A5025520245- RNA and protein synthesis mechanisms
- RNA Research and Splicing
- RNA Interference and Gene Delivery
- Advanced biosensing and bioanalysis techniques
- MicroRNA in disease regulation
- RNA modifications and cancer
- Fungal and yeast genetics research
- Bacteriophages and microbial interactions
- Genomics and Chromatin Dynamics
- RNA regulation and disease
- Traditional Chinese Medicine Studies
- DNA and Nucleic Acid Chemistry
- Molecular Biology Techniques and Applications
- Traditional Chinese Medicine Analysis
- Flavonoids in Medical Research
- Protease and Inhibitor Mechanisms
- Chromatography in Natural Products
- Bacterial Genetics and Biotechnology
- Biochemical and Molecular Research
- Plant Virus Research Studies
- Phytochemistry and Biological Activities
- Plant-derived Lignans Synthesis and Bioactivity
- Signaling Pathways in Disease
- Adenosine and Purinergic Signaling
- Morinda citrifolia extract uses
Tianjin Agricultural University
2011-2024
Merck & Co., Inc., Rahway, NJ, USA (United States)
2010-2019
United States Military Academy
2010-2016
Kettering University
1998-2010
Shanxi Provincial Cancer Hospital
2009
Rockefeller University
2005-2007
Howard Hughes Medical Institute
2006-2007
Memorial Sloan Kettering Cancer Center
2006
Cornell University
1999
Nanjing University
1995-1996
RNA silencing refers to small regulatory RNA-mediated processes that repress endogenous gene expression and defend hosts from offending viruses. As an anti-host defense mechanism, viruses encode suppressors can block pathways. Cucumber mosaic virus (CMV)-encoded 2b protein was among the first identified could inhibit post-transcriptional (PTGS), but with little or no effect on miRNA functions. The mechanisms underlying suppression of are unknown. Here, we demonstrate CMV also interferes...
Small interfering RNAs (siRNAs) and microRNAs (miRNAs) guide catalytic sequence-specific cleavage of fully or nearly complementary target mRNAs control translation and/or stability many that share 6–8 nucleotides (nt) complementarity to the siRNA miRNA 5′ end. siRNA- miRNA-containing ribonucleoprotein silencing complexes are assembled from double-stranded 21- 23-nt RNase III processing intermediates carry phosphates 2-nt overhangs with free 3′ hydroxyl groups. Despite structural symmetry a...
Abstract Small interfering RNA (siRNA)‐mediated silencing requires siRNA loading into the RNA‐induced complex (RISC). Presence of 5′‐phosphate (5′‐P) is reported to be critical for efficient RISC antisense strand (AS) by anchoring it mid‐domain Argonaute2 (Ago2) protein. Phosphorylation exogenous duplex siRNAs thought accomplished cytosolic Clp1 kinase. However, although extensive chemical modifications are essential siRNA–GalNAc conjugate activity, they can significantly impair kinase...
Elongating RNA polymerase II is targeted by macromolecular assemblies that regulate mRNA synthesis and processing. The capping apparatus the first of to act on nascent pre-mRNA. Although recruitment enzymes transcription complex dependent phosphorylation C-terminal domain Rpb1 subunit (Pol-II), there may be additional levels control coordinate with elongation. Here we show triphosphatase (Pct1) guanylyltransferase (Pce1) fission yeast bind independently elongation factor Spt5. 990-amino acid...
This study investigated the effects of thermosonication (TS) on quality blackcurrant juice, along with its physicochemical properties, bioactive compounds, antioxidant capacity, and microbiological sensory qualities. The treatments included raw juice (RJ), pasteurized (90 °C, 1 min, PJ), thermosonicated (480 W, 40 kHz at 40, 50, or 60 for 10, 20, 30, TJ). results indicated that pasteurization pH, total soluble solids, titratable acidity were not significant (p > 0.05). However,...
Effective small interfering RNA (siRNA)–mediated therapeutics require the siRNA to be delivered into cellular RNA-induced silencing complex (RISC). Quantitative information of this essential delivery step is currently inferred from efficacy gene and uptake in tissue. Here we report an approach directly quantify RISC rodents monkey. This achieved by specific immunoprecipitation tissue lysates quantification RNAs immunoprecipitates stem–loop PCR. The method, expected independent vehicle...
An interaction network connecting mRNA capping enzymes, the RNA polymerase II (Pol II) carboxyl-terminal domain (CTD), elongation factor Spt5, and Cdk7 Cdk9 protein kinases is thought to comprise a transcription checkpoint. A crux of this which regulates early has an imputed role in pre-mRNA processing via its physical association with enzymes. Schizosaccharomyces pombe Spt5 distinctive CTD composed tandem nonapeptide repeats consensus sequence 1TPAWNSGSK9. The binds enzymes substrate for...
Effective delivery of siRNA (small interfering RNA) into the cells requires translocation cytosol. One potential strategy uses cell-delivery peptides that facilitate this step. In present paper, we describe characterization an amphipathic peptide mediates uptake non-covalently bound and its subsequent release Biophysical peptide/siRNA mixtures at neutral lysosomal (acidic) pH suggested formation α-helical structure only in endosomes lysosomes. Surprisingly, even though enhanced cells, no...
Deeper knowledge of pharmacokinetic and pharmacodynamic (PK/PD) concepts for RNA therapeutics is important to streamline the drug development process rigorous selection best performing candidates. Here we characterized PK/PD relationship small interfering RNAs (siRNAs) targeting luciferase by examining siRNA concentration in plasma liver, temporal RNA-induced silencing complex binding profiles, mRNA reduction, protein inhibition measured noninvasive bioluminescent imaging. A dose-dependent...
