scholarly journals Screening for mutations in transcription factors in a Czech cohort of 170 patients with congenital and early-onset hypothyroidism: identification of a novel PAX8 mutation in dominantly inherited early-onset non-autoimmune hypothyroidism

2007 ◽  
Vol 156 (5) ◽  
pp. 521-529 ◽  
Author(s):  
Eva Al Taji ◽  
Heike Biebermann ◽  
Zdeňka Límanová ◽  
Olga Hníková ◽  
Jaroslav Zikmund ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Thyroid Gland ◽  
Dna Binding ◽  
Early Onset ◽  
Direct Sequencing ◽  
Low Frequency ◽  
Population Based ◽  
Loss Of Function ◽  
Binding Studies ◽  
Autoimmune Hypothyroidism

Objective: Mutations in NKX2.1, NKX2.5, FOXE1 and PAX8 genes, encoding for transcription factors involved in the development of the thyroid gland, have been identified in a minority of patients with syndromic and non-syndromic congenital hypothyroidism (CH). Design: In a phenotype-selected cohort of 170 Czech paediatric and adolescent patients with non-goitre CH, including thyroid dysgenesis, or non-goitre early-onset hypothyroidism, PAX8, NKX2.1, NKX2.5, FOXE1 and HHEX genes were analysed for mutations. Methods: NKX2.1, NKX2.5, FOXE1 and HHEX genes were directly sequenced in patients with syndromic CH. PAX8 mutational screening was performed in all 170 patients by single-stranded conformation polymorphism, followed by direct sequencing of samples with abnormal findings. The R52P PAX8 mutation was functionally characterized by DNA binding studies. Results: We identified a novel PAX8 mutation R52P, dominantly inherited in a three-generation pedigree and leading to non-congenital, early-onset, non-goitre, non-autoimmune hypothyroidism with gradual postnatal regression of the thyroid size and function. The R52P PAX8 mutation results in the substitution of a highly conserved residue of the DNA-binding domain with a loss-of-function effect. Conclusions: The very low frequency of genetic defects in a population-based cohort of children affected by non-goitre congenital and early-onset hypothyroidism, even in a phenotype-focussed screening study, suggests the pathogenetic role of either non-classic genetic mechanisms or the involvement of genes unknown so far. Identification of a novel PAX8 mutation in a particular variant of non-congenital early-onset hypothyroidism indicates a key function of PAX8 in the postnatal growth and functional maintenance of the thyroid gland.

scholarly journals ATRT-11. PREVALENCE OF GERMLINE VARIANTS IN SMARCB1 INCLUDING SOMATIC MOSAICISM IN AT/RT AND OTHER RHABDOID TUMORS

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii277-iii278
Author(s):  
Ryota Shirai ◽  
Tomoo Osumi ◽  
Keita Terashima ◽  
Chikako Kiyotani ◽  
Meri Uchiyama ◽  
...  
Keyword(s):  
Direct Sequencing ◽  
Snp Array ◽  
Somatic Mosaicism ◽  
Low Frequency ◽  
Rhabdoid Tumor ◽  
Loss Of Function ◽  
Germline Mosaicism ◽  
Germline Variants ◽  
Predisposition Genes ◽  
Rhabdoid Tumors

Abstract BACKGROUND Genetic hallmark of atypical teratoid/rhabdoid tumor (AT/RT) is loss-of-function variants or deletions in SMARCB1 gene on 22q11.2 chromosome, which is common to extracranial malignant rhabdoid tumors (MRT). Previous studies demonstrated that approximately one-thirds of AT/RT and extracranial MRT patients harbored germline SMARCB1 variants as the rhabdoid tumor predisposing syndrome. We studied herein intensive analysis of the SMARCB1 gene in AT/RT and extracranial MRT patients focusing on prevalence of germline genetic variants. PROCEDURE: In total, 16 patients were included. Both tumor-derived DNA and germline DNA were obtained from all patients. First, screening for SMARCB1 alterations in the tumor specimens was done by direct sequencing, ddPCR and SNP array analysis. Then, analysis of germline DNA samples focusing on the genomic abnormalities detected in the paired tumors in each case was performed. RESULTS In eight of 16 cases (50%), genomic alterations observed in the tumor-derived DNA were also detected in the germline DNA. It is worth noting that three patients had germline mosaicism. Two of three patients had mosaic deletion, including SMARCB1 region, and the average copy number of the deleted region in the SMARCB1 gene in the germline was 1.60 and 1.76. For another patient, the fraction of SMARCB1 variants in normal cells was as low as 1.7%. CONCLUSIONS Approximately half the MRT cases in this study had SMARCB1 germline alterations. Considering the presence of low-frequency mosaicisms which conventional methods might overlook, inherited germline variants in predisposition genes are more important than previously assumed for the pathogenesis of pediatric cancers.

scholarly journals Diverse motif ensembles specify non-redundant DNA binding activities of AP-1 family members in macrophages

2018 ◽  
Author(s):  
Gregory J. Fonseca ◽  
Jenhan Tao ◽  
Emma M. Westin ◽  
Sascha H. Duttke ◽  
Nathanael J. Spann ◽  
...  
Keyword(s):  
Machine Learning ◽  
Transcription Factors ◽  
Dna Binding ◽  
Specific Binding ◽  
Family Members ◽  
General Mechanism ◽  
Loss Of Function ◽  
Chip Sequencing ◽  
Collaborative Interactions ◽  
Genomic Locations

ABSTRACTMechanisms by which members of the AP-1 family of transcription factors play both redundant and non-redundant biological roles despite recognizing the same DNA sequence remain poorly understood. To address this question, we investigated the molecular functions and genome-wide DNA binding patterns of AP-1 family members in macrophages. ChIP-sequencing showed overlapping and distinct binding profiles for each factor that were remodeled following TLR4 ligation. Development of a machine learning approach that jointly weighs hundreds of DNA recognition elements yielded dozens of motifs predicted to drive factor-specific binding profiles. Machine learning-based predictions were confirmed by analysis of the effects of mutations in genetically diverse mice and by loss of function experiments. These findings provide evidence that non-redundant genomic locations of different AP-1 family members in macrophages largely result from collaborative interactions with diverse, locus-specific ensembles of transcription factors and suggest a general mechanism for encoding functional specificities of their common recognition motif.

scholarly journals Properties of transcription factors regulating interleukin-2 gene transcription through the NFAT binding site in untreated or drug- treated naive and memory T-helper cells

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2612-2621 ◽  
Author(s):  
A Mouzaki ◽  
D Rungger
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Binding Site ◽  
T Helper Cells ◽  
Interleukin 2 ◽  
Jurkat Cells ◽  
Memory Cells ◽  
T Helper ◽  
Binding Studies ◽  
Helper Cells

Abstract Combining in vitro DNA binding studies and functional transcription assays in the Xenopus oocyte, we have tested the presence and functional state of transcription factors controlling the interleukin-2 (IL-2) promoter through the NFAT binding site. In naive T-helper cells, the IL-2 gene is repressed by a silencer. After first mitogenic stimulation, this silencer becomes undetectable while an activator is newly synthesized. In resting memory cells, the activator has low DNA- binding affinity and is located in the cytoplasm. However, no silencer is formed. Upon renewed cellular activation, this pre-existing activator is again targeted to the nucleus and regains function in promoting transcription. Cyclosporin A and FK506 act on two distinct levels of the IL-2 control mechanism. They prevent nuclear transport and reactivation of the performed activator in memory cells and, in naive cells, they render the silencer resistant to displacement by the activator. DNA-binding of silencer and activator from T-helper, and NFAT-1 from Jurkat cells, requires the same three G residues, but cross- linking analyses show differences in their constituent subunits. Supershift experiments show that the activator contains fra-2 and junD, whereas the silencer reacts with none of the antibodies tested.

Regulation of segmentation and segmental identity by Drosophila homeoproteins: the role of DNA binding in functional activity and specificity

Development ◽  
1997 ◽  
Vol 124 (22) ◽  
pp. 4425-4433 ◽  
Author(s):  
M.D. Biggin ◽  
W. McGinnis
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Target Genes ◽  
Current Model ◽  
Binding Studies ◽  
Binding Model ◽  
Selective Binding ◽  
Recognition Sites

Recent advances have shed new light on how the Q50 homeoproteins act in Drosophila. These transcription factors have remarkably similar and promiscuous DNA-binding specificities in vitro; yet they each specify distinct developmental fates in vivo. One current model suggests that, because the Q50 homeoproteins have distinct biological functions, they must each regulate different target genes. According to this ‘co-selective binding’ model, significant binding of Q50 homeoproteins to functional DNA elements in vivo would be dependent upon cooperative interactions with other transcription factors (cofactors). If the Q50 homeoproteins each interact differently with cofactors, they could be selectively targeted to unique, limited subsets of their in vitro recognition sites and thus control different genes. However, a variety of experiments question this model. Molecular and genetic experiments suggest that the Q50 homeoproteins do not regulate very distinct sets of genes. Instead, they mostly control the expression of a large number of shared targets. The distinct morphogenic properties of the various Q50 homeoproteins may principally result from the different manners in which they either activate or repress these common targets. Further, in vivo binding studies indicate that at least two Q50 homeoproteins have very broad and similar DNA-binding specificities in embryos, a result that is inconsistent with the ‘co-selective binding’ model. Based on these and other data, we suggest that Q50 homeoproteins bind many of their recognition sites without the aid of cofactors. In this ‘widespread binding’ model, cofactors act mainly by helping to distinguish the way in which homeoproteins regulate targets to which they are already bound.

Abstract 14629: Rare Variants for Electrocardiographic Traits Identify Arrhythmia Susceptibility Genes

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Sean J Jurgens ◽  
Seung Hoan Choi ◽  
Christopher M Haggerty ◽  
Amelia W Hall ◽  
Jennifer Halford ◽  
...  
Keyword(s):  
Rare Variants ◽  
Sequence Data ◽  
Low Frequency ◽  
Population Based ◽  
P Wave ◽  
Cardiac Conduction ◽  
Incomplete Penetrance ◽  
Loss Of Function ◽  
Discovery Sample ◽  
Pathogenic Variants

Introduction: Electrocardiogram (ECG) intervals are quantitative and heritable endophenotypes for arrhythmias and sudden cardiac death (SCD). Studying rare sequence variation related to ECG intervals may help identify the genetic underpinnings of cardiac conduction and SCD. Methods: Using a discovery sample of 29,000 individuals with whole-genome sequences from TOPMed and a replication sample of about 100,000 individuals with whole-exome sequence data from the UK Biobank and MyCode, we examined associations between low-frequency (MAF<1%) and rare (MAF<0.1%) coding variants with 5 routinely ascertained ECG intervals (RR, P-wave, PR, QRS, and QTc intervals). We further assessed pathogenic variants in identified genes using ClinVar. Results: In low-frequency single variant analysis, we observed associations for PR interval in PAM ( P =2x10 -7 ) and MFGE8 ( P =5x10 -8 ). In gene-based tests, we identified rare coding variation associated with marked effects in established SCD genes KCNQ1, KCNH2, SCN5A and KCNE1 . For example, loss-of-function or pathogenic variants in KCNQ1 and KCNH2 were carried in 0.2% of individuals, were associated with 29 ms longer QTc intervals ( P =2x10 -82 ) and conferred up to 23-fold increased odds of marked QTc prolongation ( P =4x10 -25 ). Nevertheless, over 75% of carriers had normal QTc intervals. Similarly, loss-of-function or pathogenic variants in SCN5A , carried by 0.1% of individuals, conferred marked PR prolongation (31 ms), yet less than 30% of carriers had first-degree atrioventricular block. Discussion: This study demonstrates the value of studying ECGs in large sequenced biobanks for identifying rare variants predisposing to cardiac arrhythmias. Results define the frequency of pathogenic variation in SCD genes in the population and document incomplete penetrance of such variation. Our findings may serve as a benchmark for future population-based analyses aimed at discovering clinically actionable variants and genes.

scholarly journals Auto-inhibitory regulation of DNA binding by the C-terminal tails of the mitochondrial transcription factors Mtf1 and TFB2M

2020 ◽  
Author(s):  
Urmimala Basu ◽  
Nandini Mishra ◽  
Mohammed Farooqui ◽  
Jiayu Shen ◽  
Laura C. Johnson ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Transcription Initiation ◽  
Binding Activity ◽  
Intramolecular Interactions ◽  
Mitochondrial Rna ◽  
Binding Studies ◽  
Tail Region ◽  
Mitochondrial Transcription ◽  
Dna Binding Activity

ABSTRACTThe structurally homologous Mtf1 and TFB2M proteins serve as transcription initiation factors of the Saccharomyces cerevisiae and human mitochondrial RNA polymerases, respectively. These transcription factors directly interact with the non-template strand of the transcription bubble to drive promoter melting. Given the key roles of Mtf1 and TFB2M in promoter-specific transcription initiation, it is expected that the DNA binding activity of the mitochondrial transcription factors would be regulated to prevent DNA binding at inappropriate times. However, there is little information on how mitochondrial DNA transcription is regulated. While studying the C-tail deletion mutants of Mtf1 and TFB2M, we stumbled upon a new finding that suggested that the flexible C-tail region of these factors autoregulates their DNA binding activity. Quantitative DNA binding studies with fluorescence anisotropy-based titrations show that Mtf1 with an intact C-tail has no affinity for the DNA but the deletion of C-tail greatly increases the DNA binding affinity. Similar observations were made with TFB2M, although autoinhibition by the C-tail of TFB2M was not as absolute as in Mtf1. Analysis of available TFB2M structures show that the C-tail makes intramolecular interactions with the DNA binding groove in the free factor, which we propose masks the DNA binding activity. Further studies show that the RNA polymerase relieves autoinhibition by interacting with the C-tail and engaging it in complex formation. Thus, our biochemical and structural analysis identify previously unknown autoinhibitory and activation mechanisms of mitochondrial transcription factors that regulate the DNA binding activity and aid in specific assembly of the initiation complexes.

scholarly journals The C-terminal tails of the mitochondrial transcription factors Mtf1 and TFB2M are part of an autoinhibitory mechanism that regulates DNA binding

2020 ◽  
Vol 295 (20) ◽  
pp. 6823-6830 ◽  
Author(s):  
Urmimala Basu ◽  
Nandini Mishra ◽  
Mohammed Farooqui ◽  
Jiayu Shen ◽  
Laura C. Johnson ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Transcription Initiation ◽  
Binding Activity ◽  
Intramolecular Interactions ◽  
Mitochondrial Rna ◽  
Binding Studies ◽  
Tail Region ◽  
Mitochondrial Transcription ◽  
Dna Binding Activity

The structurally homologous Mtf1 and TFB2M proteins serve as transcription initiation factors of mitochondrial RNA polymerases in Saccharomyces cerevisiae and humans, respectively. These transcription factors directly interact with the nontemplate strand of the transcription bubble to drive promoter melting. Given the key roles of Mtf1 and TFB2M in promoter-specific transcription initiation, it can be expected that the DNA binding activity of the mitochondrial transcription factors is regulated to prevent DNA binding at inappropriate times. However, little information is available on how mitochondrial DNA transcription is regulated. While studying C-terminal (C-tail) deletion mutants of Mtf1 and TFB2M, we stumbled upon a finding that suggested that the flexible C-tail region of these factors autoregulates their DNA binding activity. Quantitative DNA binding studies with fluorescence anisotropy-based titrations revealed that Mtf1 with an intact C-tail has no affinity for DNA but deletion of the C-tail greatly increases Mtf1's DNA binding affinity. Similar observations were made with TFB2M, although autoinhibition by the C-tail of TFB2M was not as complete as in Mtf1. Analysis of available TFB2M structures disclosed that the C-tail engages in intramolecular interactions with the DNA binding groove in the free factor, which, we propose, inhibits its DNA binding activity. Further experiments showed that RNA polymerase relieves this autoinhibition by interacting with the C-tail and engaging it in complex formation. In conclusion, our biochemical and structural analyses reveal autoinhibitory and activation mechanisms of mitochondrial transcription factors that regulate their DNA binding activities and aid in specific assembly of transcription initiation complexes.

scholarly journals Properties of transcription factors regulating interleukin-2 gene transcription through the NFAT binding site in untreated or drug- treated naive and memory T-helper cells

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2612-2621 ◽  
Author(s):  
A Mouzaki ◽  
D Rungger
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Binding Site ◽  
T Helper Cells ◽  
Interleukin 2 ◽  
Jurkat Cells ◽  
Memory Cells ◽  
T Helper ◽  
Binding Studies ◽  
Helper Cells

Combining in vitro DNA binding studies and functional transcription assays in the Xenopus oocyte, we have tested the presence and functional state of transcription factors controlling the interleukin-2 (IL-2) promoter through the NFAT binding site. In naive T-helper cells, the IL-2 gene is repressed by a silencer. After first mitogenic stimulation, this silencer becomes undetectable while an activator is newly synthesized. In resting memory cells, the activator has low DNA- binding affinity and is located in the cytoplasm. However, no silencer is formed. Upon renewed cellular activation, this pre-existing activator is again targeted to the nucleus and regains function in promoting transcription. Cyclosporin A and FK506 act on two distinct levels of the IL-2 control mechanism. They prevent nuclear transport and reactivation of the performed activator in memory cells and, in naive cells, they render the silencer resistant to displacement by the activator. DNA-binding of silencer and activator from T-helper, and NFAT-1 from Jurkat cells, requires the same three G residues, but cross- linking analyses show differences in their constituent subunits. Supershift experiments show that the activator contains fra-2 and junD, whereas the silencer reacts with none of the antibodies tested.

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