TEAD transcription factors regulate gene expression through interactions with DNA and other proteins. They are crucial for the development of eukaryotic organisms and to control the expression of genes involved mostly in cell proliferation and differentiation; however, their deregulation can lead to tumorigenesis. To study the interactions of TEAD1 with M-CAT motifs and their inverted versions, the KD of each complex was determined, and H/D exchange, quantitative chemical cross-linking, molecular docking, and smFRET were utilized for structural characterization. ChIP-qPCR was employed to correlate the results with a cell line model. The results obtained showed that although the inverted motif has 10× higher KD, the same residues were affected by the presence of M-CAT in both orientations. Molecular docking and smFRET revealed that TEAD1 binds the inverted motif rotated 180°. In addition, the inverted motif was proven to be occupied by TEAD1 in Jurkat cells, suggesting that the low-affinity binding sites present in the human genome may possess biological relevance.

• MeSH
• DNA vazebné proteiny chemie   metabolismus   MeSH
• DNA chemie   metabolismus   MeSH
• jaderné proteiny chemie   metabolismus   MeSH
• Jurkat buňky MeSH
• lidé MeSH
• nukleotidové motivy MeSH
• simulace molekulového dockingu MeSH
• transkripční faktory TEA domény MeSH
• transkripční faktory chemie   metabolismus   MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

[Figure: see text].

• MeSH
• adaptorové proteiny signální transdukční chemie   metabolismus   MeSH
• DNA vazebné proteiny chemie   metabolismus   MeSH
• elongace genetické transkripce * MeSH
• exprese genu MeSH
• interakční proteinové domény a motivy genetika   MeSH
• lidé MeSH
• mapy interakcí proteinů MeSH
• molekulární modely MeSH
• mutace MeSH
• nádorové buněčné linie MeSH
• proteinové domény MeSH
• proteiny vázající RNA chemie   genetika   metabolismus   MeSH
• RNA-polymerasa II chemie   metabolismus   MeSH
• transkripční elongační faktory chemie   metabolismus   MeSH
• transkripční faktory chemie   genetika   metabolismus   MeSH
• vazba proteinů MeSH
• vnitřně neuspořádané proteiny chemie   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

The At-Hook Motif Nuclear Localized Protein (AHL) gene family encodes embryophyte-specific nuclear proteins with DNA binding activity. They modulate gene expression and affect various developmental processes in plants. We identify AHL18 (At3G60870) as a developmental modulator of root system architecture and growth. AHL18 is involved in regulation of the length of the proliferation domain and number of dividing cells in the root apical meristem and thereby, cell production. Both primary root growth and lateral root development respond according to AHL18 transcription level. The ahl18 knock-out plants show reduced root systems due to a shorter primary root and a lower number of lateral roots. This change results from a higher number of arrested and non-developing lateral root primordia (LRP) rather than from a decreased LRP initiation. The over-expression of AHL18 results in a more extensive root system, longer primary roots, and increased density of lateral root initiation events. AHL18 is thus involved in the formation of lateral roots at both LRP initiation and their later development. We conclude that AHL18 participates in modulation of root system architecture through regulation of root apical meristem activity, lateral root initiation and emergence; these correspond well with expression pattern of AHL18.

• MeSH
• Arabidopsis genetika   růst a vývoj   metabolismus   MeSH
• AT-hook motivy MeSH
• DNA vazebné proteiny chemie   genetika   metabolismus   MeSH
• kořeny rostlin genetika   růst a vývoj   metabolismus   MeSH
• mutace MeSH
• proteiny huseníčku chemie   genetika   metabolismus   MeSH
• regulace genové exprese u rostlin MeSH
• vývojová regulace genové exprese MeSH
• Publikační typ
• časopisecké články MeSH

Cruciform structures are preferential targets for many architectural and regulatory proteins, as well as a number of DNA binding proteins with weak sequence specificity. Some of these proteins are also capable of inducing the formation of cruciform structures upon DNA binding. In this paper we analyzed the amino acid composition of eighteen cruciform binding proteins of Homo sapiens. Comparison with general amino acid frequencies in all human proteins revealed unique differences, with notable enrichment for lysine and serine and/or depletion for alanine, glycine, glutamine, arginine, tyrosine and tryptophan residues. Based on bootstrap resampling and fuzzy cluster analysis, multiple molecular mechanisms of interaction with cruciform DNA structures could be suggested, including those involved in DNA repair, transcription and chromatin regulation. The proteins DEK, HMGB1 and TOP1 in particular formed a very distinctive group. Nonetheless, a strong interaction network connecting nearly all the cruciform binding proteins studied was demonstrated. Data reported here will be very useful for future prediction of new cruciform binding proteins or even construction of predictive tool/web-based application.

