Missense variants in human forkhead transcription factors reveal determinants of forkhead DNA bispecificity
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PROVIDER: PRJNA1266190 | ENA |
REPOSITORIES: ENA
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Project description:Recognition of specific DNA sequences by transcription factors (TFs) is a key step in transcriptional control of gene expression. While most forkhead (FH) TFs bind either an FKH (RYAAAYA) or an FHL (GACGC) recognition motif, some FHs can bind both motifs. Mechanisms that control whether a FH is monospecific versus bispecific have remained unknown. Screening a library of 12 reference FH proteins, 61 naturally occurring missense variants including clinical variants, and 22 designed mutant FHs for DNA binding activity using universal (“all 10-mer”) protein binding microarrays (PBMs) revealed non-DNA-contacting residues that control mono- versus bispecificity. Variation in non-DNA-contacting amino acid residues of TFs is associated with human traits and may play a role in the evolution of TF DNA binding activities and gene regulatory networks.
2025-05-27 | GSE297784 | GEO
Project description:1 Novel Missense Variants in Patients with Primary Immunodeficiency and Immune dysregulation
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Project description:Nuclear deubiquitinase BAP1 (BRCA1-Associated Protein 1) is a core component of multiprotein complexes that promote transcription by reversing the ubiquitination of histone 2A (H2A). BAP1 is a tumor suppressor gene whose germline loss-of-function variants predispose to cancer. To our knowledge, there are very rare examples of different germline variants in the same gene causing either a NDD or a tumor predisposition syndrome. Here, we report a series of 11 de novo germline heterozygous missense BAP1 variants associated with a rare syndromic neurodevelopmental disorder (NDD). Functional analysis showed that most of the variants cannot rescue the consequences of BAP1 inactivation, suggesting a loss-of-function mechanism. In T cells isolated from two affected children, H2A deubiquitination was impaired in matching peripheral blood mononuclear cells, histone H3 K27 acetylation ChIP-seq indicated that these BAP1 variants induced genome-wide chromatin state alterations, with enrichment for regulatory regions surrounding genes of the ubiquitin-proteasome system (UPS). Altogether, these results define a clinical syndrome caused by rare germline missense BAP1 variants that alter chromatin remodeling through abnormal histone ubiquitination and lead to transcriptional dysregulation of developmental genes.
2021-12-10 | GSE190394 | GEO
Project description:BRCA1 (breast cancer 1, early onset) mutations confer a high breast and ovarian cancer risk. Most of BRCA1 cancer-predisposing mutations originate truncated proteins, but missense mutations have also been detected in familial breast and ovarian cancer patients. These variants are rare and their role in cancer predisposition is often difficult to ascertain. Our purpose in the present work was to study the molecular mechanisms affected in human cells by two BRCA1 missense variants both located in the second BRCA1 BRCT domain, M1775R and A1789T. These variants were isolated from familial breast cancer patients and their role in the pathogenesis of breast cancer was also investigated by a study previously performed by our group in yeast cells. Here we present a microarray study to compare the expression profiles of HeLa cells transfected with these two variants and HeLa cells transfected with BRCA1 wild type. Data analysis was performed by evaluating three comparisons: M1775R versus wild-type (M1775RvsWT-contrast), A1789T versus wild-type (A1789TvsWT-contrast) and the mutated BRCT domain versus wild-type (MutvsWT-contrast), obtained by considering the two variants as a unique BRCT mutation. We found 173 differentially expressed genes in MutvsWT-contrast, 201 in M1775RvsWT-contrast and 313 in A1789TvsWT-contrast. Our results showed that the considered BRCA1 variants had an impact on cell processes often deregulated in cancerogenesis, such as cell cycle progression and DNA damage response and repair. Thus, our study further supports the putative role in the pathogenesis of cancer of the M1775R and A1789T BRCA1 variants from a molecular point of view.
2012-06-01 | E-MTAB-761 | biostudies-arrayexpress
Project description:Rare germline heterozygous missense variants of the BRCA1-Associated Protein 1 gene, BAP1, heterozygous missense variants cause a syndromic neurodevelopmental disorder
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Project description:Polycomb repressive complex 1 (PRC1) modifies chromatin through catalysis of histone 2A lysine 119 monoubiquitination (H2AK119ub1). RING1 and RNF2 interchangeably serve as the catalytic subunit within PRC1. Pathogenic missense variants in PRC1 core components reveal functions of these proteins that are obscured in knockout models. While Ring1a knockout models remain healthy, the microcephaly and neuropsychiatric phenotypes associated with a pathogenic RING1 missense variant implicate unappreciated functions. Using an in-vitro model of neurodevelopment, we observe that RING1 contributes to the broad placement of H2AK119ub1, and that its targets overlap with those of RNF2. PRC1 complexes harboring hypomorphic RING1 bind target loci but do not catalyze H2AK119ub1, reducing H2AK119ub1 by preventing catalytically active complexes from accessing the locus. This results in delayed DNA damage repair and cell cycle progression in neural progenitor cells (NPCs). Conversely, reduced H2AK119ub1 due to hypomorphic RING1 does not generate differential expression that impacts NPC differentiation. The greater reduction of H2AK119ub1 dosage observed in NPCs expressing hypomorphic RNF2, however, exhibited defects in both DNA repair and transcriptional regulation. These findings suggest that the DNA damage response is more sensitive to H2AK119ub1 dosage change than is regulation of gene expression.
2024-07-29 | GSE252485 | GEO
Project description:We performed a massively parallel screen in human HAP1 cells to identify loss-of-function missense variants in the key DNA mismatch repair factor MSH2. Resulting variant loss-of-function (LOF) scores are strongly concordant with previous functional evidence and available variant classification.
2020-12-23 | GSE162130 | GEO
Project description:Polycomb repressive complex 1 (PRC1) modifies chromatin through catalysis of histone 2A lysine 119 monoubiquitination (H2AK119ub1). RING1 and RNF2 interchangeably serve as the catalytic subunit within PRC1. Pathogenic missense variants in PRC1 core components reveal functions of these proteins that are obscured in knockout models. While Ring1a knockout models remain healthy, the microcephaly and neuropsychiatric phenotypes associated with a pathogenic RING1 missense variant implicate unappreciated functions. Using an in-vitro model of neurodevelopment, we observe that RING1 contributes to the broad placement of H2AK119ub1, and that its targets overlap with those of RNF2. PRC1 complexes harboring hypomorphic RING1 bind target loci but do not catalyze H2AK119ub1, reducing H2AK119ub1 by preventing catalytically active complexes from accessing the locus. This results in delayed DNA damage repair and cell cycle progression in neural progenitor cells (NPCs). Conversely, reduced H2AK119ub1 due to hypomorphic RING1 does not generate differential expression that impacts NPC differentiation. The greater reduction of H2AK119ub1 dosage observed in NPCs expressing hypomorphic RNF2, however, exhibited defects in both DNA repair and transcriptional regulation. These findings suggest that the DNA damage response is more sensitive to H2AK119ub1 dosage change than is regulation of gene expression.
2024-07-29 | GSE252449 | GEO
Project description:Polycomb repressive complex 1 (PRC1) modifies chromatin through catalysis of histone 2A lysine 119 monoubiquitination (H2AK119ub1). RING1 and RNF2 interchangeably serve as the catalytic subunit within PRC1. Pathogenic missense variants in PRC1 core components reveal functions of these proteins that are obscured in knockout models. While Ring1a knockout models remain healthy, the microcephaly and neuropsychiatric phenotypes associated with a pathogenic RING1 missense variant implicate unappreciated functions. Using an in-vitro model of neurodevelopment, we observe that RING1 contributes to the broad placement of H2AK119ub1, and that its targets overlap with those of RNF2. PRC1 complexes harboring hypomorphic RING1 bind target loci but do not catalyze H2AK119ub1, reducing H2AK119ub1 by preventing catalytically active complexes from accessing the locus. This results in delayed DNA damage repair and cell cycle progression in neural progenitor cells (NPCs). Conversely, reduced H2AK119ub1 due to hypomorphic RING1 does not generate differential expression that impacts NPC differentiation. The greater reduction of H2AK119ub1 dosage observed in NPCs expressing hypomorphic RNF2, however, exhibited defects in both DNA repair and transcriptional regulation. These findings suggest that the DNA damage response is more sensitive to H2AK119ub1 dosage change than is regulation of gene expression.
2024-07-29 | GSE252447 | GEO
Project description:The forkhead transcription factor (TF) FoxN3 recognizes both the canonical forkhead DNA motif (RYAAAYA), as well as a novel alternate motif (GACGC). These two motifs are more distinct than the typically observed primary and secondary motifs of other TFs, as the two motifs cannot be aligned and are not clearly related to each other. Amino acids at the canonical base-contacting positions in the forkhead DNA binding domain are largely conserved throughout the forkhead family, so the ability of FoxN3 and other bispecific forkhead factors to recognize the alternate site cannot be explained by amino acid substitutions at these positions. Neither the mechanism of this bispecific DNA recognition nor its regulatory function are understood. Here we show that human FoxN3 does recognize both motifs in cells. We also report the crystal structures of the FoxN3 DNA binding domain bound to the canonical forkhead binding site, and separately to the alternate DNA binding site. FoxN3 adopts a remarkably similar conformation to recognize both motifs, making contacts with different DNA bases using the same amino acids. However, the DNA structure is significantly altered between the two motifs, allowing the same domain to recognize two motifs of different lengths. Changes between the two structures in the conformation of segments of the forkhead domain outside of the direct DNA-contacting positions contribute to the ability to bind the alternate site. Swapping these segments of the domain between different forkhead proteins changes the ability of proteins to recognize the alternate site, highlighting the importance of non-base-contacting positions in determining TF-DNA binding specificity. The structures present a new mechanism by which a single DBD can recognize two divergent sequences.
2019-02-27 | GSE112672 | GEO