scholarly journals Co-SELECT reveals sequence non-specific contribution of DNA shape to transcription factor binding in vitro

2019 ◽  
Vol 47 (13) ◽  
pp. 6632-6641 ◽  
Author(s):  
Soumitra Pal ◽  
Jan Hoinka ◽  
Teresa M Przytycka
Keyword(s):  
Dna Binding ◽  
Specific Binding ◽  
Sequence Similarity ◽  
Binding Motif ◽  
Sequence Motif ◽  
Sequence Motifs ◽  
Shape Features ◽  
Dna Shape

Abstract Understanding the principles of DNA binding by transcription factors (TFs) is of primary importance for studying gene regulation. Recently, several lines of evidence suggested that both DNA sequence and shape contribute to TF binding. However, the following compelling question is yet to be considered: in the absence of any sequence similarity to the binding motif, can DNA shape still increase binding probability? To address this challenge, we developed Co-SELECT, a computational approach to analyze the results of in vitro HT-SELEX experiments for TF–DNA binding. Specifically, Co-SELECT leverages the presence of motif-free sequences in late HT-SELEX rounds and their enrichment in weak binders allows Co-SELECT to detect an evidence for the role of DNA shape features in TF binding. Our approach revealed that, even in the absence of the sequence motif, TFs have propensity to bind to DNA molecules of the shape consistent with the motif specific binding. This provides the first direct evidence that shape features that accompany the preferred sequence motifs also bestow an advantage for weak, sequence non-specific binding.

scholarly journals Co-SELECT reveals sequence non-specific contribution of DNA shape to transcription factor binding in vitro

2018 ◽  
Author(s):  
Soumitra Pal ◽  
Jan Hoinka ◽  
Teresa M. Przytycka
Keyword(s):  
Dna Binding ◽  
Sequence Similarity ◽  
Binding Motif ◽  
Sequence Motif ◽  
Shape Features ◽  
Specific Shape ◽  
Dna Shape ◽  
Specific Contribution

AbstractUnderstanding the principles of DNA binding by transcription factors (TFs) is of primary importance for studying gene regulation. Recently, several lines of evidence suggested that both DNA sequence and shape contribute to TF binding. However, the question if in the absence of any sequence similarity to the binding motif, DNA shape can still increase probability of binding was yet to be addressed.To address this challenge, we developed Co-SELECT, a computational approach to analyze the results of in vitro HT-SELEX experiments for TF-DNA binding. Specifically, the presence of motif-free sequences in late HT-SELEX rounds and their enrichment in weak binders allowed us to detect evidence for the role of DNA shape features in TF binding.Our approach revealed that, even in the absence of the sequence motif, TFs have propensity to weakly bind to DNA molecules enriched in specific shape features. Surprisingly, we also found that some properties of DNA shape contribute to promiscuous binding of all tested TF families. Strikingly, such promiscuously bound shapes correspond to the most frequent shape formed by the DNA. We propose that this promiscuous binding facilitates diffusing of TFs along the DNA molecule before it is locked in its binding site.

scholarly journals Quantitative modeling of transcription factor binding specificities using DNA shape

2015 ◽  
Vol 112 (15) ◽  
pp. 4654-4659 ◽  
Author(s):  
Tianyin Zhou ◽  
Ning Shen ◽  
Lin Yang ◽  
Namiko Abe ◽  
John Horton ◽  
...  
Keyword(s):  
Dna Binding ◽  
In Vitro Selection ◽  
Specific Binding ◽  
Support Vector ◽  
Shape Features ◽  
Basic Step ◽  
Shape Information ◽  
Dna Shape

DNA binding specificities of transcription factors (TFs) are a key component of gene regulatory processes. Underlying mechanisms that explain the highly specific binding of TFs to their genomic target sites are poorly understood. A better understanding of TF−DNA binding requires the ability to quantitatively model TF binding to accessible DNA as its basic step, before additional in vivo components can be considered. Traditionally, these models were built based on nucleotide sequence. Here, we integrated 3D DNA shape information derived with a high-throughput approach into the modeling of TF binding specificities. Using support vector regression, we trained quantitative models of TF binding specificity based on protein binding microarray (PBM) data for 68 mammalian TFs. The evaluation of our models included cross-validation on specific PBM array designs, testing across different PBM array designs, and using PBM-trained models to predict relative binding affinities derived from in vitro selection combined with deep sequencing (SELEX-seq). Our results showed that shape-augmented models compared favorably to sequence-based models. Although both k-mer and DNA shape features can encode interdependencies between nucleotide positions of the binding site, using DNA shape features reduced the dimensionality of the feature space. In addition, analyzing the feature weights of DNA shape-augmented models uncovered TF family-specific structural readout mechanisms that were not revealed by the DNA sequence. As such, this work combines knowledge from structural biology and genomics, and suggests a new path toward understanding TF binding and genome function.

