Host poly(A) polymerases PAPD5 and PAPD7 provide two layers of protection that ensure the integrity and stability of hepatitis B virus RNA

ABSTRACT To determine the influence of posttranscriptional modifications on 3′ end processing and RNA stability in plant mitochondria, peaatp9 and Oenothera atp1 transcripts were investigated for the presence and function of 3′ nonencoded nucleotides. A 3′ rapid amplification of cDNA ends approach initiated at oligo(dT)-adapter primers finds the expected poly(A) tails predominantly attached within the second stem or downstream of the double stem-loop structures at sites of previously mapped 3′ ends. Functional studies in a pea mitochondrial in vitro processing system reveal a rapid removal of the poly(A) tails up to termini at the stem-loop structure but little if any influence on further degradation of the RNA. In contrast 3′ poly(A) tracts at RNAs without such stem-loop structures significantly promote total degradation in vitro. To determine the in vivo identity of 3′ nonencoded nucleotides more accurately, pea atp9 transcripts were analyzed by a direct anchor primer ligation-reverse transcriptase PCR approach. This analysis identified maximally 3-nucleotide-long nonencoded extensions most frequently of adenosines combined with cytidines. Processing assays with substrates containing homopolymer stretches of different lengths showed that 10 or more adenosines accelerate RNA processivity, while 3 adenosines have no impact on RNA life span. Thus polyadenylation can generally stimulate the decay of RNAs, but processivity of degradation is almost annihilated by the stabilizing effect of the stem-loop structures. These antagonistic actions thus result in the efficient formation of 3′ processed and stable transcripts.

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Pudilan xiaoyan oral liquid (PDL) inhibit the replication of influenza A virus through regulating TLR3/MyD88 signaling pathway

10.21203/rs.3.rs-1205788/v1 ◽

2022 ◽

Author(s):

Zheng Zhihui ◽

Yuqian Zhang ◽

Gang Tian ◽

Zehua Wang ◽

Ronghua Wang ◽

...

Keyword(s):

Influenza Virus ◽

Influenza A Virus ◽

Influenza A ◽

Antiviral Effect ◽

Inhibitory Effect ◽

Tnf Α ◽

Oral Liquid ◽

Ifn Γ

Abstract Background Pudilan Xiaoyan Oral Liquid (PDL) as a famous Chinese patent medicine has been widely used for treating upper respiratory tract infection. However, the antiviral effect of PDL remain unclear. Here, the antiviral effect of in vitro and in vivo of PDL against influenza A virus were for the first time investigated. Methods The in vitro inhibitory effect of PDL on influenza A virus was investigated using MDCK cell model. The in vivo inhibitory effect on influenza virus pneumonia was evaluated with the ICR female mice (14-16 g) model infected by influenza A virus (A/FM/1/47, H1N1, mouse-adapted). Moreover, expression levels of inflammatory cytokines including TNF-α, IP10, IL-10, IL-1β, IL-6 and IFN-γ in lung tissue were measured by qRT-PCR. The potential mechanism of PDL against acute lung injury caused by influenza A virus was investigated by RT-PCR and Western blot. Results Our results indicated that in vitro PDL has a broad-spectrum inhibitory effect on different subtypes of influenza A viruses and in vivo PDL could dose-dependently prevent weight loss of mice, increase food intake and reduce mortality caused by influenza A H1N1 virus. Furthermore, PDL could markedly improve the acute lung injury caused by influenza A virus and significantly reduce the mRNA levels of inflammatory factors such as TNF-α, IP10, IL-10, IL-1β, IL-6, and IFN-γ. Mechanistic research indicated that the protective effect of PDL on viral pneumonia might be achieved by inhibiting TLR3/MyD88/IRAK4/TRAF3 signaling pathway. Conclusion PDL not only showed a good inhibitory effect on influenza A virus in vitro, but also exhibited a significant protective effect against lethal influenza virus infection in vivo. These findings provide evidence for the clinical treatment of influenza A virus infection with PDL.

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Relative Expression Levels Rather Than Specific Activity Plays the Major Role in Determining In Vivo AKT Isoform Substrate Specificity

Enzyme Research ◽

10.4061/2011/720985 ◽

2011 ◽

Vol 2011 ◽

pp. 1-18 ◽

Cited By ~ 8

Author(s):

Rachel S. Lee ◽

Colin M. House ◽

Briony E. Cristiano ◽

Ross D. Hannan ◽

Richard B. Pearson ◽

...

