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Gain-of-function STAT1 mutations are associated with PD-L1 overexpression and a defect in B-cell survival

2013; Elsevier BV; Volume: 131; Issue: 6 Linguagem: Inglês

10.1016/j.jaci.2013.01.004

1097-6825

Neil Romberg, Henner Morbach, Monica G. Lawrence, Sang-Hyun Kim, Insoo Kang, Steven M. Holland, Joshua D. Milner, Eric Meffre,

Immune Cell Function and Interaction

Heterozygous gain-of-function mutations in the coiled-coil domain of STAT1 were recently identified as a cause of chronic mucocutaneous candidiasis (CMC).1van de Veerdonk F.L. Plantinga T.S. Hoischen A. Smeekens S.P. Joosten L.A. Gilissen C. et al.STAT1 mutations in autosomal dominant chronic mucocutaneous candidiasis.N Engl J Med. 2011; 365: 54-61Crossref PubMed Scopus (539) Google Scholar, 2Liu L. Okada S. Kong X.F. Kreins A.Y. Cypowyj S. Abhyankar A. et al.Gain-of-function human STAT1 mutations impair IL-17 immunity and underlie chronic mucocutaneous candidiasis.J Exp Med. 2011; 208: 1635-1648Crossref PubMed Scopus (627) Google Scholar As with STAT3 mutations in hyper-IgE syndrome, the candidal susceptibility associated with gain-of-function STAT1 mutations appears secondary to TH17-cell deficiency.3Milner J.D. Brenchley J.M. Laurence A. Freeman A.F. Hill B.J. Elias K.M. et al.Impaired T(H)17 cell differentiation in subjects with autosomal dominant hyper-IgE syndrome.Nature. 2008; 452: 773-776Crossref PubMed Scopus (954) Google Scholar The mechanistic link between constitutive STAT1 activity and diminished TH17 cells is yet to be clearly defined. Here, we present a kindred with a novel gain-of-function STAT1 mutation associated with a complex clinical phenotype including candidiasis, humoral immunodeficiency, overexpression of programmed cell death protein ligand 1 (PD-L1), and increased B-cell apoptosis. We identified 4 related individuals heterozygous for a novel E235A missense mutation in a highly conserved segment of the coiled-coil domain of STAT1 (Fig 1, A). To our knowledge, this is the first mutation described in exon 9 of the STAT1α gene locus to be associated with CMC. The mutation is not present in unaffected adult family members. The index patient (II.6) is a 60-year-old woman with CMC and progressive antibody deficiency. Beginning in infancy, the patient experienced candidal infections at sites including the oral cavity, esophagus, vagina, skin, and nails. Later, as a young woman, the patient was diagnosed with IgG2 subclass deficiency that progressed to frank hypogammaglobulinemia (IgG 514, IgA 7, IgM < 10 mg/dL) and B-cell lymphopenia (2 CD20+cells/μL; normal range, 97–440) requiring intravenous immunoglobulin. Over her lifetime, the patient suffered recurrent pulmonary infections from Pseudomonas aeruginosa, Streptococcus pneumonia, Serratia species, Mycobacterium avium, and respiratory syncytial virus that resulted in severe bronchiectasis requiring lobectomy. Consistent with published reports of patients with gain-of-function STAT1 mutations, our index patient (II.6) has experienced human papillomavirus positive squamous cell carcinoma of palate, basal cell carcinoma, shingles, and fibromuscular dysplasia with carotid and celiac/splenic artery dissection. The index patient's daughter (III.2) is a 30-year-old woman with CMC, B-cell lymphopenia (6 CD20+cells/μL), and IgG2 subclass deficiency manifesting in adolescence. Despite antifungal therapy, the patient experienced candidal infections at sites including the vagina, skin, and nails. She has also experienced noncandidal infections including pneumonia, otitis media, sinusitis, and chronic bronchitis. The third and forth persons carrying the E235A allele are children of patient III.2, 2 males aged 6 weeks and 24 months (IV.1 and IV.2). They have not yet manifested symptoms of immunodeficiency. Stimulation of T cells from patients II.6 and III.2 with IL-21 significantly increased the phosphorylation of STAT1 compared with a healthy control (Fig 1, B). Furthermore, stimulation of patient PBMCs with phorbol 12-myristate 13-acetate/ionomycin demonstrated diminished IL-17–secreting CD4+ cells compared with a related healthy control (Fig 1, C). Hence, the E235A mutation confers to STAT1 a gain of function and is associated with TH17-cell deficiency. A remarkable feature of our kindred is the overexpression of PD-L1 on the surface of naive CD4+ T cells. All 4 family members carrying the E235A allele had higher PD-L1 staining compared with members without it (Fig 1, D). To investigate whether the overexpression of PD-L1 was a common feature to gain-of-function STAT1 mutations, we obtained PBMCs from 2 additional patients carrying either the I156T or the E353K missense STAT1 mutation. Both patients, unrelated to our kindred, revealed a similar increase in PD-L1 expression on their naive T cells (Fig 1, D). Recent data from mice demonstrate that the expression of PD-L1 on undifferentiated naive T cells prevents commitment to the TH17 lineage through a PD-1/PD-L1 interaction. In this context, PD-L1 expression is dependent on IL27/IL27R binding and STAT1.4Hirahara K. Ghoreschi K. Yang X.P. Takahashi H. Laurence A. Vahedi G. et al.Interleukin-27 priming of T cells controls IL-17 production in trans via induction of the ligand PD-L1.Immunity. 