The Genetic, Molecular, And Cellular Bases Of Unidentified Primary Immunodeficiencies

The immune system is inseparable from every part of human biology. From cell intrinsic mechanisms of pathogen recognition to multi-cellular interactions over vast ranges of time and space, the immune system is both essential for protection from infection and central to the pathogenesis of many diseases. Thus understanding it has long been a focus of biomedical research. While in vitro molecular, biochemical, and cellular techniques as well as complex genetically modified animal models have been developed, these approaches still only approximate true human disease and in vivo human biology. Primary immunodeficiencies are inborn genetic defects of immunity and present rare opportunities to observe, study, and understand how genetic perturbations impact human immunity directly. I therefore clinically and genetically analyzed three patient families with unidentified primary immunodeficiencies. Using whole exome sequencing coupled with in vitro and in vivo biochemical and cellular assays, I identified two novel genetic etiologies of primary immunodeficiency. I first identified de novo missense mutations in GNAI2, the gene encoding the ubiquitously expressed heterotrimeric G-protein Gαi2, in 2 families with life-threating multi-organ system autoimmunity and immunodeficiency to mucocutaneous infections. Gαi2 is essential for chemokine mediated leukocyte migration as well as regulating development, inflammation, and metabolism in the immune system and beyond. The heterozygous dominant gain-of-function patient proteins impaired chemokine signaling and chemotaxis in addition to augmenting T cell activation by constitutively activating costimulatory pathways and reducing the requirement for T cell costimulation. I also identified homozygous missense mutations in IFIH1, the gene encoding the cytosolic pattern recognition receptor of dsRNA MDA5, in the third family of study. The affected individual presented with recurrent severe respiratory infections with RNA viruses including human rhinovirus, coronaviruses (HKU1, OC43, NL63), influenza virus, and respiratory syncytial virus. The mutant protein lost the ability to bind dsRNA and failed to initiate antiviral interferon-β and pro-inflammatory NF-κB responses. Using gene knockdown and gene editing in immortalized and patient derived cell lines, I demonstrated an essential role for MDA5 in restricting rhinovirus infection in human respiratory epithelium. Thus this work demonstrates the power of human genetics to identify disease causing mutations in rare individuals and reveal how the immune system uses molecules involved in cell migration, activation, and nucleic acid sensing to robustly protect us from virus infections without causing autoimmunity