scholarly journals Nanopore Analysis of Wild-Type and Mutant Prion Protein (PrPC): Single Molecule Discrimination and PrPC Kinetics

PLoS ONE ◽  
2013 ◽  
Vol 8 (2) ◽  
pp. e54982 ◽  
Author(s):  
Nahid N. Jetha ◽  
Valentyna Semenchenko ◽  
David S. Wishart ◽  
Neil R. Cashman ◽  
Andre Marziali
Keyword(s):  
Prion Protein ◽  
Single Molecule ◽  
Wild Type

scholarly journals Targeting the DNA replication stress phenotype of KRAS mutant cancer cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Tara Al Zubaidi ◽  
O. H. Fiete Gehrisch ◽  
Marie-Michelle Genois ◽  
Qi Liu ◽  
Shan Lu ◽  
...  
Keyword(s):  
Dna Replication ◽  
Single Molecule ◽  
Replication Stress ◽  
Proton Radiation ◽  
Wild Type ◽  
Cellular Effects ◽  
Proton Treatment ◽  
Dna Replication Stress ◽  
Fork Stalling ◽  
Mutant Cells

AbstractMutant KRAS is a common tumor driver and frequently confers resistance to anti-cancer treatments such as radiation. DNA replication stress in these tumors may constitute a therapeutic liability but is poorly understood. Here, using single-molecule DNA fiber analysis, we first characterized baseline replication stress in a panel of unperturbed isogenic and non-isogenic cancer cell lines. Correlating with the observed enhanced replication stress we found increased levels of cytosolic double-stranded DNA in KRAS mutant compared to wild-type cells. Yet, despite this phenotype replication stress-inducing agents failed to selectively impact KRAS mutant cells, which were protected by CHK1. Similarly, most exogenous stressors studied did not differentially augment cytosolic DNA accumulation in KRAS mutant compared to wild-type cells. However, we found that proton radiation was able to slow fork progression and preferentially induce fork stalling in KRAS mutant cells. Proton treatment also partly reversed the radioresistance associated with mutant KRAS. The cellular effects of protons in the presence of KRAS mutation clearly contrasted that of other drugs affecting replication, highlighting the unique nature of the underlying DNA damage caused by protons. Taken together, our findings provide insight into the replication stress response associated with mutated KRAS, which may ultimately yield novel therapeutic opportunities.

scholarly journals Expression of the Prion Protein Family Member Shadoo Causes Drug Hypersensitivity That Is Diminished by the Coexpression of the Wild Type Prion Protein

2015 ◽  
Vol 291 (9) ◽  
pp. 4473-4486 ◽  
Author(s):  
Antal Nyeste ◽  
Petra Bencsura ◽  
István Vida ◽  
Zoltán Hegyi ◽  
László Homolya ◽  
...  
Keyword(s):  
Family Member ◽  
Prion Protein ◽  
Drug Hypersensitivity ◽  
Protein Family ◽  
Wild Type

scholarly journals KIF18A's neck linker permits navigation of microtubule-bound obstacles within the mitotic spindle

2019 ◽  
Vol 2 (1) ◽  
pp. e201800169 ◽  
Author(s):  
Heidi LH Malaby ◽  
Dominique V Lessard ◽  
Christopher L Berger ◽  
Jason Stumpff
Keyword(s):  
Single Molecule ◽  
Mitotic Spindle ◽  
Binding Proteins ◽  
Mitotic Chromosome ◽  
Chromosome Movement ◽  
Wild Type ◽  
Microtubule Associated Proteins ◽  
Chromosome Alignment ◽  
Associated Proteins ◽  
Fiber Dynamics

KIF18A (kinesin-8) is required for mammalian mitotic chromosome alignment. KIF18A confines chromosome movement to the mitotic spindle equator by accumulating at the plus-ends of kinetochore microtubule bundles (K-fibers), where it functions to suppress K-fiber dynamics. It is not understood how the motor accumulates at K-fiber plus-ends, a difficult feat requiring the motor to navigate protein dense microtubule tracks. Our data indicate that KIF18A's relatively long neck linker is required for the motor's accumulation at K-fiber plus-ends. Shorter neck linker (sNL) variants of KIF18A display a deficiency in accumulation at the ends of K-fibers at the center of the spindle. Depletion of K-fiber–binding proteins reduces the KIF18A sNL localization defect, whereas their overexpression reduces wild-type KIF18A's ability to accumulate on this same K-fiber subset. Furthermore, single-molecule assays indicate that KIF18A sNL motors are less proficient in navigating microtubules coated with microtubule-associated proteins. Taken together, these results support a model in which KIF18A's neck linker length permits efficient navigation of obstacles to reach K-fiber ends during mitosis.

