Finding 1: Because of our interest in patients with symptoms of atopy in the context of unidentified syndromes, we have been studying a series of families with cold urticaria, antibody deficiency, autoimmunity and granulomatous disease segregating in an autosomal dominant pattern. We have named this syndrome PLAID, as described in our report in the New England Journal of Medicine. Patients have low IgM and IgA, low NK cells, normal T cells and low to absent class-switched B cells. In collaboration with Dr. Daniel Kastner's lab, we found that the disease is caused by genomic deletions in PLCG2. The deletions are in an autoregulatory domain, leading to gain of PLCG2 enzymatic function but loss of distal signaling function, resulting in defective calcium flux and ERK phosphorylation in response to surface receptor cross-linking in B and NK cells, poor in vitro expansion and switched immunoglobulin secretion in response to SAC and CpG, and abnormal B-cell receptor editing in B-cells. We have found that the cold spontaneously activates certain cells in these patients, such as B cells and mast cells. We have since found that cold also activates neutrophils and monocytes from these patients, and have further described the neonatal onset rash, and subsequent adult-onset granulomatous disease in 7 patients with PLAID, which may be due to the cooler temperatures in the skin activating neutrophils and monocytes. In addition, we have found 4 new families with symptoms identical to PLAID, but who lack any mutations or deletions in PLCG2, suggesting a set of genes beyond PLCG2 may be capable of causing PLAID. A report describing these findings was published in JAMA Dermatology. In addition, in collaboration with Sue Pierce's lab (Laboratory of Immunogenetics, NIAID), we have also shown that the cSH2 domain which is mutated in PLAID may be responsible for stabilizing the B-cell receptor signalosome. Numerous proximal signaling molecules fail to activated normally with B-cell receptor cross-linking, despite the fact that in theory PLCG2 function is downstream of those molecules. The results of this study were published in Science Signaling. Finding 2: In our study of patients with immune deficiencies and atopic phenotypes, we have developed a flow-based assay for assessing multiple STAT responses to individual cytokines. Using this tool, in collaboration with Dr. Steve Holland's lab, we have found a family with invasive cryptosporidial infections, bronchiectasis and antibody deficiency with marked elevations in serum IgE. We found absent IL-21 signaling in these patients. In collaboration with Christoph Klein at the Gene Center of the University of Munich (LMU), we found a 6 bp in-frame deletion in the IL-21 receptor in these patients. Dr. Klein had followed two patients with similar phenotypes and found an independent mutation in the IL-21 receptor. The report of ours and Dr. Klein's patients was published in the Journal of Experimental Medicine, with an additional patient we identified subsequently in collaboration with Helen Su's lab (Laboratory of Host Defenses, NIAID) and published as part of a case series in Current Opinion in Pediatrics. The clinical and laboratory overlap between STAT3 loss-of-function patients, STAT1 gain-of-function patients, and IL-21r loss of function patients can be gleaned from these various reports, and provides the basis for ongoing investigation into these common pathways. This has included work we performed in collaboration with Warren Leonard's lab showing the effects of IL-21 signaling and activity when either STAT1 or STAT3 has gain or loss of function in patients. This work was published in PNAS. Finding 3: On a related topic, using whole exome sequencing on a cohort of patients with autoimmunity, lymphoproliferation, immune deficiency and short stature in collaboration with Helen Su and Mike Lenardo (Laboratory of Immunology, NIAID), we identified 4 patients with novel mutations in STAT3 that largely segregated with disease. At the same time an addition 9 patients from 8 different families were identified by our collaborators Megan Cooper at Washington University, Lisa Forbes and Asbjorg Stray-Pedersen at Baylor, and Sophie Hambleton at Newcastle University. We found that these mutations led to gain of STAT3 function, in contrast to AD-HIES patients who have loss of STAT3 function. We further showed