A5019902933- RNA and protein synthesis mechanisms
- CRISPR and Genetic Engineering
- Advanced biosensing and bioanalysis techniques
- RNA Research and Splicing
- RNA Interference and Gene Delivery
- RNA modifications and cancer
- MicroRNA in disease regulation
- Nitric Oxide and Endothelin Effects
- Cancer-related molecular mechanisms research
- RNA regulation and disease
- Insect symbiosis and bacterial influences
- ATP Synthase and ATPases Research
- Bacteriophages and microbial interactions
- Cancer, Hypoxia, and Metabolism
- Genomics and Phylogenetic Studies
- Congenital heart defects research
- interferon and immune responses
- Renin-Angiotensin System Studies
- Epigenetics and DNA Methylation
- Electron Spin Resonance Studies
- Plant Virus Research Studies
- Hormonal Regulation and Hypertension
- Mitochondrial Function and Pathology
- Innovation and Socioeconomic Development
- Chromosomal and Genetic Variations
New England Biolabs (United States)
2012-2024
New England Biolabs (China)
2023
University of Toronto
1999-2013
Mount Sinai Hospital
2008
University Health Network
2008
Lunenfeld-Tanenbaum Research Institute
2008
Hospital for Sick Children
2008
University of Massachusetts Chan Medical School
2005-2007
St. Michael's Hospital
1999-2003
Emory University
1999
Abstract Bacterial Cas9 nucleases from type II CRISPR-Cas antiviral defence systems have been repurposed as genome editing tools. Although these proteins are found in many microbes, only a handful of variants used for applications. Here, we use bioinformatic and biochemical analyses to explore this largely uncharacterized diversity. We apply cell-free screens assess the protospacer adjacent motif (PAM) guide RNA (gRNA) requirements 79 proteins, thus identifying at least 7 distinct gRNA...
The basis for the endothelial cell-restricted expression of nitric-oxide synthase (eNOS) is not known. While transgenic promoter/reporter mice demonstrated endothelium cell-specific eNOS expression, we found robust episomal constructs in cell types that do express native transcript. To explore mechanism underlying this differential activity pattern chromatin-versus episome-based promoters, examined methylation status 5'-regulatory sequences human gene. DNA differed dramatically between and...
APOBEC3G is an antiviral host factor capable of inhibiting the replication both exogenous and endogenous retroviruses as well hepatitis B, a DNA virus that replicates through RNA intermediate. To gain insight into mechanism whereby restricts retroviral replication, we investigated subcellular localization protein. Herein, report localizes to mRNA processing (P) bodies, cytoplasmic compartments involved in degradation storage nontranslating mRNAs. Biochemical analysis revealed...
T4 RNA ligases are commonly used to attach adapters RNAs, but large differences in ligation efficiency make detection and quantitation problematic. We developed a selection strategy using random RNAs combination with high-throughput sequencing gain insight into the of ligating pre-adenylated DNA 3′-ends. After analyzing biases sequence, secondary structure RNA-adapter cofold structure, we conclude that do not show significant primary sequence preference substrates, biased against structural...
High-throughput sequencing (HTS) has become a powerful tool for the detection of and sequence characterization microRNAs (miRNA) other small RNAs (sRNA). Unfortunately, use HTS data to determine relative quantity different miRNAs in sample been shown be inconsistent with quantitative PCR Northern Blot results. Several recent studies have concluded that major contributor this inconsistency is bias introduced during construction sRNA libraries primarily derived from adaptor ligation steps,...
A comprehensive analysis of the structure neuronal nitric oxide synthase (nNOS; EC 1.14.13.39 ) mRNA species revealed NOS1 to be most structurally diverse human gene described date in terms promoter usage. Nine unique exon 1 variants are variously used for transcript initiation tissues, and each is expressed from a 5′-flanking region. The dependence on genomic regions control transcription cell-specific fashion burdens transcripts with complex 5′-mRNA leader sequences. Elaborate splicing...
Endothelial nitric-oxide synthase (eNOS) mRNA levels are abnormal in diseases of the cardiovascular system, but changes gene expression cannot be accounted for by transcription alone. We found evidence existence an antisense (sONE) that is derived from a unit (NOS3AS) on opposite DNA strand which human eNOS (NOS3) transcribed at chromosome 7q36. The genes oriented tail-to-tail configuration, and mRNAs encoding sONE complementary 662 nucleotides. could detected variety cell types, both vivo...
Small regulatory RNA repertoires in biological samples are heterogeneous mixtures that may include species arising from varied biosynthetic pathways and modification events. profiling discovery approaches ought to capture molecules a way is representative of expression level. It follows the effects modifications on representation should be minimized. The collection high-quality, data, therefore, will highly dependent bias-free sample manipulation advance quantification. We examined impact...
Cas9 nuclease is the key effector of type II CRISPR adaptive immune systems found in bacteria. The can be programmed by a single guide RNA (sgRNA) to cleave DNA sequence-specific manner. This property has led its widespread adoption as genome editing tool research laboratories and holds great promise for biotechnological therapeutic applications. general mechanistic features catalysis homologs are comparable; however, high degree diversity exists among protein sequences, which may result...
T4 RNA ligases 1 and 2 are useful tools for analysis. Their use upstream of analyses such as high-throughput sequencing microarrays has recently increased their importance. The truncated form ligase 2, comprising amino acids 1-249 (T4 Rnl2tr), is an attractive tool attachment adapters or labels to 3'-ends. Compared 1, Rnl2tr a decreased ability ligate 5'-PO4 ends in single-stranded ligations, compared the full-length Rnl2, activity joining 5'-adenylated combination these properties allows...
Human endothelial nitric oxide synthase (eNOS) mRNA is highly stable in cells (ECs). Posttranscriptional regulation of eNOS stability an important component regulation, especially under hypoxic conditions. Here, we show that the human 3' untranslated region (3' UTR) contains multiple, evolutionarily conserved pyrimidine (C and CU)-rich sequence elements are both necessary sufficient for stabilization. Importantly, RNA immunoprecipitations electrophoretic mobility shift assays (EMSAs)...
ABSTRACT CRISPR-Cas12a (Cpf1) are RNA-guided nuclease effectors of acquired immune response that act in their native organisms by cleaving targeted DNA sequences. Like CRISPR-Cas9 targeting enzymes, Cas12a orthologs have been repurposed for genome editing non-native and manipulation vitro . Recent studies shown activation via guide RNA-target pairing causes multiple turnover, non-specific ssDNA degradation trans , after single turnover on-target cleavage cis We find the activity affects RNA...
Article21 October 2022Open Access Source DataTransparent process A new family of CRISPR-type V nucleases with C-rich PAM recognition Tomas Urbaitis CasZyme, Vilnius, Lithuania Institute Biotechnology, Vilnius University, Contribution: Conceptualization, Data curation, Formal analysis, Validation, Investigation, Visualization, Methodology, Writing - original draft, review & editing Search for more papers by this author Giedrius Gasiunas Corresponding Author [email protected]...
Endothelium-derived nitric oxide (NO) is primarily attributable to constitutive expression of the endothelial synthase (eNOS) gene. Although a more comprehensive understanding transcriptional regulation eNOS emerging with respect in vitro regulatory pathways, their relevance vivo warrants assessment. In this regard, promoter-reporter insertional transgenic murine lines were created containing 5,200 bp native promoter directing transcription nuclear-localized β-galactosidase. Examination...
Expression of the neuronal nitric-oxide synthase (nNOS) mRNA is subject to complex cell-specific transcriptional regulation, which mediated by alternative promoters. Unexpectedly, we identified a 89-nucleotide alternatively spliced exon located in 5'-untranslated region between 1 variants and common 2 that contains translational initiation codon. Alternative splicing events do not affect open reading frame are distinctly uncommon mammals; therefore, assessed its functional relevance....