scholarly journals RNA polymerase errors cause splicing defects and can be regulated by differential expression of RNA polymerase subunits

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Lucas B Carey
Keyword(s):  
Rna Polymerase ◽  
Differential Expression ◽  
Dna Polymerase ◽  
Error Rate ◽  
Single Cells ◽  
Coding Sequences ◽  
Polymerase Fidelity ◽  
Splicing Defects ◽  
Cellular Phenotypes ◽  
Rna Polymerase Subunits

Errors during transcription may play an important role in determining cellular phenotypes: the RNA polymerase error rate is >4 orders of magnitude higher than that of DNA polymerase and errors are amplified >1000-fold due to translation. However, current methods to measure RNA polymerase fidelity are low-throughout, technically challenging, and organism specific. Here I show that changes in RNA polymerase fidelity can be measured using standard RNA sequencing protocols. I find that RNA polymerase is error-prone, and these errors can result in splicing defects. Furthermore, I find that differential expression of RNA polymerase subunits causes changes in RNA polymerase fidelity, and that coding sequences may have evolved to minimize the effect of these errors. These results suggest that errors caused by RNA polymerase may be a major source of stochastic variability at the level of single cells.

Oligo(A)-stimulated Tetrahymena rDNA synthesis in vitro

1981 ◽  
Vol 59 (6) ◽  
pp. 396-403 ◽  
Author(s):  
Peter R. Ganz ◽  
Gyorgy B. Kiss ◽  
Ronald E. Pearlman
Keyword(s):  
Rna Polymerase ◽  
Dna Synthesis ◽  
Dna Polymerase ◽  
Ribosomal Rna ◽  
Replication Proteins ◽  
General Applicability ◽  
In Vitro Synthesis ◽  
Restriction Fragments ◽  
Double Stranded Dna

The synthesis of Tetrahymena rDNA has been examined using purified DNA polymerase and partially purified preparations of homologous replication enzymes (fraction IV). DNA synthesis with purified DNA polymerase alone was less than that with fraction IV enzymes. This suggested that there were additional factors in fraction IV other than DNA polymerase which contributed to or enhanced rDNA synthesis in vitro. Neither hybridization of rDNA with Tetrahymena ribosomal RNA nor preincubation of rDNA with homologous or heterologous RNA polymerase served to stimulate in vitro synthesis by fraction IV enzymes. However, when rDNA was hybridized with oligoriboadenylate, DNA synthesis using fraction IV was stimulated approximately 4- to 4.5-fold over 150 min of incubation, relative to a similarly treated but unhybridized rDNA control. Using oligoriboadenylate-hybridized EcoR1 and HindIII restriction fragments of rDNA to localize the synthesis most of the in vitro synthesis occurred within a 2.4 × 106 Mr fragment encompassing the centre of the rDNA molecule. The approach of hybridizing a synthetic homooligoribonucleotide primer to double-stranded DNA should prove to be of general applicability in designing similar template–primers in other systems for the purpose of isolating replication proteins.

Linear plasmids that integrate into mitochondrial DNA in Neurospora

Genome ◽  
1989 ◽  
Vol 31 (1) ◽  
pp. 155-159 ◽  
Author(s):  
H. Bertrand ◽  
A. J. F. Griffiths
Keyword(s):  
Mitochondrial Dna ◽  
Rna Polymerase ◽  
Dna Polymerase ◽  
Inverted Repeats ◽  
Insertion Sequences ◽  
Long Terminal Repeats ◽  
Linear Plasmids ◽  
Mt Dna ◽  
Common Features ◽  
Mitochondrial Chromosome

In some field isolates of Neurospora from Hawaii and India, senescence is induced by integration of linear DNA plasmids, kalilo and maranhar, respectively, into mitochondrial (mt) DNA. Although the two plasmids show little homology at the DNA level, both have inverted long terminal repeats, and each potentially encodes a DNA polymerase and a RNA polymerase. Both plasmids generate very long inverted repeats of mtDNA at their ends upon integration into mitochondrial chromosomes. Hence, they appear to integrate by a mechanism that involves pairing of both ends of the plasmid with short stretches of homologous nucleotide sequences in mtDNA. This recombinogenic association apparently generates an origin for an unscheduled round of replication of mtDNA. In the process, the resulting two copies of the mitochondrial chromosome are joined to opposite ends of the plasmid. A model for the senescence-associated accumulation of mtDNAs with plasmid insertion sequences is proposed on the basis of common features that characterize senescence in a variety of filamentous fungi.Key words: Neurospora, senescence, plasmids, mitochondria.

