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 found that very small cooling perturbations could lead to massive activation of the enzyme and, in collaboration with Dr Peter Gierschik, Ulm, we constructed a map of the necessary PLCG2 amino acid residues which lead to the striking cold sensitivity. Furthermore, it was established that point mutations in mice who develop an auto inflammatory disease cause similar cold responsiveness, buttressing the idea that page deletions are not needed to cause the cold phenotype. However, we also show that S707Y, the point mutation in APLAID, does not cause cold sensitivity, despite the auto inflammation seen, which is consistent with the patient phenotype. A report describing these findings was published in Cellular Signaling. Finding 2: Because of our discovery of patients with gain-of-function mutations in STAT3 which lead to significant autoimmunity and inflammation, and because of our interest in mir17-92, which is unregulated by STAT3 to suppress a variety of inhibitory pathways in cellular signaling, we collaborated with the Kastner lab, NHGRI, in the investigation of patients with auto inflammation and autoimmunity who have haploinsufficiency in A20, a negative regulator of NFkB signaling which is a target of mir17-92. We showed the cellular inflammatory consequences of these mutations, which in some cases had overlap with the STAT3GOF patients. The results were published in Nature Genetics. Finding 3: In an ongoing collaboration with Jean Laurent Casanova, Rockefeller University, we have been studying patients with gain of function mutations in STAT1. Phenotyping, with special attention to allergy, and cellular investigations are ongoing, however, a large global survey of STAT1GOF patients, including substantial amounts of phenotypic data from the large population followed here, was published in Blood, showing the broad spectrum of disease including infections, autoimmunity, neoplasms, vascular abnormalities, and other manifestations. Finding 4: In studying patients with GATA3 haploinsufficiency, we wished to determine the cellular and phenotypic consequences of haploinsufficiency in such a key Th2-promoting gene highly associated with atopy. We found that allergic sensitization could occur normally, and that there were minimal cellular changes associated with Th2 defects. Thus while the haploinsufficiency has a substantial effect on multiple organ systems including hearing and endocrine, there was little immunologic consequence. These findings have substantial implications for our studies allelic expression of gata3, as well as the impact of gene dose effects in gata3 on allergy risk. 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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