be measured in more than 400 well-characterized SLE patients and matched healthy donors. The study-population will allow comparing samples from clinical versus preclinical patients.Following hypotheses will be explored: i) Binding of IgG antibodies to microparticle DNA occurs in a substantial fraction of SLE patients, but not in healthy individuals, ii) DNA content in circulating microparticles is increased in SLE patients and correlates with binding of antibodies to microparticle DNA and reduced DNASE1L3 activity, and iii) Antibodies to microparticles-associated DNA and chromatin develop early in the disease and correlate with or even predict its severity.Objective 2 explores the role and therapeutic utility of DNASE1L3 in murine models of SSc.The effect of DNASE1L3-deficiency on clinical and immunological parameters will be assessed in the bleomycin-induced mouse model, and in a genetic model with tight skin1 mice. To test the therapeutic utility of DNASE1L3, mice will be injected with an adenoviral vector encoding human DNASE1L3.In detail following hypotheses will be explored: i) DNASE1L3 deficient mice develop a more aggravated form of experimental SSc than wild type mice, ii) Increased microparticle DNA content, binding of antibodies to microparticle DNA and reduced DNASE1L3 activity correlate with diseases severity in experimental SSc, and iii) DNASE1L3 delivery prevents and reverts experimental SSc.