Keyword(s):

Substrate Specificity ◽

Specific Activity ◽

Dominant Role ◽

Protein Substrates ◽

Cellular Processes ◽

Disease States ◽

The Individual

The AKT protooncogene mediates many cellular processes involved in normal development and disease states such as cancer. The three structurally similar isoforms: AKT1, AKT2, and AKT3 exhibit both functional redundancy and isoform-specific functions; however the basis for their differential signalling remains unclear. Here we show that in vitro, purified AKT3 is ∼47-fold more active than AKT1 at phosphorylating peptide and protein substrates. Despite these marked variations in specific activity between the individual isoforms, a comprehensive analysis of phosphorylation of validated AKT substrates indicated only subtle differences in signalling via individual isoforms in vivo. Therefore, we hypothesise, at least in this model system, that relative tissue/cellular abundance, rather than specific activity, plays the dominant role in determining AKT substrate specificity in situ.

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Small cis-Acting Sequences That Specify Secondary Structures in a Chloroplast mRNA Are Essential for RNA Stability and Translation

Molecular and Cellular Biology ◽

10.1128/mcb.19.12.8479 ◽

1999 ◽

Vol 19 (12) ◽

pp. 8479-8491 ◽

Cited By ~ 45

Author(s):

David C. Higgs ◽

Risa S. Shapiro ◽

Karen L. Kindle ◽

David B. Stern

Keyword(s):

Untranslated Region ◽

Rna Stability ◽

Dimethyl Sulfate ◽

Secondary Structures ◽

Rna Modification ◽

Range Interaction ◽

Stem Loop ◽

Chloroplast Mrna

ABSTRACT Nucleus-encoded proteins interact with cis-acting elements in chloroplast transcripts to promote RNA stability and translation. We have analyzed the structure and function of three such elements within the Chlamydomonas petD 5′ untranslated region; petD encodes subunit IV of the cytochromeb 6/f complex. These elements were delineated by linker-scanning mutagenesis, and RNA secondary structures were investigated by mapping nuclease-sensitive sites in vitro and by in vivo dimethyl sulfate RNA modification. Element I spans a maximum of 8 nucleotides (nt) at the 5′ end of the mRNA; it is essential for RNA stability and plays a role in translation. This element appears to form a small stem-loop that may interact with a previously described nucleus-encoded factor to block 5′→3′ exoribonucleolytic degradation. Elements II and III, located in the center and near the 3′ end of the 5′ untranslated region, respectively, are essential for translation, but mutations in these elements do not affect mRNA stability. Element II is a maximum of 16 nt in length, does not form an obvious secondary structure, and appears to bind proteins that protect it from dimethyl sulfate modification. Element III spans a maximum of 14 nt and appears to form a stem-loop in vivo, based on dimethyl sulfate modification and the sequences of intragenic suppressors of element III mutations. Furthermore, mutations in element II result in changes in the RNA structure near element III, consistent with a long-range interaction that may promote translation.

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Host poly(A) polymerases PAPD5 and PAPD7 provide two layers of protection that ensure the integrity and stability of hepatitis B virus RNA

Journal of Virology ◽

10.1128/jvi.00574-21 ◽

2021 ◽

Author(s):

Fei Liu ◽

Amy C. H. Lee ◽

Fang Guo ◽

Andrew S. Kondratowicz ◽

Holly M. Micolochick Steuer ◽

...

Keyword(s):

