Pathological actions of STAT4, MTOR and STAT3 in human T cell differentiation

Abstract

There is an expanding catalogue of immune deficiency diseases with inflammatory and autoimmune manifestations. Understanding the genetic basis of this paradoxical association could enhance our understanding of the pathophysiology of not only rare immune deficiency syndromes, but also more common inflammatory or autoimmune diseases. We aimed to elucidate mechanism of such diseases using two approaches. First, a discovery project in which we identified a proband who presented with both inflammation and recurrent infections, including bronchiectasis by the age of 4 years, which was associated with hypergammaglobulinaemia. Whole exome sequencing revealed novel mutations in STAT4 and MTOR. Until now, there has been no human genetic analysis supported by functional studies to determine the role of STAT4 or MTOR in Th1 effector differentiation. The STAT4 mutation (affecting the DNA binding domain) results in prolonged STAT4 phosphorylation and nuclear retention, thus conferring a gain-of-function phenotype. We show that this promotes excessive Th1 and follicular helper T cell (TFH) formation. The MTOR mutation affects the negative regulatory domain, and is therefore also gain-of-function. We found biochemical evidence for cross-regulation between STAT4 and mTOR that explained accentuated TFH and Th1 formation. In this case, enhanced TFH formation might help explain the aberrant antibody response observed in the proband. The second approach examined T cell differentiation in patients with known defects in STAT3. Loss-of-function (LoF) mutations in STAT3 result in autosomal dominant hyper IgE syndrome (ADHIES), and these patients present with unexplained atopic manifestations. Mouse studies have revealed that cytokine signalling via STAT3 promotes IL10 production by Th2 cells that have regulatory role. We found that suboptimal STAT3 signalling in humans also compromises formation of inducible IL10 production by human Th2 cells, which might explain this accentuated atopic phenotype in ADHIES. More recently, patients with STAT3 gain-of-function (GoF) mutations have been reported in which antibody deficiency occurs concurrently with organ-specific autoimmunity. We observed that suboptimal STAT3 signalling promotes CD4+ T cell exhaustion marked by PD1 and CD57 expression. Since CD57+ PD1high cells constitute a significant subset of human TFH cells, we proceeded to characterize this population in detail. Interestingly, we show that this subset contains CD4+ T cells with cytotoxic gene expression signature and activity. Interestingly, cytotoxicity is attenuated in CD57+ TFH cells when compared with their circulating counterparts. STAT3 LoF results in expansion of circulating CD57+ cells, but does not increase the cytotoxic fraction above the proportion normally observed in tonsil. By contrast, STAT3 GoF enhances the cytotoxic fraction in CD57+ CD4+ T cells in blood, with cytotoxic action against autologous B cells. Since STAT3 is known to promote CXCR5 expression and abundance of TFH cells, concurrent promotion of cytotoxicity within TFH cells might provide an additional mechanism for defects in antibody production within germinal centres of patients with enhanced STAT3 activity. These findings point to events in germinal centres that can explain the paradoxical association between antibody deficiency and autoimmunity. Taken together, these approaches provide insight into new mechanisms to explain concurrent immune deficiency and autoimmunity. Show more