Role of the Ribonuclease ONCONASE in miRNA Biogenesis and tRNA Processing: Focus on Cancer and Viral Infections

The majority of transcribed RNAs do not codify for proteins, nevertheless they display crucial regulatory functions by affecting the cellular protein expression profile. MicroRNAs (miRNAs) and transfer RNA-derived small RNAs (tsRNAs) are effectors of interfering mechanisms, so that their biogenesis is a tightly regulated process. Onconase (ONC) is an amphibian ribonuclease known for cytotoxicity against tumors and antiviral activity. Additionally, ONC administration in patients resulted in clinical effectiveness and in a well-tolerated feature, at least for lung carcinoma and malignant mesothelioma. Moreover, the ONC therapeutic effects are actually potentiated by cotreatment with many conventional antitumor drugs. This review not only aims to describe the ONC activity occurring either in different tumors or in viral infections but also to analyze the molecular mechanisms underlying ONC pleiotropic and cellular-specific effects. In cancer, data suggest that ONC affects malignant phenotypes by generating tRNA fragments and miRNAs able to downregulate oncogenes expression and upregulate tumor-suppressor proteins. In cells infected by viruses, ONC hampers viral spread by digesting the primer tRNAs necessary for viral DNA replication. In this scenario, new therapeutic tools might be developed by exploiting the action of ONC-elicited RNA derivatives

Asthma and poly(ADP-ribose) polymerase inhibition: a new therapeutic approach

Raffaela Zaffini, Giovanni Gotte, Marta Menegazzi Department of Neuroscience, Biomedicine and Movement Science, Biochemistry Section, University of Verona, Verona, Italy Abstract: Asthma is a chronic lung disease affecting people of all ages worldwide, and it frequently begins in childhood. Because of its chronic nature, it is characterized by pathological manifestations, including airway inflammation, remodeling, and goblet cell hyperplasia. Current therapies for asthma, including corticosteroids and beta-2 adrenergic agonists, are directed toward relieving the symptoms of the asthmatic response, with poor effectiveness against the underlying causes of the disease. Asthma initiation and progression depends on the T helper (Th) 2 type immune response carried out by a complex interplay of cytokines, such as interleukin (IL) 4, IL5, and IL13, and the signal transducer and activator of transcription 6. Much of the data resulting from different laboratories support the role of poly(ADP-ribose) polymerase (PARP) 1 and PARP14 activation in asthma. Indeed, PARP enzymes play key roles in the regulation and progression of the inflammatory asthma process because they affect the expression of genes and chemokines involved in the immune response. Consistently, PARP inhibition achievable either upon genetic ablation or by using pharmacological agents has shown a range of therapeutic effects against the disease. Indeed, in the last two decades, several preclinical studies highlighted the protective effects of PARP inhibition in various animal models of asthma. PARP inhibitors showed the ability to reduce the overall lung inflammation acting with a specific effect on immune cell recruitment and through the modulation of asthma-associated cytokines production. PARP inhibition has been shown to affect the Th1–Th2 balance and, at least in some aspects, the airway remodeling. In this review, we summarize and discuss the steps that led PARP inhibition to become a possible future therapeutic strategy against allergic asthma. Keywords: allergic airway disease, PARP1, PARP14, remodeling, STAT6, Th1–Th2 balance, Th2 respons

Protective role of st. John’s wort and its components hyperforin and hypericin against diabetes through inhibition of inflammatory signaling: Evidence from in vitro and in vivo studies

Diabetes mellitus is a very common chronic disease with progressively increasing prevalence. Besides the well-known autoimmune and inflammatory pathogenesis of type 1 diabetes, in many people, metabolic changes and inappropriate lifestyle favor a subtle chronic inflammatory state that contributes to development of insulin resistance and progressive loss of β-cell function and mass, eventually resulting in metabolic syndrome or overt type 2 diabetes. In this paper, we review the anti-inflammatory effects of the extract of Hypericum perforatum L. (St. John’s wort, SJW) and its main active ingredients firstly in representative pathological situations on inflammatory basis and then in pancreatic β cells and in obese or diabetic animal models. The simultaneous and long-lasting inhibition of signal transducer and activator of transcription (STAT)-1, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and mitogen-activated protein kinases (MAPKs)/c-jun N-terminal kinase (JNK) signaling pathways involved in pro-inflammatory cytokine-induced β-cell dysfunction/death and insulin resistance make SJW particularly suitable for both preventive and therapeutic use in metabolic diseases. Hindrance of inflammatory cytokine signaling is likely dependent on the hyperforin content of SJW extract, but recent data reveal that hypericin can also exert relevant protective effects, mediated by activation of the cyclic adenosine monophosphate (cAMP)/protein kinase cAMP-dependent (PKA)/adenosine monophosphate activated protein kinase (AMPK) pathway, against high-fat-diet-induced metabolic abnormalities. Actually, the mechanisms of action of the two main components of SJW appear complementary, strengthening the efficacy of the plant extract. Careful quantitative analysis of SJW components and suitable dosage, with monitoring of possible drug–drug interaction in a context of remarkable tolerability, are easily achievable pre-requisites for forthcoming clinical applications

Signal transducer and activator of transcription-1 localizes to the mitochondria and modulates mitophagy