Hepatitis B ◽

Small Molecule ◽

Dominant Role ◽

Regulatory Element ◽

Rna Metabolism ◽

Host Factors ◽

Health Concern ◽

Genetic Studies

Noncanonical poly(A) polymerases PAPD5 and PAPD7 (PAPD5/7) stabilize HBV RNA via the interaction with the viral post-transcriptional regulatory element (PRE), representing new antiviral targets to control HBV RNA metabolism, HBsAg production and viral replication. Inhibitors targeting these proteins are being developed as antiviral therapies, therefore it is important to understand how PAPD5/7 coordinate to stabilize HBV RNA. Here, we utilized a potent small-molecule AB-452 as a chemical probe, along with genetic analyses to dissect the individual roles of PAPD5/7 in HBV RNA stability. AB-452 inhibits PAPD5/7 enzymatic activities and reduces HBsAg both in vitro (EC 50 ranged from 1.4 to 6.8 nM) and in vivo by 0.93 log 10 . Our genetic studies demonstrate that the stem-loop alpha sequence within PRE is essential for both maintaining HBV poly(A) tail integrity and determining sensitivity towards the inhibitory effect of AB-452. Although neither single knock-out (KO) of PAPD5 nor PAPD7 reduces HBsAg RNA and protein production, PAPD5 KO does impair poly(A) tail integrity and confers partial resistance to AB-452. In contrast, PAPD7 KO did not result in any measurable changes within the HBV poly(A) tails, but cells with both PAPD5 and PAPD7 KO show reduced HBsAg production and conferred complete resistance to AB-452 treatment. Our results indicate that PAPD5 plays a dominant role in stabilizing viral RNA by protecting the integrity of its poly(A) tail, while PAPD7 serves as a second line of protection. These findings inform PAPD5 targeted therapeutic strategies and open avenues for further investigating PAPD5/7 in HBV replication. Importance Chronic hepatitis B affects more than 250 million patients and is a major public health concern worldwide. HBsAg plays a central role in maintaining HBV persistence and as such, therapies that aim at reducing HBsAg through destabilizing or degrading HBV RNA have been extensively investigated. Besides directly degrading HBV transcripts through antisense oligonucleotides or RNA silencing technologies, small-molecule compounds targeting host factors such as the noncanonical poly(A) polymerase PAPD5 and PAPD7 have been reported to interfere with HBV RNA metabolism. Herein, our antiviral and genetic studies using relevant HBV infection and replication models further characterize the interplays between the cis-element within the viral sequence and the trans-elements from the host factors. PAPD5/7 targeting inhibitors, with oral bioavailability, thus represent an opportunity to reduce HBsAg through destabilizing HBV RNA.

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Opposing roles for Egalitarian and Staufen in transport, anchoring and localization of oskar mRNA in the Drosophila oocyte

PLoS Genetics ◽

10.1371/journal.pgen.1009500 ◽

2021 ◽

Vol 17 (4) ◽

pp. e1009500

Author(s):

Sabine Mohr ◽

Andrew Kenny ◽

Simon T. Y. Lam ◽

Miles B. Morgan ◽

Craig A. Smibert ◽

...

Keyword(s):

Binding Sites ◽

Inhibitory Effect ◽

Multiple Roles ◽

Rna Transport ◽

Nurse Cells ◽

Stem Loop ◽

Stem Loop Structure ◽

Dependent Inhibition

Localization of oskar mRNA includes two distinct phases: transport from nurse cells to the oocyte, a process typically accompanied by cortical anchoring in the oocyte, followed by posterior localization within the oocyte. Signals within the oskar 3’ UTR directing transport are individually weak, a feature previously hypothesized to facilitate exchange between the different localization machineries. We show that alteration of the SL2a stem-loop structure containing the oskar transport and anchoring signal (TAS) removes an inhibitory effect such that in vitro binding by the RNA transport factor, Egalitarian, is elevated as is in vivo transport from the nurse cells into the oocyte. Cortical anchoring within the oocyte is also enhanced, interfering with posterior localization. We also show that mutation of Staufen recognized structures (SRSs), predicted binding sites for Staufen, disrupts posterior localization of oskar mRNA just as in staufen mutants. Two SRSs in SL2a, one overlapping the Egalitarian binding site, are inferred to mediate Staufen-dependent inhibition of TAS anchoring activity, thereby promoting posterior localization. The other three SRSs in the oskar 3’ UTR are also required for posterior localization, including two located distant from any known transport signal. Staufen, thus, plays multiple roles in localization of oskar mRNA.

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Biologically Active Constituents of a Polyphenol Extract from Geranium sanguineum L. with Anti-Influenza Activity

Zeitschrift für Naturforschung C ◽

10.1515/znc-2006-7-807 ◽

2006 ◽

Vol 61 (7-8) ◽

pp. 508-516 ◽

Cited By ~ 17

Author(s):

Assen Pantev ◽

Stefka Ivancheva ◽

Lidija Staneva ◽

Julia Serkedjieva

Keyword(s):

Antiviral Effect ◽

Influenza Viruses ◽

Biologically Active ◽

Chemical Components ◽

Aerial Roots ◽

Influenza Activity ◽

Etoac Fraction ◽

The Individual

From the aerial roots of the medicinal plant Geranium sanguineum L. a polyphenol-rich extract with strong anti-influenza activity has been isolated. To investigate its active fractions, the extract was partitioned by solvents with increasing polarity. The n-BuOH fraction contained the majority of the in vitro antiviral activity; the EtOAc fraction was the most effective one in vivo. A bioassay-directed fractionation of the n-BuOH and EtOAc fractions was performed to obtain information about the nature of the chemical components of the plant extract, responsible for the antiviral effect. The individual constituents were identified by spectroscopic methods and comparison with authentic samples and by HPLC. The cell-toxic and virus-inhibitory effects of the fractions and some individual polyphenol compounds, found in Geranium sanguineum L., were studied using the replication of representative influenza viruses in cell cultures. This study showed that the presence of a variety of biologically active compounds as well as the possible synergistic interactions between them seem to be decisive for the overall antiviral effect.