2012; 36: 1017-1030Abstract Full Text Full Text PDF PubMed Scopus (206) Google Scholar Accordingly, constitutively active STAT1 molecules in subjects carrying gain-of-function STAT1 mutations may be responsible for increased PD-L1 expression on naive T cells, thereby discouraging differentiation into TH17 cells. As previously described, the clinical phenotype of patients with gain-of-function STAT1 mutations is quite broad and can include candidiasis, antithyroid autoimmunity, squamous cell carcinoma, and vascular anomalies. In this issue, 2 reports show broader phenotypes to gain-of-function STAT1 mutations with an immunodysregulation polyendocrinopathy enteropathy X-linked syndrome (IPEX)-like autoimmune syndrome in one report and disseminated coccidioidomycosis and histoplasmosis in the other. Here, we report a not yet appreciated feature associated with gain-of-function STAT1 mutations: humoral immunodeficiency. STAT1's function as a key modulator of cell death is thoroughly described. Perhaps the best illustration of this is growth arrest of the STAT1-negative U3A fibroblast line upon transfection with wild-type STAT1α and treatment with IFN-γ.5Chin Y.E. Kitagawa M. Su W.C. You Z.H. Iwamoto Y. Fu X.Y. Cell growth arrest and induction of cyclin-dependent kinase inhibitor p21 WAF1/CIP1 mediated by STAT1.Science. 1996; 272: 719-722Crossref PubMed Scopus (743) Google Scholar The IFN-γ receptor requires STAT1 for intracellular signaling. Moreover, transfection of the same cell line with constitutively activated STAT1 initiates caspase-mediated apoptosis.6Sironi J.J. Ouchi T. STAT1-induced apoptosis is mediated by caspases 2, 3, and 7.J Biol Chem. 2004; 279: 4066-4074Crossref PubMed Scopus (100) Google Scholar Related experiments implicate STAT1 activation during apoptosis in B-cell lymphoma cells.7Niitsu N. Higashihara M. Honma Y. Human B-cell lymphoma cell lines are highly sensitive to apoptosis induced by all-trans retinoic acid and interferon-gamma.Leuk Res. 2002; 26: 745-755Abstract Full Text Full Text PDF PubMed Scopus (14) Google Scholar Interestingly, progressive B-cell lymphopenia is a remarkable feature of patients (II.6 and III.2) in our kindred, suggesting a defect in cell survival. Indeed, CD19+ B cells from subjects carrying the E235A allele appear apoptotic with increased Annexin V staining (Fig 2, A, top row) and elevated caspase activity (Fig 2, B). In culture for 24 hours, B cells from patient III.2 demonstrated even greater Annexin V and considerable 7-AAD staining, evidence of accelerated cell death (Fig 2, A, middle row). The patient's B cells were only partially rescued by the stimulation of their B-cell receptors (Fig 2, A, bottom row). We also found enhanced caspase activity in B cells from the 2 additional patients who were heterozygous for either the E353K or the I156T missense STAT1 mutation (Fig 2, B). B-cell lymphopenia was a significant finding in the former patient (39 cells/μL) but not the latter (335 cells/μL). Altogether our data reveal that gain-of-function STAT1 mutations increase B-cell apoptosis. Over time, this may result in B-cell lymphopenia and antibody deficiency. In summary, we identified individuals heterozygous for gain-of-function STAT1 mutation with 2 unappreciated features. The first is the overexpression of PD-L1 on naive T cells, which provides a general mechanism for how constitutively active STAT1 blocks the development of the TH17 lineage. The second feature, accelerated B-cell apoptosis that may result in progressive B-cell lymphopenia and humoral immunodeficiency, further broadens the clinical phenotype associated with gain-of-function STAT1 mutations. We acknowledge Dr O'Shea of the National Institutes of Health for sharing his unpublished manuscript with our group and thank the patients and their family members for making this work possible. CorrectionJournal of Allergy and Clinical ImmunologyVol. 132Issue 6PreviewWith regard to the June 2013 article entitled "Gain-of-function STAT1 mutations are associated with PD-L1 overexpression and a defect in B-cell survival" (J Allergy and Clin Immunol 2013;131:1691-3), the new gain-of-function STAT1 mutation that the authors describe at nucleotide position 705 that is displayed in Fig 1, A results in a E235G amino acid substitution, not a E235A substitution as was reported in the original publication. The authors regret the error. Full-Text PDF