Theoretical analysis of RNA polymerase fidelity: a steady-state copolymerization approach

2021 ◽  
Author(s):  
Wenbo Fu ◽  
Qiushi Li ◽  
Yongshun Song ◽  
Yaogen Shu ◽  
Zhongcan Ouyang ◽  
...  
Keyword(s):  
Steady State ◽  
Dna Replication ◽  
Theoretical Analysis ◽  
Rna Polymerase ◽  
Higher Order ◽  
Simplified Models ◽  
Rna Transcription ◽  
Dna Transcription ◽  
Polymerase Fidelity ◽  
Neighbor Effects

Abstract The fidelity of DNA transcription catalyzed by RNA polymerase (RNAP) has long been an important issue in biology. Experiments have revealed that RNAP can incorporate matched nucleotides selectively and proofread the incorporated mismatched nucleotides. However, systematic theoretical researches on RNAP fidelity are still lacking. In the last decade, several theories on RNA transcription have been proposed, but they only handled highly simplified models without considering the high-order neighbor effects and the oligonucleotides cleavage both of which are critical for the overall fidelity. In this paper, we regard RNA transcription as a binary copolymerization process and calculate the transcription fidelity by the steady-state copolymerization theory recently proposed by us for DNA replication. With this theory, the more realistic models considering higher-order neighbor effects, oligonucleotides cleavage, multi-step incorporation and multi-step cleavage can be rigorously handled.

scholarly journals Big knowledge from big data in functional genomics

2017 ◽  
Vol 1 (3) ◽  
pp. 245-248 ◽  
Author(s):  
Chris P. Ponting
Keyword(s):  
Big Data ◽  
Molecular Interactions ◽  
Human Population ◽  
Single Cells ◽  
Human Biology ◽  
Reverse Causation ◽  
Downstream Effects ◽  
Dna Variants ◽  
Cellular Phenotypes ◽  
Why Questions

With so much genomics data being produced, it might be wise to pause and consider what purpose this data can or should serve. Some improve annotations, others predict molecular interactions, but few add directly to existing knowledge. This is because sequence annotations do not always implicate function, and molecular interactions are often irrelevant to a cell's or organism's survival or propagation. Merely correlative relationships found in big data fail to provide answers to the Why questions of human biology. Instead, those answers are expected from methods that causally link DNA changes to downstream effects without being confounded by reverse causation. These approaches require the controlled measurement of the consequences of DNA variants, for example, either those introduced in single cells using CRISPR/Cas9 genome editing or that are already present across the human population. Inferred causal relationships between genetic variation and cellular phenotypes or disease show promise to rapidly grow and underpin our knowledge base.

scholarly journals Single-cell tumor-immune microenvironment of BRCA1/2 mutated high-grade serous ovarian cancer

2021 ◽  
Author(s):  
Inga-Maria Launonen ◽  
Nuppu Lyytikäinen ◽  
Julia Casado ◽  
Ella Anttila ◽  
Angéla Szabó ◽  
...  
Keyword(s):  
Tumor Microenvironment ◽  
Single Cell ◽  
Single Cells ◽  
High Grade ◽  
Prognostic Role ◽  
Immune Microenvironment ◽  
Proteomic Data ◽  
Tumor Immune Microenvironment ◽  
Cellular Phenotypes

Abstract The majority of high-grade serous ovarian cancers (HGSCs) are deficient in homologous recombination (HR) DNA repair, most commonly due to mutations or hypermethylation of the BRCA1/2 genes. We aimed to discover how BRCA1/2 mutations shape the cellular phenotypes and spatial interactions of the tumor microenvironment. Using a highly multiplex immunofluorescence and image analysis we generated spatial proteomic data for 21 markers in 124,623 single cells from 112 tumor cores originating from 31 tumors with BRCA1/2 mutation (BRCA1/2mut), and from 13 tumors without alterations in HR genes (HRwt). We identified a phenotypically distinct tumor microenvironment in the BRCA1/2mut tumors with evidence of increased immunosurveillance. Importantly, we found an opposing prognostic role of a proliferative tumor-cell phenotypic subpopulation in the HR-genotypes, which associated with enhanced spatial tumor-immune interactions by the CD8+ and CD4+T-cells in BRCA1/2mut tumors. The single-cell spatial landscapes indicate distinct patterns of spatial immunosurveillance with the premise to improve immunotherapeutic strategies and patient stratification in HGSC.

scholarly journals DEsingle for detecting three types of differential expression in single-cell RNA-seq data

2017 ◽  
Author(s):  
Zhun Miao ◽  
Ke Deng ◽  
Xiaowo Wang ◽  
Xuegong Zhang
Keyword(s):  
Single Cell ◽  
Differential Expression ◽  
Negative Binomial ◽  
Single Cells ◽  
R Package ◽  
Supplementary Information ◽  
Binomial Model ◽  
Supplementary Data ◽  
Rna Seq ◽  
Real Zeros

AbstractSummaryThe excessive amount of zeros in single-cell RNA-seq data include “real” zeros due to the on-off nature of gene transcription in single cells and “dropout” zeros due to technical reasons. Existing differential expression (DE) analysis methods cannot distinguish these two types of zeros. We developed an R package DEsingle which employed Zero-Inflated Negative Binomial model to estimate the proportion of real and dropout zeros and to define and detect 3 types of DE genes in single-cell RNA-seq data with higher accuracy.Availability and ImplementationThe R package DEsingle is freely available at https://github.com/miaozhun/DEsingle and is under Bioconductor’s consideration [email protected] informationSupplementary data are available at bioRxiv online.

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