• MeSH
• aminokyseliny chemie   MeSH
• chromatin MeSH
• chromozomální proteiny, nehistonové chemie   MeSH
• DNA vazebné proteiny chemie   MeSH
• DNA-topoisomerasy I chemie   MeSH
• konformace nukleové kyseliny MeSH
• lidé MeSH
• onkogenní proteiny chemie   MeSH
• protein HMGB1 chemie   MeSH
• proteiny vázající poly-ADP-ribosu chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

The limited information available on the structure of complexes involving transcription factors and cognate DNA response elements represents a major obstacle in the quest to understand their mechanism of action at the molecular level. We implemented a concerted structural proteomics approach, which combined hydrogen-deuterium exchange (HDX), quantitative protein-protein and protein-nucleic acid cross-linking (XL), and homology analysis, to model the structure of the complex between the full-length DNA binding domain (DBD) of Forkhead box protein O4 (FOXO4) and its DNA binding element (DBE). The results confirmed that FOXO4-DBD assumes the characteristic forkhead topology shared by these types of transcription factors, but its binding mode differs significantly from those of other members of the family. The results showed that the binding interaction stabilized regions that were rather flexible and disordered in the unbound form. Surprisingly, the conformational effects were not limited only to the interface between bound components, but extended also to distal regions that may be essential to recruiting additional factors to the transcription machinery. In addition to providing valuable new insights into the binding mechanism, this project provided an excellent evaluation of the merits of structural proteomics approaches in the investigation of systems that are not directly amenable to traditional high-resolution techniques.

• MeSH
• DNA vazebné proteiny chemie   metabolismus   MeSH
• DNA chemie   metabolismus   MeSH
• hmotnostní spektrometrie MeSH
• molekulární struktura MeSH
• responzivní elementy MeSH
• transkripční faktory chemie   metabolismus   MeSH
• vodík-deuteriová výměna MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

Topoisomerase II (TOP2) relieves torsional stress by forming transient cleavage complex intermediates (TOP2ccs) that contain TOP2-linked DNA breaks (DSBs). While TOP2ccs are normally reversible, they can be "trapped" by chemotherapeutic drugs such as etoposide and subsequently converted into irreversible TOP2-linked DSBs. Here, we have quantified etoposide-induced trapping of TOP2ccs, their conversion into irreversible TOP2-linked DSBs, and their processing during DNA repair genome-wide, as a function of time. We find that while TOP2 chromatin localization and trapping is independent of transcription, it requires pre-existing binding of cohesin to DNA. In contrast, the conversion of trapped TOP2ccs to irreversible DSBs during DNA repair is accelerated 2-fold at transcribed loci relative to non-transcribed loci. This conversion is dependent on proteasomal degradation and TDP2 phosphodiesterase activity. Quantitative modeling shows that only two features of pre-existing chromatin structure-namely, cohesin binding and transcriptional activity-can be used to predict the kinetics of TOP2-induced DSBs.

• MeSH
• chromozomy genetika   MeSH
• DNA vazebné proteiny chemie   genetika   MeSH
• DNA-topoisomerasy typu II chemie   genetika   MeSH
• DNA chemie   genetika   MeSH
• dvouřetězcové zlomy DNA * MeSH
• etoposid chemie   MeSH
• genetická transkripce MeSH
• genová konverze genetika   MeSH
• HCT116 buňky MeSH
• inhibitory topoisomerasy II chemie   farmakologie   MeSH
• kinetika MeSH
• lidé MeSH
• multiproteinové komplexy chemie   genetika   MeSH
• oprava DNA genetika   MeSH
• proteiny vázající poly-ADP-ribosu chemie   genetika   MeSH
• torze mechanická MeSH
• translokace genetická genetika   MeSH
• zlomy chromozomů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH

Human telomeric repeat binding factors TRF1 and TRF2 along with TIN2 form the core of the shelterin complex that protects chromosome ends against unwanted end-joining and DNA repair. We applied a single-molecule approach to assess TRF1-TIN2-TRF2 complex formation in solution at physiological conditions. Fluorescence cross-correlation spectroscopy was used to describe the complex assembly by analyzing how coincident fluctuations of differently labeled TRF1 and TRF2 correlate when they move together through the confocal volume of the microscope. We observed, at the single-molecule level, that TRF1 effectively substitutes TRF2 on TIN2. We assessed also the effect of another telomeric factor TPP1 that recruits telomerase to telomeres. We found that TPP1 upon binding to TIN2 induces changes that expand TIN2 binding capacity, such that TIN2 can accommodate both TRF1 and TRF2 simultaneously. We suggest a molecular model that explains why TPP1 is essential for the stable formation of TRF1-TIN2-TRF2 core complex.

• MeSH
• DNA vazebné proteiny chemie   metabolismus   MeSH
• lidé MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• protein TRF1 genetika   metabolismus   MeSH
• protein TRF2 genetika   metabolismus   MeSH
• proteinové domény MeSH
• proteiny vázající telomery genetika   metabolismus   MeSH
• telomerasa metabolismus   MeSH
• telomery metabolismus   MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Nucleoid-associated proteins belong to a group of small but abundant proteins in bacterial cells. These transcription regulators are responsible for many important cellular processes and also are involved in pathogenesis of bacteria. The best-known nucleoid-associated proteins, such as HU, FIS, H-NS, and IHF, are often discussed. The most important findings in research concerning HU protein are described in this mini review. Its roles in DNA compaction, shape modulation, and negative supercoiling induction have been studied intensively. HU protein regulates bacteria survival, growth, SOS response, virulence genes expression, cell division, and many other cell processes. Elucidating the mechanism of HU protein action has been the subject of many research projects. This mini review provides a comprehensive overview of the HU protein.