that the phenotypes were likely due to suppression of STAT5 signaling leading to impaired regulatory T-cell formation and likely the short stature-- mimicking phenotypes seen in patients with loss of STAT5b function. Our results were published in Blood. Finding 4: Using whole exome sequencing, we found recessive mutations in phosphoglucomutase 3 (PGM3) which lead to a syndrome of severe allergy, immune deficiency and neurocognitive abnormalities. The patients have severe atopic dermatitis and high IgE, as well as high IgA and IgG, low CD8 and memory B-cell numbers, dysmyelination, and abnormally increased Th2 and Th17 cytokine production. The mutations in PGM3 lead to a reduction in normal protein glycosylation and segregate perfectly with disease in two different multiplex families affected. Of significant importance, exogenous N-acetyl glucosamine, a substrate upstream of PGM3, was able to correct an observed deficit in cytoplasmic UDP-N-acetyl glucosamine, suggesting that exogenous sugars and/or sugar amines might be able to correct the defect found in these patients. The results were published in the Journal of Allergy and Clinical Immunology. In addition, because of our finding that exogenous N-acetyl glucosamine could correct some of the cellular defects seen in PGM3 deficiency, we now have an approved protocol to administer oral n-acetylglucosamine and uridine supplements to patients with PGM3 deficiency. Accrual is beginning at the time of this report. Finding 5: In our efforts to describe familial inheritance of allergic disease, we have identified 25 families with an autosomal dominant inheritance of elevated basal tryptase levels in the absence of any signs of mastocytosis. Affected family members have cutaneous flushing, episodic gastrointestinal pain, food allergy and anaphylaxis, and, in a subset of patients, connective tissue abnormalties reminiscent of Ehlers-Danlos syndrome, hypermobility type. Patient basophils have a degranulation defect, perhaps due to anergy ex-vivo, and in the multiple families in whom bone marrow biopsies were done, there was a significant elevation in mast cell number, despite the complete lack of morphologic evidence or scale of mastocytosis. A report describing our experience with the first 9 families was published in the Journal of Allergy and Clinical Immunology. We have subsequently found an additional 30+ families, accruing 160 affected patients. The genetic underpinnings of this disorder are being deciphered at this time.

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Natarajan, Mukil; Hsu, Amy P; Weinreich, Michael A et al. (2018) Aspergillosis, eosinophilic esophagitis, and allergic rhinitis in signal transducer and activator of transcription 3 haploinsufficiency. J Allergy Clin Immunol 142:993-997.e3
Lyons, Jonathan J; Milner, Joshua D (2018) Primary atopic disorders. J Exp Med 215:1009-1022
Sabato, Vito; Chovanec, Jack; Faber, Margaretha et al. (2018) First Identification of an Inherited TPSAB1 Quintuplication in a Patient with Clonal Mast Cell Disease. J Clin Immunol 38:457-459
Lyons, Jonathan J; Stotz, Stephanie C; Chovanec, Jack et al. (2018) A common haplotype containing functional CACNA1H variants is frequently coinherited with increased TPSAB1 copy number. Genet Med 20:503-512
Milner, Joshua D (2018) TCR Signaling Abnormalities in Human Th2-Associated Atopic Disease. Front Immunol 9:719
Schussler, Edith; Yang, Amy; Lyons, Jonathan J et al. (2018) Persistent tryptase elevation in a patient with Gaucher disease. J Allergy Clin Immunol Pract 6:697-699
Valent, Peter; Akin, Cem; Bonadonna, Patrizia et al. (2018) Mast cell activation syndrome: Importance of consensus criteria and call for research. J Allergy Clin Immunol 142:1008-1010
Oh, Kyu-Seon; Gottschalk, Rachel A; Lounsbury, Nicolas W et al. (2018) Dual Roles for Ikaros in Regulation of Macrophage Chromatin State and Inflammatory Gene Expression. J Immunol 201:757-771
Ma, Chi A; Xi, Liqiang; Cauff, Brian et al. (2017) Somatic STAT5b gain-of-function mutations in early onset nonclonal eosinophilia, urticaria, dermatitis, and diarrhea. Blood 129:650-653
Schwerd, Tobias; Twigg, Stephen R F; Aschenbrenner, Dominik et al. (2017) A biallelic mutation in IL6ST encoding the GP130 co-receptor causes immunodeficiency and craniosynostosis. J Exp Med 214:2547-2562

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