Other search tools

About this data

The publication data currently available has been vetted by Vanderbilt faculty, staff, administrators and trainees. The data itself is retrieved directly from NCBI's PubMed and is automatically updated on a weekly basis to ensure accuracy and completeness.

If you have any questions or comments, please contact us.

Results: 1 to 10 of 403

Publication Record

Connections

Human DNA polymerase η accommodates RNA for strand extension.
Su Y, Egli M, Guengerich FP
(2017) J Biol Chem 292: 18044-18051
MeSH Terms: Base Pair Mismatch, DNA Primers, DNA Replication, DNA-Directed DNA Polymerase, Deoxyguanosine, Electrophoretic Mobility Shift Assay, Humans, Kinetics, Nucleic Acid Heteroduplexes, Nucleic Acid Hybridization, Oligodeoxyribonucleotides, Oligoribonucleotides, Pyrimidine Dimers, RNA, Recombinant Proteins, Reverse Transcription, Substrate Specificity, Transcription Elongation, Genetic
Show Abstract · Added March 14, 2018
Ribonucleotides are the natural analogs of deoxyribonucleotides, which can be misinserted by DNA polymerases, leading to the most abundant DNA lesions in genomes. During replication, DNA polymerases tolerate patches of ribonucleotides on the parental strands to different extents. The majority of human DNA polymerases have been reported to misinsert ribonucleotides into genomes. However, only PrimPol, DNA polymerase α, telomerase, and the mitochondrial human DNA polymerase (hpol) γ have been shown to tolerate an entire RNA strand. Y-family hpol η is known for translesion synthesis opposite the UV-induced DNA lesion cyclobutane pyrimidine dimer and was recently found to incorporate ribonucleotides into DNA. Here, we report that hpol η is able to bind DNA/DNA, RNA/DNA, and DNA/RNA duplexes with similar affinities. In addition, hpol η, as well as another Y-family DNA polymerase, hpol κ, accommodates RNA as one of the two strands during primer extension, mainly by inserting dNMPs opposite unmodified templates or DNA lesions, such as 8-oxo-2'-deoxyguanosine or cyclobutane pyrimidine dimer, even in the presence of an equal amount of the DNA/DNA substrate. The discovery of this RNA-accommodating ability of hpol η redefines the traditional concept of human DNA polymerases and indicates potential new functions of hpol η .
© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.
0 Communities
1 Members
0 Resources
18 MeSH Terms
A Polymerase With Potential: The Fe-S Cluster in Human DNA Primase.
Holt ME, Salay LE, Chazin WJ
(2017) Methods Enzymol 595: 361-390
MeSH Terms: Catalytic Domain, DNA, DNA Primase, DNA Primers, DNA Replication, Humans, Iron-Sulfur Proteins, Oxidation-Reduction, RNA, Sequence Analysis
Show Abstract · Added March 24, 2018
Replication of DNA in eukaryotes is primarily executed by the combined action of processive DNA polymerases δ and ɛ. These enzymes cannot initiate synthesis of new DNA without the presence of a primer on the template ssDNA. The primers on both the leading and lagging strands are generated by DNA polymerase α-primase (pol-prim). DNA primase is a DNA-dependent RNA polymerase that synthesizes the first ~10 nucleotides and then transfers the substrate to polymerase α to complete primer synthesis. The mechanisms governing the coordination and handoff between primase and polymerase α are largely unknown. Isolated DNA primase contains a [4Fe-4S] cluster that has been shown to serve as a redox switch modulating DNA binding affinity. This discovery suggests a mechanism for modulating the priming activity of primase and handoff to polymerase α. In this chapter, we briefly discuss the current state of knowledge of primase structure and function, including the role of its iron-sulfur cluster. This is followed by providing the methods for expressing, purifying, and biophysically/structurally characterizing primase and its iron-sulfur cluster-containing domain, p58C.
© 2017 Elsevier Inc. All rights reserved.
0 Communities
1 Members
0 Resources
10 MeSH Terms
Polymerase Bypass of N(6)-Deoxyadenosine Adducts Derived from Epoxide Metabolites of 1,3-Butadiene.
Kotapati S, Wickramaratne S, Esades A, Boldry EJ, Quirk Dorr D, Pence MG, Guengerich FP, Tretyakova NY
(2015) Chem Res Toxicol 28: 1496-507
MeSH Terms: Butadienes, Chromatography, High Pressure Liquid, DNA, DNA Adducts, DNA Primers, DNA Replication, DNA-Directed DNA Polymerase, Deoxyadenosines, Epoxy Compounds, Humans, Kinetics, Oligodeoxyribonucleotides, Spectrometry, Mass, Electrospray Ionization
Show Abstract · Added March 14, 2018
N(6)-(2-Hydroxy-3-buten-1-yl)-2'-deoxyadenosine (N(6)-HB-dA I) and N(6),N(6)-(2,3-dihydroxybutan-1,4-diyl)-2'-deoxyadenosine (N(6),N(6)-DHB-dA) are exocyclic DNA adducts formed upon alkylation of the N(6) position of adenine in DNA by epoxide metabolites of 1,3-butadiene (BD), a common industrial and environmental chemical classified as a human and animal carcinogen. Since the N(6)-H atom of adenine is required for Watson-Crick hydrogen bonding with thymine, N(6)-alkylation can prevent adenine from normal pairing with thymine, potentially compromising the accuracy of DNA replication. To evaluate the ability of BD-derived N(6)-alkyladenine lesions to induce mutations, synthetic oligodeoxynucleotides containing site-specific (S)-N(6)-HB-dA I and (R,R)-N(6),N(6)-DHB-dA adducts were subjected to in vitro translesion synthesis in the presence of human DNA polymerases β, η, ι, and κ. While (S)-N(6)-HB-dA I was readily bypassed by all four enzymes, only polymerases η and κ were able to carry out DNA synthesis past (R,R)-N(6),N(6)-DHB-dA. Steady-state kinetic analyses indicated that all four DNA polymerases preferentially incorporated the correct base (T) opposite (S)-N(6)-HB-dA I. In contrast, hPol β was completely blocked by (R,R)-N(6),N(6)-DHB-dA, while hPol η and κ inserted A, G, C, or T opposite the adduct with similar frequency. HPLC-ESI-MS/MS analysis of primer extension products confirmed that while translesion synthesis past (S)-N(6)-HB-dA I was mostly error-free, replication of DNA containing (R,R)-N(6),N(6)-DHB-dA induced significant numbers of A, C, and G insertions and small deletions. These results indicate that singly substituted (S)-N(6)-HB-dA I lesions are not miscoding, but that exocyclic (R,R)-N(6),N(6)-DHB-dA adducts are strongly mispairing, probably due to their inability to form stable Watson-Crick pairs with dT.
0 Communities
1 Members
0 Resources
13 MeSH Terms
Heparan Sulfate Proteoglycans Are Important for Islet Amyloid Formation and Islet Amyloid Polypeptide-induced Apoptosis.
Oskarsson ME, Singh K, Wang J, Vlodavsky I, Li JP, Westermark GT
(2015) J Biol Chem 290: 15121-32
MeSH Terms: Amyloid, Animals, Apoptosis, Base Sequence, CHO Cells, Cells, Cultured, Cricetinae, Cricetulus, DNA Primers, Glucuronidase, Heparan Sulfate Proteoglycans, Humans, Islet Amyloid Polypeptide, Islets of Langerhans, Mice, Mice, Inbred C57BL, Mice, Transgenic, Real-Time Polymerase Chain Reaction
Show Abstract · Added June 6, 2017
Deposition of β cell toxic islet amyloid is a cardinal finding in type 2 diabetes. In addition to the main amyloid component islet amyloid polypeptide (IAPP), heparan sulfate proteoglycan is constantly present in the amyloid deposit. Heparan sulfate (HS) side chains bind to IAPP, inducing conformational changes of the IAPP structure and an acceleration of fibril formation. We generated a double-transgenic mouse strain (hpa-hIAPP) that overexpresses human heparanase and human IAPP but is deficient of endogenous mouse IAPP. Culture of hpa-hIAPP islets in 20 mm glucose resulted in less amyloid formation compared with the amyloid load developed in cultured islets isolated from littermates expressing human IAPP only. A similar reduction of amyloid was achieved when human islets were cultured in the presence of heparin fragments. Furthermore, we used CHO cells and the mutant CHO pgsD-677 cell line (deficient in HS synthesis) to explore the effect of cellular HS on IAPP-induced cytotoxicity. Seeding of IAPP aggregation on CHO cells resulted in caspase-3 activation and apoptosis that could be prevented by inhibition of caspase-8. No IAPP-induced apoptosis was seen in HS-deficient CHO pgsD-677 cells. These results suggest that β cell death caused by extracellular IAPP requires membrane-bound HS. The interaction between HS and IAPP or the subsequent effects represent a possible therapeutic target whose blockage can lead to a prolonged survival of β cells.
© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.
1 Communities
0 Members
0 Resources
18 MeSH Terms
Preserving Mafa expression in diabetic islet β-cells improves glycemic control in vivo.
Matsuoka TA, Kaneto H, Kawashima S, Miyatsuka T, Tochino Y, Yoshikawa A, Imagawa A, Miyazaki J, Gannon M, Stein R, Shimomura I
(2015) J Biol Chem 290: 7647-57
MeSH Terms: Animals, Base Sequence, Blood Glucose, DNA Primers, Diabetes Mellitus, Experimental, Humans, Islets of Langerhans, Maf Transcription Factors, Large, Mice, Mice, Inbred C57BL, Real-Time Polymerase Chain Reaction
Show Abstract · Added September 28, 2015
The murine Mafa transcription factor is a key regulator of postnatal islet β-cell activity, affecting insulin transcription, insulin secretion, and β-cell mass. Human MAFA expression is also markedly decreased in islet β-cells of type 2 diabetes mellitus (T2DM) patients. Moreover, levels are profoundly reduced in db/db islet β-cells, a mouse model of T2DM. To examine the significance of this key islet β-cell-enriched protein to glycemic control under diabetic conditions, we generated transgenic mice that conditionally and specifically produced Mafa in db/db islet β-cells. Sustained expression of Mafa resulted in significantly lower plasma glucose levels, higher plasma insulin, and augmented islet β-cell mass. In addition, there was increased expression of insulin, Slc2a2, and newly identified Mafa-regulated genes involved in reducing β-cell stress, like Gsta1 and Gckr. Importantly, the levels of human GSTA1 were also compromised in T2DM islets. Collectively, these results illustrate how consequential the reduction in Mafa activity is to islet β-cell function under pathophysiological conditions.
© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.
1 Communities
1 Members
0 Resources
11 MeSH Terms
miR-216a regulates snx5, a novel notch signaling pathway component, during zebrafish retinal development.
Olena AF, Rao MB, Thatcher EJ, Wu SY, Patton JG
(2015) Dev Biol 400: 72-81
MeSH Terms: Analysis of Variance, Animals, Cloning, Molecular, DNA Primers, Gene Expression Profiling, Gene Expression Regulation, Developmental, Gene Knockdown Techniques, Image Processing, Computer-Assisted, Immunoblotting, In Situ Hybridization, Intracellular Signaling Peptides and Proteins, Membrane Proteins, MicroRNAs, Microarray Analysis, Models, Biological, Receptors, Notch, Retina, Signal Transduction, Sorting Nexins, Ubiquitin-Protein Ligases, Zebrafish, Zebrafish Proteins
Show Abstract · Added February 4, 2016
Precise regulation of Notch signaling is essential for normal vertebrate development. Mind bomb (Mib) is a ubiquitin ligase that is required for activation of Notch by Notch׳s ligand, Delta. Sorting Nexin 5 (SNX5) co-localizes with Mib and Delta complexes and has been shown to directly bind to Mib. We show that microRNA-216a (miR-216a) is expressed in the retina during early development and regulates snx5 to precisely regulate Notch signaling. miR-216a and snx5 have complementary expression patterns. Knocking down miR-216a and/or overexpression of snx5 resulted in increased Notch activation. Conversely, knocking down snx5 and/or miR-216a overexpression caused a decrease in Notch activation. We propose a model in which SNX5, precisely controlled by miR-216a, is a vital partner of Mib in promoting endocytosis of Delta and subsequent activation of Notch signaling.
Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
0 Communities
1 Members
0 Resources
22 MeSH Terms
Insulin-independent role of adiponectin receptor signaling in Drosophila germline stem cell maintenance.
Laws KM, Sampson LL, Drummond-Barbosa D
(2015) Dev Biol 399: 226-36
MeSH Terms: Adipocytes, Animals, Animals, Genetically Modified, Cloning, Molecular, DNA Primers, Drosophila, Drosophila Proteins, Female, Gene Expression Regulation, Developmental, Germ Cells, Image Processing, Computer-Assisted, Insulin, Microscopy, Fluorescence, Ovary, Receptors, Adiponectin, Reverse Transcriptase Polymerase Chain Reaction, Signal Transduction, Stem Cells
Show Abstract · Added March 19, 2017
Adipocytes have key endocrine roles, mediated in large part by secreted protein hormones termed adipokines. The adipokine adiponectin is well known for its role in sensitizing peripheral tissues to insulin, and several lines of evidence suggest that adiponectin might also modulate stem cells/precursors. It remains unclear, however, how adiponectin signaling controls stem cells and whether this role is secondary to its insulin-sensitizing effects or distinct. Drosophila adipocytes also function as an endocrine organ and, although no obvious adiponectin homolog has been identified, Drosophila AdipoR encodes a well-conserved homolog of mammalian adiponectin receptors. Here, we generate a null AdipoR allele and use clonal analysis to demonstrate an intrinsic requirement for AdipoR in germline stem cell (GSC) maintenance in the Drosophila ovary. AdipoR null GSCs are not fully responsive to bone morphogenetic protein ligands from the niche and have a slight reduction in E-cadherin levels at the GSC-niche junction. Conversely, germline-specific overexpression of AdipoR inhibits natural GSC loss, suggesting that reduction in adiponectin signaling might contribute to the normal decline in GSC numbers observed over time in wild-type females. Surprisingly, AdipoR is not required for insulin sensitization of the germline, leading us to speculate that insulin sensitization is a more recently acquired function than stem cell regulation in the evolutionary history of adiponectin signaling. Our findings establish Drosophila female GSCs as a new system for future studies addressing the molecular mechanisms whereby adiponectin receptor signaling modulates stem cell fate.
Copyright © 2015 Elsevier Inc. All rights reserved.
0 Communities
1 Members
0 Resources
18 MeSH Terms
Human PrimPol is a highly error-prone polymerase regulated by single-stranded DNA binding proteins.
Guilliam TA, Jozwiakowski SK, Ehlinger A, Barnes RP, Rudd SG, Bailey LJ, Skehel JM, Eckert KA, Chazin WJ, Doherty AJ
(2015) Nucleic Acids Res 43: 1056-68
MeSH Terms: DNA Primase, DNA Primers, DNA Replication, DNA-Binding Proteins, DNA-Directed DNA Polymerase, Humans, Mitochondrial Proteins, Multifunctional Enzymes, Mutagenesis, Proliferating Cell Nuclear Antigen, Protein Interaction Domains and Motifs, Replication Protein A
Show Abstract · Added January 20, 2015
PrimPol is a recently identified polymerase involved in eukaryotic DNA damage tolerance, employed in both re-priming and translesion synthesis mechanisms to bypass nuclear and mitochondrial DNA lesions. In this report, we investigate how the enzymatic activities of human PrimPol are regulated. We show that, unlike other TLS polymerases, PrimPol is not stimulated by PCNA and does not interact with it in vivo. We identify that PrimPol interacts with both of the major single-strand binding proteins, RPA and mtSSB in vivo. Using NMR spectroscopy, we characterize the domains responsible for the PrimPol-RPA interaction, revealing that PrimPol binds directly to the N-terminal domain of RPA70. In contrast to the established role of SSBs in stimulating replicative polymerases, we find that SSBs significantly limit the primase and polymerase activities of PrimPol. To identify the requirement for this regulation, we employed two forward mutation assays to characterize PrimPol's replication fidelity. We find that PrimPol is a mutagenic polymerase, with a unique error specificity that is highly biased towards insertion-deletion errors. Given the error-prone disposition of PrimPol, we propose a mechanism whereby SSBs greatly restrict the contribution of this enzyme to DNA replication at stalled forks, thus reducing the mutagenic potential of PrimPol during genome replication.
© The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.
1 Communities
3 Members
0 Resources
12 MeSH Terms
Induction of diverse cardiac cell types by reprogramming fibroblasts with cardiac transcription factors.