• MeSH
• bakteriální proteiny chemie   genetika   metabolismus   MeSH
• DNA bakterií metabolismus   MeSH
• DNA vazebné proteiny chemie   genetika   metabolismus   MeSH
• DNA chemie   MeSH
• posttranslační úpravy proteinů MeSH
• regulace genové exprese u bakterií fyziologie   MeSH
• vazba proteinů MeSH
• virulence genetika   fyziologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

NExo is an enzyme from Neisseria meningitidis that is specialized in the removal of the 3'-phosphate and other 3'-lesions, which are potential blocks for DNA repair. NExo is a highly active DNA 3'-phosphatase, and although it is from the class II AP family it lacks AP endonuclease activity. In contrast, the NExo homologue NApe, lacks 3'-phosphatase activity but is an efficient AP endonuclease. These enzymes act together to protect the meningococcus from DNA damage arising mainly from oxidative stress and spontaneous base loss. In this work, we present crystal structures of the specialized 3'-phosphatase NExo bound to DNA in the presence and absence of a 3'-phosphate lesion. We have outlined the reaction mechanism of NExo, and using point mutations we bring mechanistic insights into the specificity of the 3'-phosphatase activity of NExo. Our data provide further insight into the molecular origins of plasticity in substrate recognition for this class of enzymes. From this we hypothesize that these specialized enzymes lead to enhanced efficiency and accuracy of DNA repair and that this is important for the biological niche occupied by this bacterium.

• MeSH
• bakteriální proteiny chemie   MeSH
• DNA vazebné proteiny chemie   MeSH
• DNA-lyasa (apurinová nebo apyrimidinová) chemie   MeSH
• DNA chemie   MeSH
• endonukleasy metabolismus   MeSH
• exodeoxyribonukleasy chemie   MeSH
• katalytická doména MeSH
• konformace proteinů MeSH
• krystalografie rentgenová MeSH
• mutace MeSH
• mutageneze cílená MeSH
• Neisseria meningitidis enzymologie   genetika   MeSH
• oprava DNA * MeSH
• oxidační stres MeSH
• poškození DNA MeSH
• substrátová specifita MeSH
• vazba proteinů MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: De novo mutations in PURA have recently been described to cause PURA syndrome, a neurodevelopmental disorder characterised by severe intellectual disability (ID), epilepsy, feeding difficulties and neonatal hypotonia. OBJECTIVES: To delineate the clinical spectrum of PURA syndrome and study genotype-phenotype correlations. METHODS: Diagnostic or research-based exome or Sanger sequencing was performed in individuals with ID. We systematically collected clinical and mutation data on newly ascertained PURA syndrome individuals, evaluated data of previously reported individuals and performed a computational analysis of photographs. We classified mutations based on predicted effect using 3D in silico models of crystal structures of Drosophila-derived Pur-alpha homologues. Finally, we explored genotype-phenotype correlations by analysis of both recurrent mutations as well as mutation classes. RESULTS: We report mutations in PURA (purine-rich element binding protein A) in 32 individuals, the largest cohort described so far. Evaluation of clinical data, including 22 previously published cases, revealed that all have moderate to severe ID and neonatal-onset symptoms, including hypotonia (96%), respiratory problems (57%), feeding difficulties (77%), exaggerated startle response (44%), hypersomnolence (66%) and hypothermia (35%). Epilepsy (54%) and gastrointestinal (69%), ophthalmological (51%) and endocrine problems (42%) were observed frequently. Computational analysis of facial photographs showed subtle facial dysmorphism. No strong genotype-phenotype correlation was identified by subgrouping mutations into functional classes. CONCLUSION: We delineate the clinical spectrum of PURA syndrome with the identification of 32 additional individuals. The identification of one individual through targeted Sanger sequencing points towards the clinical recognisability of the syndrome. Genotype-phenotype analysis showed no significant correlation between mutation classes and disease severity.

• MeSH
• abnormality očí genetika   MeSH
• DNA vazebné proteiny chemie   genetika   MeSH
• genetické asociační studie MeSH
• lidé MeSH
• mentální retardace genetika   MeSH
• mutace * MeSH
• novorozenec MeSH
• obličej abnormality   MeSH
• proteiny Drosophily chemie   genetika   MeSH
• strukturní homologie proteinů MeSH
• svalová hypotonie etiologie   genetika   MeSH
• syndrom MeSH
• těhotenství MeSH
• transkripční faktory chemie   genetika   MeSH
• Check Tag
• lidé MeSH
• novorozenec MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH