Molecular mechanism and structural basis of gain-of-function of STAT1 caused by pathogenic R274Q mutation
Ryoji Fujiki1, Atsushi Hijikata2, Tsuyoshi Shirai2
1From the Department of Technology Development, Kazusa DNA Research Institute, 2-6-7 Kazusa-Kamatari, Kisarazu-Shi, Chiba-Ken, 292-0818, fujiki@kazusa.or.jp.
Abstract:
Gain-of-function (GOF) mutations in the STAT1 gene are critical for the onset of chronic mucocutaneous candidiasis (CMC) disease. However, the molecular basis for the gain of STAT1 function remains largely unclear. Here, we investigated the structural features of STAT1 GOF residues to better understand the impact of these pathogenic mutations. We constructed STAT1 alanine mutants of the α3 helix residues of the coiled-coil domain, which are frequently found in CMC pathogenic mutations, and measured their transcriptional activities. Most of the identified GOF residues were located inside the coiled-coil domain stem structure or at the protein surface of the anti-parallel dimer interface. Unlike those, Arg-274 was adjacent to the DNA-binding domain. In addition, Arg-274 was found to functionally interact with Gln-441 in the DNA-binding domain. Because Gln-441 is located at the anti-parallel dimer contact site, Gln-441 reorientation by Arg-274 mutation probably impedes formation of the dimer. Further, the statistical analysis of RNA-seq data with STAT1-deficient epithelial cells and primary T cells from a CMC patient revealed that the R274Q mutation affected gene expression levels of 66 and 76 non-overlapping RefSeq genes, respectively. Because their transcription levels were only slightly modulated by wild-type STAT1, we concluded that the R274Q mutation increased transcriptional activity but did not change dramatically the repertoire of STAT1 targets. Hence, we provide a novel mechanism of STAT1 GOF triggered by a CMC pathogenic mutation.
Insights
Gain-of-function mutations in Signal Transducer and Activator of Transcription 1 (STAT1) cause chronic mucocutaneous candidiasis. A novel mechanism reveals how the R274Q mutation enhances STAT1 activity by disrupting dimer formation.
Area of Science:
- Immunology
- Genetics
- Molecular Biology
Background:
- Gain-of-function (GOF) mutations in Signal Transducer and Activator of Transcription 1 (STAT1) are implicated in chronic mucocutaneous candidiasis (CMC).
- The precise molecular mechanisms underlying STAT1 GOF in CMC remain incompletely understood.
- Structural insights into pathogenic STAT1 mutations are crucial for elucidating disease pathogenesis.
Purpose of the Study:
- To investigate the structural and functional impact of STAT1 GOF residues, particularly those in the coiled-coil domain, on STAT1 function.
- To elucidate the molecular basis of STAT1 gain-of-function in chronic mucocutaneous candidiasis.
- To identify novel mechanisms driving STAT1 GOF mutations.
Main Methods:
- Construction and transcriptional activity measurement of STAT1 alanine mutants at α3 helix residues.
- Structural analysis of STAT1 GOF residues within the coiled-coil domain and at dimer interfaces.
- RNA-sequencing analysis of STAT1-deficient epithelial cells and primary T cells from CMC patients with the R274Q mutation.
Main Results:
- Identified STAT1 GOF residues located within the coiled-coil domain or at the anti-parallel dimer interface.
- The Arg-274 mutation was found adjacent to the DNA-binding domain and functionally interacted with Gln-441, potentially impeding dimer formation.
- RNA-seq analysis showed the R274Q mutation altered expression of specific RefSeq genes, indicating increased transcriptional activity without a broad change in STAT1 targets.
Conclusions:
- The Arg-274 mutation in STAT1 provides a novel mechanism for gain-of-function by disrupting dimer formation and modulating transcriptional activity.
- This study enhances understanding of the molecular basis of STAT1 GOF in chronic mucocutaneous candidiasis.
- The findings highlight the importance of structural integrity at the dimer interface for normal STAT1 function.
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