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The Effect of Barbituric Acid Derivatives on Platelet Function in Vitro and in Vivo

Thrombosis and Haemostasis ◽

10.1055/s-0038-1649080 ◽

1973 ◽

Vol 30 (02) ◽

pp. 315-326

Author(s):

J. Heinz Joist ◽

Jean-Pierre Cazenave ◽

J. Fraser Mustard

Keyword(s):

Platelet Aggregation ◽

Platelet Function ◽

Barbituric Acid ◽

Platelet Rich Plasma ◽

Inhibitory Effect ◽

Primary Response ◽

Sodium Pentobarbital ◽

Barbituric Acid Derivatives

SummarySodium pentobarbital (SPB) and three other barbituric acid derivatives were found to inhibit platelet function in vitro. SPB had no effect on the primary response to ADP of platelets in platelet-rich plasma (PRP) or washed platelets but inhibited secondary aggregation induced by ADP in human PRP. The drug inhibited both phases of aggregation induced by epinephrine. SPB suppressed aggregation and the release reaction induced by collagen or low concentrations of thrombin, and platelet adherence to collagen-coated glass tubes. The inhibition by SPB of platelet aggregation was readily reversible and isotopically labeled SPB did not become firmly bound to platelets. No inhibitory effect on platelet aggregation induced by ADP, collagen, or thrombin could be detected in PRP obtained from rabbits after induction of SPB-anesthesia.

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Inhibition of Human and Animal Platelet Adhesiveness to Glass Bead Columns by Adenosine, Dipyridamole, Chlorpromazine and Acetylsalicylic Acid

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648055 ◽

1976 ◽

Vol 36 (02) ◽

pp. 401-410 ◽

Cited By ~ 6

Author(s):

Buichi Fujttani ◽

Toshimichi Tsuboi ◽

Kazuko Takeno ◽

Kouichi Yoshida ◽

Masanao Shimizu

Keyword(s):

Guinea Pig ◽

Acetylsalicylic Acid ◽

Guinea Pigs ◽

Glass Bead ◽

Animal Species ◽

Inhibitory Effect ◽

Rabbit Platelet ◽

Platelet Adhesiveness

SummaryThe differences among human, rabbit and guinea-pig platelet adhesiveness as for inhibitions by adenosine, dipyridamole, chlorpromazine and acetylsalicylic acid are described, and the influence of measurement conditions on platelet adhesiveness is also reported. Platelet adhesiveness of human and animal species decreased with an increase of heparin concentrations and an increase of flow rate of blood passing through a glass bead column. Human and rabbit platelet adhesiveness was inhibited in vitro by adenosine, dipyridamole and chlorpromazine, but not by acetylsalicylic acid. On the other hand, guinea-pig platelet adhesiveness was inhibited by the four drugs including acetylsalicylic acid. In in vivo study, adenosine, dipyridamole and chlorpromazine inhibited platelet adhesiveness in rabbits and guinea-pigs. Acetylsalicylic acid showed the inhibitory effect in guinea-pigs, but not in rabbits.

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Phenolic Acids Exert Anticholinesterase and Cognition-Improving Effects

Current Alzheimer Research ◽

10.2174/1567205014666171128102557 ◽

2018 ◽

Vol 15 (6) ◽

pp. 531-543 ◽

Cited By ~ 4

Author(s):

Dominik Szwajgier ◽

Ewa Baranowska-Wojcik ◽

Kamila Borowiec

Keyword(s):

Phenolic Acids ◽

Ex Vivo ◽

Amyloid Β ◽

Inhibitory Effect ◽

In Vivo Studies ◽

Amyloid Β Peptide ◽

Beneficial Effects ◽

Aβ Fibrils

Numerous authors have provided evidence regarding the beneficial effects of phenolic acids and their derivatives against Alzheimer's disease (AD). In this review, the role of phenolic acids as inhibitors of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) is discussed, including the structure-activity relationship. In addition, the inhibitory effect of phenolic acids on the formation of amyloid β-peptide (Aβ) fibrils is presented. We also cover the in vitro, ex vivo, and in vivo studies concerning the prevention and treatment of the cognitive enhancement.

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