scholarly journals Limited effects of m6A modification on mRNA partitioning into stress granules

2021 ◽  
Author(s):  
Anthony Khong ◽  
Tyler Matheny ◽  
Thao Ngoc Huynh ◽  
Vincent Babl ◽  
Roy Parker
Keyword(s):  
Regression Analysis ◽  
Linear Regression ◽  
Single Molecule ◽  
Linear Regression Analysis ◽  
Stress Granules ◽  
Multiple Linear Regression Analysis ◽  
Minor Role ◽  
Wild Type ◽  
Specific Mrnas ◽  
A Minor

Recent studies have argued that the m6A modification of mRNAs promotes mRNA recruitment to stress granules through the interaction with YTHDF proteins (Anders et al., 2018; Ries et al., 2019). However, mRNAs that contain multiple m6A modified sites partition similarly into stress granules in both wild-type and m6A-deficient cells by single-molecule FISH suggesting m6A modifications play a minor role in mRNA partitioning into stress granules. Moreover, multiple linear regression analysis suggests m6A modification plays a minimal role in stress granule recruitment. Finally, the artificial tethering of 25 YTHDF proteins on reporter mRNAs leads to only a modest increase in mRNA partitioning to stress granules. These results indicate m6A modification makes a small, but measurable, contribution to recruiting specific mRNAs to stress granules.

scholarly journals Analysis of the Complete Genome Sequence of a Novel, Pseudorabies Virus Strain Isolated in Southeast Europe

2019 ◽  
Vol 2019 ◽  
pp. 1-12
Author(s):  
Zsolt Csabai ◽  
Dóra Tombácz ◽  
Zoltán Deim ◽  
Michael Snyder ◽  
Zsolt Boldogkői
Keyword(s):  
Single Molecule ◽  
Base Composition ◽  
Pseudorabies Virus ◽  
Point Mutations ◽  
Wild Type Virus ◽  
Economic Losses ◽  
Wild Type ◽  
Southeast Europe ◽  
Long Read ◽  
National Programs

Background. Pseudorabies virus (PRV) is the causative agent of Aujeszky’s disease giving rise to significant economic losses worldwide. Many countries have implemented national programs for the eradication of this virus. In this study, long-read sequencing was used to determine the nucleotide sequence of the genome of a novel PRV strain (PRV-MdBio) isolated in Serbia.Results. In this study, a novel PRV strain was isolated and characterized. PRV-MdBio was found to exhibit similar growth properties to those of another wild-type PRV, the strain Kaplan. Single-molecule real-time (SMRT) sequencing has revealed that the new strain differs significantly in base composition even from strain Kaplan, to which it otherwise exhibits the highest similarity. We compared the genetic composition of PRV-MdBio to strain Kaplan and the China reference strain Ea and obtained that radical base replacements were the most common point mutations preceding conservative and silent mutations. We also found that the adaptation of PRV to cell culture does not lead to any tendentious genetic alteration in the viral genome.Conclusion. PRV-MdBio is a wild-type virus, which differs in base composition from other PRV strains to a relatively large extent.

scholarly journals A Transposon Story: From TE Content to TE Dynamic Invasion of Drosophila Genomes Using the Single-Molecule Sequencing Technology from Oxford Nanopore

Cells ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 1776
Author(s):  
Mourdas Mohamed ◽  
Nguyet Thi-Minh Dang ◽  
Yuki Ogyama ◽  
Nelly Burlet ◽  
Bruno Mugat ◽  
...  
Keyword(s):  
Single Molecule ◽  
Wild Type ◽  
Sequencing Data ◽  
Short Read ◽  
Short Read Sequencing ◽  
Sequencing Technologies ◽  
Oxford Nanopore ◽  
In The Wild ◽  
Successive Generations ◽  
Type Strains

