Dominant negative effects of Weaver syndrome-associated EZH2 variants [RNA-seq]
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PROVIDER: PRJNA1291488 | ENA |
REPOSITORIES: ENA
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Project description:Dominant negative effects of Weaver syndrome-associated EZH2 variants
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Project description:Heterozygous missense mutations in EZH2 cause Weaver syndrome (WS), a developmental disorder characterized by intellectual disability and overgrowth. EZH2encodes the enzymatic subunit of Polycomb Repressive Complex 2 (PRC2), which mediates mono-, di-, and tri-methylation of histone H3 lysine 27 (H3K27me1/2/3). Most WS-associated EZH2 variants lack functional characterization but are presumed loss of function. However, the lack of early truncating mutations in EZH2 led us to hypothesise a dominant-negative mechanism for WS, which was supported by our structural analysis of all known WS-associated EZH2 variants. We isogenically modelled 10 representative variants in embryonic stem cells and showed they reduce global H3K27me2/3 with concomitant increases in H3K27ac and chromatin decompaction. Notably, the pattern of H3K27me2/3 reductions indicated dominant-negative interference on PRC2 activity, even when WS-variants were expressed atlow levels. RNA-seq identified weakly Polycomb-bound genes that lose canonical PRC1 (cPRC1) occupancy and become derepressed, including several phenotypically relevant growth control genes. Comparative analysis of a gain-of-function EZH2 variant causing growth restriction revealed reciprocal chromatin and transcriptional changes compared to WS-associated variants. Taken together, our findings support a model where EZH2 variants associated with opposing developmental growth syndromes affect not only H3K27me3, but also intergenic H3K27me2, chromatin architecture, and cPRC1 recruitment.
2025-09-03 | GSE302575 | GEO
Project description:Heterozygous missense mutations in EZH2 cause Weaver syndrome (WS), a developmental disorder characterized by intellectual disability and overgrowth. EZH2 encodes the enzymatic subunit of Polycomb Repressive Complex 2 (PRC2), which mediates mono-, di-, and tri-methylation of histone H3 lysine 27 (H3K27me1/2/3). Most WS-associated EZH2 variants lack functional characterization but are presumed loss of function. However, the lack of early truncating mutations in EZH2 led us to hypothesise a dominant-negative mechanism for WS, which was supported by our structural analysis of all known WS-associated EZH2 variants. We isogenically modelled 10 representative variants in embryonic stem cells and showed they reduce global H3K27me2/3 with concomitant increases in H3K27ac and chromatin decompaction. Notably, the pattern of H3K27me2/3 reductions indicated dominant- negative interference on PRC2 activity, even when WS-variants were expressed at low levels. RNA-seq identified weakly Polycomb-bound genes that lose canonical PRC1 (cPRC1) occupancy and become derepressed, including several phenotypically relevant growth control genes. Comparative analysis of a gain-of-function EZH2 variant causing growth restriction revealed reciprocal chromatin and transcriptional changes compared to WS-associated variants. Taken together, our findings support a model where EZH2 variants associated with opposing developmental growth syndromes affect not only H3K27me3, but also intergenic H3K27me2, chromatin architecture, and cPRC1 recruitment.
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Project description:Expression profiles from Rag1 mutant and Rag1/Ezh2 double mutant double negative thymocytes Keywords = Rag1 Keywords = Ezh2 Keywords = Thymocyte Keywords: repeat sample
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Project description:Heterozygous in-frame mutations in coding regions of human STAT3 underlie the only known autosomal dominant form of hy- per IgE syndrome (AD HIES). About 5% of familial cases remain unexplained. The mutant proteins are loss-of-function and dominant-negative when tested following overproduction in re- cipient cells. However, the production of mutant proteins has not been detected and quantified in the cells of heterozygous pa- tients. We report a deep intronic heterozygous STAT3 mutation, c.1282-89C>T, in 7 relatives with AD HIES. This mutation creates a new exon in the STAT3 complementary DNA, which, when over- expressed, generates a mutant STAT3 protein (D427ins17) that is loss-of-function and dominant-negative in terms of tyrosine phos- phorylation, DNA binding, and transcriptional activity. In immor- talized B cells from these patients, the D427ins17 protein was 2 kDa larger and 4-fold less abundant than wild-type STAT3, on mass spectrometry. The patients’ primary B and T lymphocytes responded poorly to STAT3-dependent cytokines. These findings are reminiscent of the impaired responses of leukocytes from other patients with AD HIES due to typical STAT3 coding muta- tions, providing further evidence for the dominance of the mutant intronic allele. These findings highlight the importance of sequenc- ing STAT3 introns in patients with HIES without candidate variants in coding regions and essential splice sites. They also show that AD HIES-causing STAT3 mutant alleles can be dominant-negative even if the encoded protein is produced in significantly smaller amounts than wild-type STAT3.
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