Dissecting phenotypic heterogeneity in endosomal TLR-driven autoimmunity using human UNC93B1 allelic variants

Some people get lupus because of damaging changes to just one gene among the ~20,000 in our genome. Studying the immune systems of these individuals can teach us important and general lessons that apply to all people living with lupus. Dr. Huizar’s work centers on a gene called UNC93B1, which helps prevent our immune system from being activated by pieces of our own cells (specifically, RNA and DNA, aka nucleic acids). People and mice with mutations in the UNC93B1 gene develop lupus and related autoimmune conditions. Fascinatingly, mutations in different parts of the gene cause different patterns of disease in mice, including the development of antibodies against distinct “self” proteins. Dr. Huizar hopes to use these mutations to understand how individual people with lupus can experience their condition in such vastly different ways, including why certain organs become damaged and not others.

What this means for people with lupus: The results of this work will teach us how genetic mutations influence the autoantibody response, and bring us closer to personalizing lupus therapies to meet the specific needs of individual patients.

Human missense variants in the TLR-regulating chaperone protein UNC93B1 are associated with systemic lupus, cutaneous lupus and neuroinflammatory disorders, and are sufficient to drive systemic autoimmune disease when knocked-in to mice. Synthesizing the spectrum of 13 reported human UNC93B1 allelic variants, we selected 3 representative variant alleles in endosomal loop 1, cytoplasmic loop 6 or the cytoplasmic C terminus of UNC93B1 for further mechanistic investigation in 3 corresponding knock-in mouse lines. We used a phage-display peptide library spanning the entire mouse proteome to identify novel self antigens targeted by autoreactive serum IgG in these mice (Phage-display Immunoprecipitation-Sequencing, PhIPseq), and have found that the top antigenic targets are distinct across the 3 lines. Underpinning this difference in the “autoreactome” is a second interesting observation: while every allele enhances macrophage TLR7 and TLR9 responses, only the loop 1 and C-terminal alleles enhance B cell responses. On this basis, we hypothesize that human UNC93B1missense variants modulate innate nucleic acid sensing in a cell-type-specific manner, and that the pattern of cell types activated may determine the autoantibody repertoire and organ tropism. In the proposed work, I will 1) characterize the serum autoantibody response to Unc93b1-allele-specific self antigens, 2) define the cell-type-specific requirements for mutant Unc93b1 expression vis a vis allele-specific autoantibody production and disease, and 3) identify the B cells reactive to allele-specific self-antigens, clone their B cell receptors and compare the tissues targeted by the encoded antibodies ex vivo and in vivo. Overall, this work will apply new technologies to probe novel, patient-derived mutations in a heretofore understudied axis of innate immunity and lupus biology – UNC93B1-mediated TLR regulation – with the goal of defining the mechanistic underpinnings of phenotypic heterogeneity in SLE.