<i>Saccharomyces cerevisiae</i> Cet1p catalyzes the first step of mRNA capping, hydrolysis γ phosphate triphosphate-terminated RNA to form a 5′ diphosphate end. The triphosphatase activity is magnesium-dependent and has turnover number 1 s<sup>−1</sup>. Here we show that purified recombinant possesses robust ATPase (<i>K</i> <sub>m</sub> = 2.8 μm; <i>V</i> <sub>max</sub> 25 s<sup>−1</sup>) in presence manganese. Cobalt also an effective cofactor, but magnesium, calcium, copper, zinc are not....
RNA polymerase II (pol II) is subject to an early elongation delay induced by negative factors Spt5/Spt4 and NELF, which overcome the positive factor P-TEFb (Cdk9/cyclin T), a protein kinase that phosphorylates pol C-terminal domain (CTD) transcription Spt5. Although rationale for this arrest restart unclear, recent studies suggest connection mRNA capping, coupled via physical functional interactions between cap-forming enzymes, CTD-PO<sub>4</sub>, Here we identify novel interaction fission...
The carboxyl-terminal domain (CTD) of elongating RNA polymerase II serves as a landing pad for macromolecular assemblies that regulate mRNA synthesis and processing. capping apparatus is the first to act on nascent pre-mRNA one which binding catalytic components most clearly dependent CTD phosphorylation. present study highlights distinctive strategy cap targeting in fission yeast whereby triphosphatase (Pct1) guanylyltransferase (Pce1) enzymes do not interact physically with each other (as...
Schizosaccharomyces pombe Cdk9/Pch1 protein kinase is a functional ortholog of the essential Saccharomyces cerevisiae Bur1/Bur2 and putative metazoan P-TEFb (Cdk9/cyclin T). SpCdk9/Pch1 phosphorylates carboxyl-terminal domain (CTD) S. transcription elongation factor Spt5, which consists 18 tandem repeats nonapeptide consensus sequence 1TPAWNSGSK9. We document divalent cation dependence specificity SpCdk9/Pch1, its NTP specificity, Spt5-CTD phosphorylation on number nonamer repeats, for...
Cyclin-dependent kinase 9 (Cdk9) of fission yeast is an essential ortholog metazoan positive transcription elongation factor b (P-TEFb), which proposed to coordinate capping and RNA polymerase II (Pol II) transcripts. Here we show that Cdk9 activated phosphorylate Pol the Spt5 by Csk1, one two CDK-activating kinases (CAKs). Activation depends on T-loop residue Thr-212. The other CAK-Mcs6, component IIH (TFIIH)-cannot activate Cdk9. Consistent with specificities CAKs in vitro, activity...
T4 RNA ligase 1 (Rnl1) exemplifies an ATP-dependent family that includes fungal tRNA (Trl1) and a putative baculovirus ligase. Rnl1 acts via covalent enzyme-AMP intermediate generated by attack of Lys-99 Nζ on the α phosphorus ATP. Mutation abolishes activity. Here we tested effects alanine mutations at 19 conserved positions in thereby identified 9 new residues essential for activity: Arg-54, Lys-75, Phe-77, Gly-102, Lys-119, Glu-227, Gly-228, Lys-240, Lys-242. Seven are located within...
A primary consideration when developing lipid nanoparticle (LNP) based small interfering RNA (siRNA) therapeutics is formulation polydispersity or heterogeneity. The level of heterogeneity physicochemical properties within a pharmaceutical batch could greatly affect the bioperformance, quality, and ability manufacturer to consistently control reproduce formulations. This article studied in size, composition, vitro performance siRNA containing LNPs, by conducting preparative scale...
RNA triphosphatase catalyzes the first step in mRNA cap formation which entails cleavage of β–γ phosphoanhydride bond triphosphate-terminated to yield a diphosphate end that is then capped with GMP by guanylyltransferase. Here we characterize 303 amino acid (Pct1p) encoded fission yeast Schizosaccharomycespombe. Pct1p hydrolyzes γ phosphate poly(A) presence magnesium. also ATP ADP and Pi manganese or cobalt (Km = 19 µM ATP; kcat 67 s–1). Hydrolysis 1 mM inhibited increasing potency inorganic...
<i>Saccharomyces cerevisiae</i> Cet1p is the prototype of a family metal-dependent RNA 5′-triphosphatases/NTPases encoded by fungi and DNA viruses; defined conserved sequence motifs A, B, C. We tested effects 12 alanine substitutions 16 conservative modifications at 18 positions motifs. Eight residues were identified as important for triphosphatase activity. These Glu-305, Glu-307, Phe-310 in motif A (IELEMKF); Arg-454 Lys-456 B (RTK); Glu-492, Glu-494, Glu-496 C (EVELE). Four acidic...
The RNA triphosphatase component (CaCet1p) of the mRNA capping apparatus pathogenic fungus Candida albicans differs mechanistically and structurally from mammals. Hence, CaCet1p is an attractive antifungal target. Here we identify a C-terminal catalytic domain residue 257 to 520 characterize manganese-dependent cobalt-dependent NTPase activity intrinsic CaCet1p. can be exploited screen in vitro for inhibitors. amino acids that comprise active site were identified by alanine-scanning...
The Spt5-Spt4 complex regulates early transcription elongation by RNA polymerase II and has an imputed role in pre-mRNA processing via its physical association with mRNA capping enzymes. Here we characterize the Schizosaccharomyces pombe core as a heterodimer map trypsin-resistant Spt4-binding domain within Spt5 subunit. A genetic analysis of Spt4 S. revealed it to be inessential for growth at 25°C–30°C but critical 37°C. These results echo conditional spt4 Δ phenotype budding yeast, where...