Nam YJ, Lubczyk C, Bhakta M, Zang T, Fernandez-Perez A, McAnally J, Bassel-Duby R, Olson EN, Munshi NV
(2014) Development 141: 4267-78
MeSH Terms: Action Potentials, Analysis of Variance, Animals, Basic Helix-Loop-Helix Transcription Factors, Cell Differentiation, DNA Primers, Embryonic Induction, Fibroblasts, GATA4 Transcription Factor, Green Fluorescent Proteins, Heart, Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels, Immunohistochemistry, MEF2 Transcription Factors, Mice, Myocytes, Cardiac, Real-Time Polymerase Chain Reaction, T-Box Domain Proteins, Transcription Factors
Show Abstract · Added April 2, 2019
Various combinations of cardiogenic transcription factors, including Gata4 (G), Hand2 (H), Mef2c (M) and Tbx5 (T), can reprogram fibroblasts into induced cardiac-like myocytes (iCLMs) in vitro and in vivo. Given that optimal cardiac function relies on distinct yet functionally interconnected atrial, ventricular and pacemaker (PM) cardiomyocytes (CMs), it remains to be seen which subtypes are generated by direct reprogramming and whether this process can be harnessed to produce a specific CM of interest. Here, we employ a PM-specific Hcn4-GFP reporter mouse and a spectrum of CM subtype-specific markers to investigate the range of cellular phenotypes generated by reprogramming of primary fibroblasts. Unexpectedly, we find that a combination of four transcription factors (4F) optimized for Hcn4-GFP expression does not generate beating PM cells due to inadequate sarcomeric protein expression and organization. However, applying strict single-cell criteria to GHMT-reprogrammed cells, we observe induction of diverse cellular phenotypes, including those resembling immature forms of all three major cardiac subtypes (i.e. atrial, ventricular and pacemaker). In addition, we demonstrate that cells induced by GHMT are directly reprogrammed and do not arise from an Nxk2.5(+) progenitor cell intermediate. Taken together, our results suggest a remarkable degree of plasticity inherent to GHMT reprogramming and provide a starting point for optimization of CM subtype-specific reprogramming protocols.
© 2014. Published by The Company of Biologists Ltd.
0 Communities
1 Members
0 Resources
MeSH Terms
Biochemical analysis of six genetic variants of error-prone human DNA polymerase ι involved in translesion DNA synthesis.
Kim J, Song I, Jo A, Shin JH, Cho H, Eoff RL, Guengerich FP, Choi JY
(2014) Chem Res Toxicol 27: 1837-52
MeSH Terms: Amino Acid Substitution, Base Sequence, DNA, DNA Primers, DNA-Directed DNA Polymerase, Fluorescence Polarization, Guanosine, Humans, Kinetics, Magnesium, Manganese, Protein Binding, Protein Structure, Tertiary, Recombinant Proteins
Show Abstract · Added January 20, 2015
DNA polymerase (pol) ι is the most error-prone among the Y-family polymerases that participate in translesion synthesis (TLS). Pol ι can bypass various DNA lesions, e.g., N(2)-ethyl(Et)G, O(6)-methyl(Me)G, 8-oxo-7,8-dihydroguanine (8-oxoG), and an abasic site, though frequently with low fidelity. We assessed the biochemical effects of six reported genetic variations of human pol ι on its TLS properties, using the recombinant pol ι (residues 1-445) proteins and DNA templates containing a G, N(2)-EtG, O(6)-MeG, 8-oxoG, or abasic site. The Δ1-25 variant, which is the N-terminal truncation of 25 residues resulting from an initiation codon variant (c.3G > A) and also is the formerly misassigned wild-type, exhibited considerably higher polymerase activity than wild-type with Mg(2+) (but not with Mn(2+)), coinciding with its steady-state kinetic data showing a ∼10-fold increase in kcat/Km for nucleotide incorporation opposite templates (only with Mg(2+)). The R96G variant, which lacks a R96 residue known to interact with the incoming nucleotide, lost much of its polymerase activity, consistent with the kinetic data displaying 5- to 72-fold decreases in kcat/Km for nucleotide incorporation opposite templates either with Mg(2+) or Mn(2+), except for that opposite N(2)-EtG with Mn(2+) (showing a 9-fold increase for dCTP incorporation). The Δ1-25 variant bound DNA 20- to 29-fold more tightly than wild-type (with Mg(2+)), but the R96G variant bound DNA 2-fold less tightly than wild-type. The DNA-binding affinity of wild-type, but not of the Δ1-25 variant, was ∼7-fold stronger with 0.15 mM Mn(2+) than with Mg(2+). The results indicate that the R96G variation severely impairs most of the Mg(2+)- and Mn(2+)-dependent TLS abilities of pol ι, whereas the Δ1-25 variation selectively and substantially enhances the Mg(2+)-dependent TLS capability of pol ι, emphasizing the potential translational importance of these pol ι genetic variations, e.g., individual differences in TLS, mutation, and cancer susceptibility to genotoxic carcinogens.
0 Communities
1 Members
0 Resources
14 MeSH Terms