The Nuclear Pore Complex Mediates Binding of the Mig1 Repressor to Target Promoters

All eukaryotic cells alter their transcriptional program in response to the sugar glucose. In Saccharomyces cerevisiae, the best-studied downstream effector of this response is the glucose-regulated repressor Mig1. We show here that nuclear pore complexes also contribute to glucose-regulated gene expression. NPCs participate in glucose-responsive repression by physically interacting with Mig1 and mediating its function independently of nucleocytoplasmic transport. Surprisingly, despite its abundant presence in the nucleus of glucose-grown nup120Δ or nup133Δ cells, Mig1 has lost its ability to interact with target promoters. The glucose repression defect in the absence of these nuclear pore components therefore appears to result from the failure of Mig1 to access its consensus recognition sites in genomic DNA. We propose that the NPC contributes to both repression and activation at the level of transcription

The Transcription Factor Gcr1 Stimulates Cell Growth by Participating in Nutrient-Responsive Gene Expression on a Global Level

Transcriptomic reprogramming is critical to the coordination between growth and cell cycle progression in response to changing extracellular conditions. In Saccharomyces cerevisiae, the transcription factor Gcr1 contributes to this coordination by supporting maximum expression of G1 cyclins in addition to regulating both glucose-induced and glucose-repressed genes. We report here the comprehensive genome-wide expression profiling of gcr1Δ cells. Our data show that reduced expression of ribosomal protein genes in gcr1Δ cells is detectable both 20 min after glucose addition and in steady-state cultures of raffinose-grown cells, showing that this defect is not the result of slow growth or growth on a repressing sugar. However, the large cell phenotype of the gcr1Δ mutant occurs only in the presence of repressing sugars. GCR1 deletion also results in aberrant derepression of numerous glucose repressed loci; glucose-grown gcr1Δ cells actively respire, demonstrating that this global alteration in transcription corresponds to significant changes at the physiological level. These data offer an insight into the coordination of growth and cell division by providing an integrated view of the transcriptomic, phenotypic, and metabolic consequences of GCR1 deletion

The Nuclear Pore Complex Mediates Binding of the Mig1 Repressor to Target Promoters

All eukaryotic cells alter their transcriptional program in response to the sugar glucose. In Saccharomyces cerevisiae, the best-studied downstream effector of this response is the glucose-regulated repressor Mig1. We show here that nuclear pore complexes also contribute to glucose-regulated gene expression. NPCs participate in glucose-responsive repression by physically interacting with Mig1 and mediating its function independently of nucleocytoplasmic transport. Surprisingly, despite its abundant presence in the nucleus of glucose-grown nup120Δ or nup133Δ cells, Mig1 has lost its ability to interact with target promoters. The glucose repression defect in the absence of these nuclear pore components therefore appears to result from the failure of Mig1 to access its consensus recognition sites in genomic DNA. We propose that the NPC contributes to both repression and activation at the level of transcription

Coiled Coil Structures and Transcription: An Analysis of the \u3ci\u3eS. cerevisiae\u3c/i\u3e coilome

The α-helical coiled coil is a simple but widespread motif that is an integral feature of many cellular structures. Coiled coils allow monomeric building blocks to form complex assemblages that can serve as molecular motors and springs. Previous parametrically delimited analyses of the distribution of coiled coils in the genomes of diverse organisms, including Escherichia coli, Saccharomyces cerevisiae, Arabidopsis thaliana, Caenorhabditis elegans and Homo sapiens, have identified conserved biological processes that make use of this versatile motif. Here we present a comprehensive inventory of the set of coiled coil proteins in S. cerevisiae by combining multiple coiled coil prediction algorithms with extensive literature curation. Our analysis of this set of proteins, which we call the coilome, reveals a wider role for this motif in transcription than was anticipated, particularly with respect to the category that includes nucleocytoplasmic shuttling factors involved in transcriptional regulation. We also show that the constitutively nuclear yeast transcription factor Gcr1 is homologous to the mammalian transcription factor MLL3, and that two coiled coil domains conserved between these homologs are important for Gcr1 dimerization and function. These data support the hypothesis that coiled coils are required to assemble structures essential for proper functioning of the transcriptional machinery

Glucose-Responsive Regulators of Gene Expression in Saccharomyces cerevisiae Function at the Nuclear Periphery via a Reverse Recruitment Mechanism

Regulation of gene transcription is a key feature of developmental, homeostatic, and oncogenic processes. The reverse recruitment model of transcriptional control postulates that eukaryotic genes become active by moving to contact transcription factories at nuclear substructures; our previous work showed that at least some of these factories are tethered to nuclear pores. We demonstrate here that the nuclear periphery is the site of key events in the regulation of glucose-repressed genes, which together compose one-sixth of the Saccharomyces cerevisiae genome. We also show that the canonical glucose-repressed gene SUC2 associates tightly with the nuclear periphery when transcriptionally active but is highly mobile when repressed. Strikingly, SUC2 is both derepressed and confined to the nuclear rim in mutant cells where the Mig1 repressor is nuclear but not perinuclear. Upon derepression all three subunits (α, β, and γ) of the positively acting Snf1 kinase complex localize to the nuclear periphery, resulting in phosphorylation of Mig1 and its export to the cytoplasm. Reverse recruitment therefore appears to explain a fundamental pathway of eukaryotic gene regulation

Glucose-Responsive Regulators of Gene Expression in \u3ci\u3eSaccharomyces cerevisiae\u3c/i\u3e Function at the Nuclear Periphery via a Reverse Recruitment Mechanism

Regulation of gene transcription is a key feature of developmental, homeostatic, and oncogenic processes. The reverse recruitment model of transcriptional control postulates that eukaryotic genes become active by moving to contact transcription factories at nuclear substructures; our previous work showed that at least some of these factories are tethered to nuclear pores. We demonstrate here that the nuclear periphery is the site of key events in the regulation of glucose-repressed genes, which together compose one-sixth of the Saccharomyces cerevisiae genome. We also show that the canonical glucose-repressed gene SUC2 associates tightly with the nuclear periphery when transcriptionally active but is highly mobile when repressed. Strikingly, SUC2 is both derepressed and confined to the nuclear rim in mutant cells where the Mig1 repressor is nuclear but not perinuclear. Upon derepression all three subunits (α, β, and γ) of the positively acting Snf1 kinase complex localize to the nuclear periphery, resulting in phosphorylation of Mig1 and its export to the cytoplasm. Reverse recruitment therefore appears to explain a fundamental pathway of eukaryotic gene regulation

Widespread transcription at neuronal activity-regulated enhancers

We used genome-wide sequencing methods to study stimulus-dependent enhancer function in mouse cortical neurons. We identified ,12,000 neuronal activity-regulated enhancers that are bound by the general transcriptional co-activator CBP in an activity-dependent manner. A function of CBP at enhancers may be to recruit RNA polymerase II (RNAPII), as we also observed activity-regulated RNAPII binding to thousands of enhancers. Notably, RNAPII at enhancers transcribes bi-directionally a novel class of enhancer RNAs (eRNAs) within enhancer domains defined by the presence of histone H3 monomethylated at lysine 4. The level of eRNA expression at neuronal enhancers positively correlates with the level of messenger RNA synthesis at nearby genes, suggesting that eRNA synthesis occurs specifically at enhancers that are actively engaged in promoting mRNA synthesis. These findings reveal that a widespread mechanism of enhancer activation involves RNAPII binding and eRNA synthesis

Diagnostic utility and reporting recommendations for clinical DNA methylation episignature testing in genetically undiagnosed rare diseases

Purpose: This study aims to assess the diagnostic utility and provide reporting recommendations for clinical DNA methylation episignature testing based on the cohort of patients tested through the EpiSign Clinical Testing Network. Methods: The EpiSign assay utilized unsupervised clustering techniques and a support vector machine–based classification algorithm to compare each patient's genome-wide DNA methylation profile with the EpiSign Knowledge Database, yielding the result that was reported. An international working group, representing distinct EpiSign Clinical Testing Network health jurisdictions, collaborated to establish recommendations for interpretation and reporting of episignature testing. Results: Among 2399 cases analyzed, 1667 cases underwent a comprehensive screen of validated episignatures, imprinting, and promoter regions, resulting in 18.7% (312/1667) positive reports. The remaining 732 referrals underwent targeted episignature analysis for assessment of sequence or copy-number variants (CNVs) of uncertain significance or for assessment of clinical diagnoses without confirmed molecular findings, and 32.4% (237/732) were positive. Cases with detailed clinical information were highlighted to describe various utility scenarios for episignature testing. Conclusion: Clinical DNA methylation testing including episignatures, imprinting, and promoter analysis provided by an integrated network of clinical laboratories enables test standardization and demonstrates significant diagnostic yield and clinical utility beyond DNA sequence analysis in rare diseases.</p

Diagnostic utility and reporting recommendations for clinical DNA methylation episignature testing in genetically undiagnosed rare diseases

Purpose: This study aims to assess the diagnostic utility and provide reporting recommendations for clinical DNA methylation episignature testing based on the cohort of patients tested through the EpiSign Clinical Testing Network. Methods: The EpiSign assay utilized unsupervised clustering techniques and a support vector machine–based classification algorithm to compare each patient's genome-wide DNA methylation profile with the EpiSign Knowledge Database, yielding the result that was reported. An international working group, representing distinct EpiSign Clinical Testing Network health jurisdictions, collaborated to establish recommendations for interpretation and reporting of episignature testing. Results: Among 2399 cases analyzed, 1667 cases underwent a comprehensive screen of validated episignatures, imprinting, and promoter regions, resulting in 18.7% (312/1667) positive reports. The remaining 732 referrals underwent targeted episignature analysis for assessment of sequence or copy-number variants (CNVs) of uncertain significance or for assessment of clinical diagnoses without confirmed molecular findings, and 32.4% (237/732) were positive. Cases with detailed clinical information were highlighted to describe various utility scenarios for episignature testing. Conclusion: Clinical DNA methylation testing including episignatures, imprinting, and promoter analysis provided by an integrated network of clinical laboratories enables test standardization and demonstrates significant diagnostic yield and clinical utility beyond DNA sequence analysis in rare diseases.</p

Diagnostic utility and reporting recommendations for clinical DNA methylation episignature testing in genetically undiagnosed rare diseases

Purpose: This study aims to assess the diagnostic utility and provide reporting recommendations for clinical DNA methylation episignature testing based on the cohort of patients tested through the EpiSign Clinical Testing Network. Methods: The EpiSign assay utilized unsupervised clustering techniques and a support vector machine–based classification algorithm to compare each patient's genome-wide DNA methylation profile with the EpiSign Knowledge Database, yielding the result that was reported. An international working group, representing distinct EpiSign Clinical Testing Network health jurisdictions, collaborated to establish recommendations for interpretation and reporting of episignature testing. Results: Among 2399 cases analyzed, 1667 cases underwent a comprehensive screen of validated episignatures, imprinting, and promoter regions, resulting in 18.7% (312/1667) positive reports. The remaining 732 referrals underwent targeted episignature analysis for assessment of sequence or copy-number variants (CNVs) of uncertain significance or for assessment of clinical diagnoses without confirmed molecular findings, and 32.4% (237/732) were positive. Cases with detailed clinical information were highlighted to describe various utility scenarios for episignature testing. Conclusion: Clinical DNA methylation testing including episignatures, imprinting, and promoter analysis provided by an integrated network of clinical laboratories enables test standardization and demonstrates significant diagnostic yield and clinical utility beyond DNA sequence analysis in rare diseases.</p