Involvement of the consensus sequence motif at coil 2b in the assembly and stability of vimentin filaments

1992 ◽  
Vol 102 (1) ◽  
pp. 31-41 ◽  
Author(s):  
P.D. Kouklis ◽  
P. Traub ◽  
S.D. Georgatos
Keyword(s):  
Consensus Sequence ◽  
Sequence Motif ◽  
Sequence Motifs ◽  
3T3 Cells ◽  
Filament Assembly ◽  
In Vitro Assembly ◽  
Vimentin Filaments

Nearly all intermediate filament (IF) proteins share two sequence motifs located at the N- and the C-terminal ends of their helical rod domain (‘coil 1a’ and ‘coil 2b’, respectively). To examine the structural role of the coil 2b motif, we have performed in vitro assembly studies and in vivo microinjection experiments employing two site-specific reagents: (a) a 20-residue synthetic peptide (C-2) representing the conserved motif itself and (b) a monoclonal antibody (anti-IFA) that recognises an epitope within the conserved coil 2b sequence. We demonstrate here that vimentin protofilaments, when induced to assemble in the presence of C-2 or anti-IFA, show a lower propensity to polymerise and yield various abberant structures. The few filaments that are formed under these conditions appear much shorter than normal IFs and are unravelled or aggregated. Furthermore, when preformed vimentin filaments are exposed to C-2 or anti-IFA, most of the normal IFs are converted into shorter filamentous forms that possess an abberant morphology. None of these effects is seen when vimentin subunits are coincubated with control peptides. Microinjection of anti-IFA into the cytoplasm of interphasic 3T3 cells provokes collapse of vimentin IFs into a juxtanuclear mass and formation of numerous amorphous aggregates distributed throughout the cytoplasm. These two effects are not seen when the anti-IFA is microinjected into the cell nucleus. Our results provide experimental evidence supporting previous suggestions for a role for the conserved coil 2b sequence in filament assembly. We propose that this region is interacting with other sites along the vimentin molecule and that these interactions are essential for proper protofilament-protofilament alignment and filament stability.

scholarly journals Cooperative Activation of Human Papillomavirus Type 8 Gene Expression by the E2 Protein and the Cellular Coactivator p300

2002 ◽  
Vol 76 (21) ◽  
pp. 11042-11053 ◽  
Author(s):  
Andreas Müller ◽  
Andreas Ritzkowsky ◽  
Gertrud Steger
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Specific Binding ◽  
Regulatory Region ◽  
Keratinocyte Differentiation ◽  
E2 Protein ◽  
Protein Protein Interaction ◽  
Cooperative Activation

ABSTRACT The E2 proteins of papillomaviruses (PV) bind to the coactivator CBP/p300 as do many other transcription factors, but the precise role of CBP/p300 in E2-specific functions is not yet understood. We show that the E2 protein of human PV type 8 (HPV8) directly binds to p300. Activation of HPV8 gene expression by low amounts of HPV8 E2 was stimulated up to sevenfold by coexpression of p300. The interaction between E2 and p300 may play a role in differentiation-dependent activation of PV gene expression, since we can show that the expression level of p300 increases during keratinocyte differentiation. Surprisingly, sequence-specific binding of E2 to its recognition sites within the regulatory region of HPV8 is not necessary for this cooperation, indicating that E2 can be recruited to the promoter via protein-protein interaction. HPV8 E2 binds via its N-terminal activation domain (AD), its C-terminal DNA binding domain (DBD), and its internal hinge region to p300 in vitro. Transient-transfection assays revealed that the AD is necessary and sufficient for cooperative activation with p300. However, we provide evidence that the interaction of the hinge and the DBD of HPV8 E2 with p300 may contribute. Our data suggest an important role of p300 in regulation of HPV8 gene expression and reveal a new mechanism by which E2 may be recruited to a promoter to activate transcription without sequence specific DNA binding.

scholarly journals Transcription factors recognize DNA shape without nucleotide recognition

2017 ◽  
Author(s):  
Md. Abul Hassan Samee ◽  
Benoit G. Bruneau ◽  
Katherine S. Pollard
Keyword(s):  
Transcription Factors ◽  
Binding Sites ◽  
De Novo ◽  
Sequence Information ◽  
Sequence Motif ◽  
Sequence Motifs ◽  
Shape Features ◽  
Sequence Recognition ◽  
Helical Twist ◽  
Dna Shape

