Transcriptional Profiling of STAT1 Gain-of-Function Reveals Common and Mutation-Specific Fingerprints

Abstract

STAT1 gain-of-function (GOF) is a primary immunodeficiency typically characterized by chronic mucocutaneous candidiasis (CMC), recurrent respiratory infections, and autoimmunity. Less commonly, also immunodysregulation polyendocrinopathy enteropathy X-linked (IPEX)-like syndromes with CMC, and combined immunodeficiency without CMC have been described. Recently, our group and others have shown that different mutation-specific mechanisms underlie STAT1 GOF in vitro, including faster nuclear accumulation (R274W), and reduced mobility (R321, N574I) to near immobility in the nucleus (T419R) upon IFNγ stimulation. In this work, we evaluated the transcriptomic fingerprint of the aforementioned STAT1 GOF mutants (R274W, R321S, T419R, and N574I) relative to STAT1 wild-type upon IFNγ stimulation in an otherwise isogenic cell model. The majority of genes up-regulated in wild-type STAT1 cells were significantly more up-regulated in cells expressing GOF mutants, except for T419R. In addition to the common interferon regulated genes (IRG), STAT1 GOF mutants up-regulated an additional set of genes, that were in part shared with other GOF mutants or mutation-specific. Overall, R274W and R321S transcriptomes clustered with STAT1 WT, while T419R and N574I had a more distinct fingerprint. We observed reduced frequency of canonical IFNγ activation site (GAS) sequences in promoters of genes up-regulated by all the STAT1 GOF mutants, suggesting loss of DNA binding specificity for the canonical GAS consensus. Interestingly, the T419R mutation, expected to directly increase the affinity for DNA, showed the most pronounced effects on the transcriptome. T419R STAT1 dysregulated more non-IRG than the other GOF mutants and fewer GAS or degenerate GAS promotor sequences could be found in the promoter regions of these genes. In conclusion, our work confirms hyperactivation of common sets of IFNγ-induced genes in STAT1 GOF with additional dysregulation of mutation-specific genes, in line with the earlier observed mutation-specific mechanisms. Binding to more degenerate GAS sequences is proposed as a mechanism toward transcriptional dysregulation in R274W, R321S, and N574I. For T419R, an increased interaction with the DNA is suggested to result in a broader and less GAS-specific response. Our work indicates that multiple routes leading to STAT1 GOF are associated with common and private transcriptomic fingerprints, which may contribute to the phenotypic variation observed in vivo.

Case report: Myocarditis in congenital STAT1 gain-of function

“…We report a case of a 55-year-old man with a de novo congenital STAT1 gain-of-function mutation (c963A>T, p.R321S), as previously reported and validated ( Figure 1A ) ( 1 , 6 – 8 ). The patient presented with chronic mucocutaneous candidiasis (CMC) ( Figure 1B ), complicated by esophageal stenosis, respiratory infections, and bronchiectasis.…”

Application of Bioinformatics Tools for the Prediction of Helper MicroRNAs for Improvement of Oncolytic Virus Efficacy

“…It is a transcription mediator of IFNs. STAT1 is activated by JAKs and is an important inhibitory protein in the reovirus replication cycle [21,34,35].…”

The Goldilocks Principle of JAKs and STATs: Gain-of function mutations, loss-of-function mutations, and their clinical consequences

“…There also exists the concept that DNA binding speci city decreases with certain STAT1-GOF mutations as fewer GAS motifs are detectable in promoters of STAT1-GOF-speci c genes. The ratio of GAS present to GAS absent in promoters of STAT1-regulated genes decreased signi cantly for the STAT1-T419R variant in ChiP-Seq experiments [102]. In RNA-Seq analyses, dysregulated gene transcription caused by the STAT1 variants resulted in different mutation-speci c ngerprints and offers a possible explanation for the wide range of phenotypic variation.…”

“…These ndings are complemented by the observation of high STAT1 mRNA levels [95,99,100]. Other studies suggest premature nuclear import with normal phosphorylation/dephosphorylation rate [101], increased nuclear accumulation (R247W), decreased mobility (R321, N571I), or immobility in the nucleus (T419R) as the cause of STAT1 hyperactivation [102]. There also exists the concept that DNA binding speci city decreases with certain STAT1-GOF mutations as fewer GAS motifs are detectable in promoters of STAT1-GOF-speci c genes.…”