Transposable elements (TEs) are the main components of genomes. However, due to their repetitive nature, they are very difficult to study using data obtained with short-read sequencing technologies. Here, we describe an efficient pipeline to accurately recover TE insertion (TEI) sites and sequences from long reads obtained by Oxford Nanopore Technology (ONT) sequencing. With this pipeline, we could precisely describe the landscapes of the most recent TEIs in wild-type strains of Drosophila melanogaster and Drosophila simulans. Their comparison suggests that this subset of TE sequences is more similar than previously thought in these two species. The chromosome assemblies obtained using this pipeline also allowed recovering piRNA cluster sequences, which was impossible using short-read sequencing. Finally, we used our pipeline to analyze ONT sequencing data from a D. melanogaster unstable line in which LTR transposition was derepressed for 73 successive generations. We could rely on single reads to identify new insertions with intact target site duplications. Moreover, the detailed analysis of TEIs in the wild-type strains and the unstable line did not support the trap model claiming that piRNA clusters are hotspots of TE insertions.

P4-124: Comparison of gene expression profiles of prion protein-deficient and wild type neuronal cell lines of mice

2006 ◽  
Vol 2 ◽  
pp. S552-S552
Author(s):  
Boe-Hyun Kim ◽  
Jae-Il Kim ◽  
Eun-Kyoung Choi ◽  
Richard I. Carp ◽  
Yong-Sun Kim
Keyword(s):  
Gene Expression ◽  
Prion Protein ◽  
Cell Lines ◽  
Expression Profiles ◽  
Neuronal Cell ◽  
Gene Expression Profiles ◽  
Wild Type ◽  
Neuronal Cell Lines

scholarly journals Single-molecule imaging of translational output from individual RNA granules in neurons

2012 ◽  
Vol 23 (5) ◽  
pp. 918-929 ◽  
Author(s):  
Vedakumar Tatavarty ◽  
Marius F. Ifrim ◽  
Mikhail Levin ◽  
George Korza ◽  
Elisa Barbarese ◽  
...  
Keyword(s):  
Single Molecule ◽  
Fluorescent Protein ◽  
Fragile X ◽  
Single Molecule Imaging ◽  
Wild Type ◽  
Rna Molecules ◽  
Temporal Regulation ◽  
Rna Granules ◽  
Mental Retardation Protein ◽  
Insight Into

Dendritic RNAs are localized and translated in RNA granules. Here we use single-molecule imaging to count the number of RNA molecules in each granule and to record translation output from each granule using Venus fluorescent protein as a reporter. For RNAs encoding activity-regulated cytoskeletal-associated protein (ARC) or fragile X mental retardation protein (FMRP), translation events are spatially clustered near individual granules, and translational output from individual granules is either sporadic or bursty. The probability of bursty translation is greater for Venus-FMRP RNA than for Venus-ARC RNA and is increased in Fmr1-knockout neurons compared to wild-type neurons. Dihydroxyphenylglycine (DHPG) increases the rate of sporadic translation and decreases bursty translation for Venus-FMRP and Venus-ARC RNAs. Single-molecule imaging of translation in individual granules provides new insight into molecular, spatial, and temporal regulation of translation in granules.

scholarly journals Bottom-up single-molecule strategy for understanding subunit function of tetrameric β-galactosidase

2018 ◽  
Vol 115 (33) ◽  
pp. 8346-8351 ◽  
Author(s):  
Xiang Li ◽  
Yu Jiang ◽  
Shaorong Chong ◽  
David R. Walt
Keyword(s):  
Single Molecule ◽  
Wild Type ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Single Molecule Level ◽  
Subunit Function ◽  
Enzyme Molecules ◽  
Individual Enzyme ◽  
Kinetics Of

In this paper, we report an example of the engineered expression of tetrameric β-galactosidase (β-gal) containing varying numbers of active monomers. Specifically, by combining wild-type and single-nucleotide polymorphism plasmids at varying ratios, tetrameric β-gal was expressed in vitro with one to four active monomers. The kinetics of individual enzyme molecules revealed four distinct populations, corresponding to the number of active monomers in the enzyme. Using single-molecule-level enzyme kinetics, we were able to measure an accurate in vitro mistranslation frequency (5.8 × 10−4 per base). In addition, we studied the kinetics of the mistranslated β-gal at the single-molecule level.

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