AbstractWe hypothesized that transcription factors (TFs) recognize DNA shape without nucleotide sequence recognition. Motivating an independent role for shape, many TF binding sites lack a sequence-motif, DNA shape adds specificity to sequence-motifs, and different sequences can encode similar shapes. We therefore asked if binding sites of a TF are enriched for specific patterns of DNA shape-features, e.g., helical twist. We developed ShapeMF, which discovers these shape-motifs de novo without taking sequence information into account. We find that most TFs assayed in ENCODE have shape-motifs and bind regulatory regions recognizing shape-motifs in the absence of sequence-motifs. When shape- and sequence-recognition co-occur, the two types of motifs can be overlapping, flanking, or separated by consistent spacing. Shape-motifs are prevalent in regions co-bound by multiple TFs. Finally, TFs with identical sequence motifs have different shape-motifs, explaining their binding at distinct locations. These results establish shape-motifs as drivers of TF-DNA recognition complementary to sequence-motifs.

Integrated requirement of non-specific and sequence-specific DNA binding in MYC-driven transcription

2020 ◽  
Author(s):  
Paola Pellanda ◽  
Mattia Dalsass ◽  
Marco Filipuzzi ◽  
Alessia Loffreda ◽  
Alessandro Verrecchia ◽  
...  
Keyword(s):  
Dna Binding ◽  
Transcriptional Activation ◽  
Specific Binding ◽  
Regulatory Elements ◽  
Chromatin Interaction ◽  
Binding Motif ◽  
Sequence Motifs ◽  
Binding Modes ◽  
Transcriptional Responses ◽  
Sequence Recognition

AbstractEukaryotic transcription factors recognize specific DNA sequence motifs, but are also endowed with generic, non-specific DNA-binding activity: how these binding modes are integrated to determine select transcriptional outputs remains unresolved. We designed mutants of the MYC transcription factor bearing substitutions in residues that contact either the DNA backbone or specific bases within the consensus binding motif (E-box), and profiled their DNA-binding and gene-regulatory activities in murine cells. Our data reveal that non-specific DNA binding is required for MYC to engage onto active regulatory elements in the genome, preceding sequence recognition; beyond merely stabilizing MYC onto select target loci, sequence-specific binding contributes to its precise positioning and – most unexpectedly – to transcriptional activation per se. In particular, at any given binding intensity, promoters targeted via the cognate DNA motif were more frequently activated by MYC. Hence, seemingly promiscuous chromatin interaction profiles actually encompass diverse DNA-binding modalities, driving defined, sequence-dependent transcriptional responses.

scholarly journals DNA sequence+shape kernel enables alignment-free modeling of transcription factor binding

2016 ◽  
Author(s):  
Wenxiu Ma ◽  
Lin Yang ◽  
Remo Rohs ◽  
William Stafford Noble
Keyword(s):  
Dna Binding ◽  
Binding Affinity ◽  
Dna Sequence ◽  
Supplementary Information ◽  
Sequence Motifs ◽  
Genomic Context ◽  
Shape Information ◽  
Dna Shape ◽  
Better Than

AbstractMotivationTranscription factors (TFs) bind to specific DNA sequence motifs. Several lines of evidence suggest that TF-DNA binding is mediated in part by properties of the local DNA shape: the width of the minor groove, the relative orientations of adjacent base pairs, etc. Several methods have been developed to jointly account for DNA sequence and shape properties in predicting TF binding affinity. However, a limitation of these methods is that they typically require a training set of aligned TF binding sites.ResultsWe describe a sequence+shape kernel that leverages DNA sequence and shape information to better understand protein-DNA binding preference and affinity. This kernel extends an existing class of k-mer based sequence kernels, based on the recently described di-mismatch kernel. Using three in vitro benchmark datasets, derived from universal protein binding microarrays (uPBMs), genomic context PBMs (gcPBMs) and SELEX-seq data, we demonstrate that incorporating DNA shape information improves our ability to predict protein-DNA binding affinity. In particular, we observe that (1) the k-spectrum+shape model performs better than the classical k-spectrum kernel, particularly for small k values; (2) the di-mismatch kernel performs better than the k-mer kernel, for larger k; and (3) the di-mismatch+shape kernel performs better than the di-mismatch kernel for intermediate k values.AvailabilityThe software is available at https://bitbucket.org/wenxiu/[email protected], [email protected] informationSupplementary data are available at Bioinformatics online.

On the Role of Transferrin in 67Ga Uptake by Tumor Cells

1988 ◽  
Vol 27 (04) ◽  
pp. 151-153
Author(s):  
P. Thouvenot ◽  
F. Brunotte ◽  
J. Robert ◽  
L. J. Anghileri
Keyword(s):  
Specific Binding ◽  
Subcellular Fractionation ◽  
Animal Blood ◽  
Tumor Bearing ◽  
Cell Structures ◽  
The Difference ◽  
Sarcoma Cells ◽  
In Vitro Uptake

In vitro uptake of 67Ga-citrate and 59Fe-citrate by DS sarcoma cells in the presence of tumor-bearing animal blood plasma showed a dramatic inhibition of both 67Ga and 59Fe uptakes: about ii/io of 67Ga and 1/5o of the 59Fe are taken up by the cells. Subcellular fractionation appears to indicate no specific binding to cell structures, and the difference of binding seems to be related to the transferrin chelation and transmembrane transport differences

scholarly journals Splicing factor SRSF1 promotes breast cancer progression via oncogenic splice switching of PTPMT1

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Jun-Xian Du ◽  
Yi-Hong Luo ◽  
Si-Jia Zhang ◽  
Biao Wang ◽  
Cong Chen ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Progression ◽  
High Risk Group ◽  
Clinical Samples ◽  
Binding Motif ◽  
Ki 67 ◽  
Tissue Samples

Abstract Background Intensive evidence has highlighted the effect of aberrant alternative splicing (AS) events on cancer progression when triggered by dysregulation of the SR protein family. Nonetheless, the underlying mechanism in breast cancer (BRCA) remains elusive. Here we sought to explore the molecular function of SRSF1 and identify the key AS events regulated by SRSF1 in BRCA. Methods We conducted a comprehensive analysis of the expression and clinical correlation of SRSF1 in BRCA based on the TCGA dataset, Metabric database and clinical tissue samples. Functional analysis of SRSF1 in BRCA was conducted in vitro and in vivo. SRSF1-mediated AS events and their binding motifs were identified by RNA-seq, RNA immunoprecipitation-PCR (RIP-PCR) and in vivo crosslinking followed by immunoprecipitation (CLIP), which was further validated by the minigene reporter assay. PTPMT1 exon 3 (E3) AS was identified to partially mediate the oncogenic role of SRSF1 by the P-AKT/C-MYC axis. Finally, the expression and clinical significance of these AS events were validated in clinical samples and using the TCGA database. Results SRSF1 expression was consistently upregulated in BRCA samples, positively associated with tumor grade and the Ki-67 index, and correlated with poor prognosis in a hormone receptor-positive (HR+) cohort, which facilitated proliferation, cell migration and inhibited apoptosis in vitro and in vivo. We identified SRSF1-mediated AS events and discovered the SRSF1 binding motif in the regulation of splice switching of PTPMT1. Furthermore, PTPMT1 splice switching was regulated by SRSF1 by binding directly to its motif in E3 which partially mediated the oncogenic role of SRSF1 by the AKT/C-MYC axis. Additionally, PTPMT1 splice switching was validated in tissue samples of BRCA patients and using the TCGA database. The high-risk group, identified by AS of PTPMT1 and expression of SRSF1, possessed poorer prognosis in the stage I/II TCGA BRCA cohort. Conclusions SRSF1 exerts oncogenic roles in BRCA partially by regulating the AS of PTPMT1, which could be a therapeutic target candidate in BRCA and a prognostic factor in HR+ BRCA patient.

scholarly journals Genome-Wide Binding Analyses of HOXB1 Revealed a Novel DNA Binding Motif Associated with Gene Repression

2021 ◽  
Vol 9 (1) ◽  
pp. 6
Author(s):  
Narendra Pratap Singh ◽  
Bony De Kumar ◽  
Ariel Paulson ◽  
Mark E. Parrish ◽  
Carrie Scott ◽  
...  
Keyword(s):  
Dna Binding ◽  
Target Genes ◽  
Embryonic Stem ◽  
Binding Motif ◽  
Binding Assays ◽  
Histone Marks ◽  
Genome Wide ◽  
Hox Cofactors

Knowledge of the diverse DNA binding specificities of transcription factors is important for understanding their specific regulatory functions in animal development and evolution. We have examined the genome-wide binding properties of the mouse HOXB1 protein in embryonic stem cells differentiated into neural fates. Unexpectedly, only a small number of HOXB1 bound regions (7%) correlate with binding of the known HOX cofactors PBX and MEIS. In contrast, 22% of the HOXB1 binding peaks display co-occupancy with the transcriptional repressor REST. Analyses revealed that co-binding of HOXB1 with PBX correlates with active histone marks and high levels of expression, while co-occupancy with REST correlates with repressive histone marks and repression of the target genes. Analysis of HOXB1 bound regions uncovered enrichment of a novel 15 base pair HOXB1 binding motif HB1RE (HOXB1 response element). In vitro template binding assays showed that HOXB1, PBX1, and MEIS can bind to this motif. In vivo, this motif is sufficient for direct expression of a reporter gene and over-expression of HOXB1 selectively represses this activity. Our analyses suggest that HOXB1 has evolved an association with REST in gene regulation and the novel HB1RE motif contributes to HOXB1 function in part through a repressive role in gene expression.

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