The signal transducer and activator of transcription (STAT) proteins are latent transcription factors that have been shown to be involved in cell proliferation, development, apoptosis, and autophagy. STAT proteins undergo activation by phosphorylation at tyrosine 701 and serine 727 where they translocate to the nucleus to regulate gene expression. STAT1 has been shown to be involved in promoting apoptotic cell death in response to cardiac ischemia/reperfusion and has recently been shown by our laboratory to be involved in negatively regulating autophagy. These processes are thought to promote cell death and restrict cell survival leading to the generation of an infarct. Here we present data that shows STAT1 localizes to the mitochondria and co-immunoprecipitates with LC3. Furthermore, electron microscopy studies also reveal mitochondria from ex vivo I/R treated hearts of STAT1KO mice contained within a double membrane autophagosome indicating that STAT1 may be involved in negatively regulating mitophagy. This is the first description of STAT1 being localized to the mitochondria and also having a role in mitophagy

Rapid inactivation of NOS-I by lipopolysaccharide plus interferon-gamma-induced tyrosine phosphorylation.

Human astrocytoma T67 cells constitutively express a neuronal NO synthase (NOS-I) and, following administration of lipopolysaccharide (LPS) plus interferon-gamma (IFNgamma), an inducible NOS isoform (NOS-II). Previous results indicated that a treatment of T67 cells with the combination of LPS plus IFNgamma, by affecting NOS-I activity, also inhibited NO production in a very short time. Here, we report that under basal conditions, a NOS-I protein of about 150 kDa was weakly and partially tyrosine-phosphorylated, as verified by immunoprecipitation and Western blotting. Furthermore, LPS plus IFNgamma increased the tyrosine phosphorylation of NOS-I, with a concomitant inhibition of its enzyme activity. The same effect was observed in the presence of vanadate, an inhibitor of phosphotyrosine-specific phosphatases. On the contrary, genistein, an inhibitor of protein-tyrosine kinases, reduced tyrosine phosphorylation of NOS-I, enhancing its enzyme activity. Finally, using reverse transcriptase-polymerase chain reaction, we have observed that a suboptimal induction of NOS-II mRNA expression in T67 cells was enhanced by vanadate (or L-NAME) and inhibited by genistein. Because exogenous NO has been found to suppress NOS-II expression, the decrease of NO production that we have obtained from the inactivation of NOS-I by LPS/IFNgamma-induced tyrosine phosphorylation provides the best conditions for NOS-II expression in human astrocytoma T67 cells

The Stimulation of Inducible Nitric-oxide Synthase by the Prion Protein Fragment 106–126 in Human Microglia Is Tumor Necrosis Factor-α-dependent and Involves p38 Mitogen-activated Protein Kinase

A synthetic peptide consisting of amino acid residues 106-126 of the human prion protein (PrP-(106--126)) has been previously demonstrated to be neurotoxic and to induce microglial activation. The present study investigated the expression of the inducible form of the nitric-oxide synthase (NOS-II) in human microglial cells treated with PrP-(106--126). Using reverse transcriptase-polymerase chain reaction, we found that PrP-(106--126) induces NOS-II gene expression after 24 h of treatment and that this effect is accompanied by a peak of nuclear factor kappa B (NF-kappa B) binding at 30 min as evaluated by electrophoretic mobility shift assay. Since our previous data demonstrated tumor necrosis factor-alpha (TNF-alpha) to be a potent inducer of NOS-II in these cells, we analyzed the expression of this cytokine in PrP-(106--126)-treated microglia. PrP-(106--126) caused the release of TNF-alpha as detected by enzyme-linked immunosorbent assay, and a blocking antibody, anti-TNF-alpha, abolished NOS-II induction elicited by this peptide. Moreover, PrP-(106-126) activates p38 mitogen-activated protein kinase, and the inhibition of this pathway determines the ablation of NF-kappa B binding induced by this fragment peptide

Direct interation of garcinol and related polyisoprenylated benzophenones of Garcinia cambogia fruits with the transcription factor STAT-1 as a likely mechanism of their inhibitory effect on cytokine signaling pathways.

Garcinol (1), a polyisoprenylated benzophenone occurring in Garcinia species, has been reported to exert anti-inflammatory activity in LPS-stimulated macrophages, through inhibition of NF-κB and/or JAK/STAT-1 activation. In order to provide deeper insight into its effects on the cytokine signaling pathway and to clarify the underlying molecular mechanisms, 1 was isolated from the fruits of Garcinia cambogia along with two other polyisoprenylated benzophenones, guttiferones K (2) and guttiferone M (3), differing from each other in their isoprenyl moieties and their positions on the benzophenone core. The affinities of 1-3 for the STAT-1 protein have been evaluated by surface plasmon resonance and molecular docking studies and resulted in KD values in the micromolar range. Consistent with the observed high affinity toward the STAT-1 protein, garcinol and guttiferones K and M were able to modulate cytokine signaling in different cultured cell lines, mainly by inhibiting STAT-1 nuclear transfer and DNA binding, as assessed by an electrophorectic mobility shift assay

The genetic basis of diurnal preference in Drosophila melanogaster.

BACKGROUND: Most animals restrict their activity to a specific part of the day, being diurnal, nocturnal or crepuscular. The genetic basis underlying diurnal preference is largely unknown. Under laboratory conditions, Drosophila melanogaster is crepuscular, showing a bi-modal activity profile. However, a survey of strains derived from wild populations indicated that high variability among individuals exists, including flies that are nocturnal. RESULTS: Using a highly diverse population, we performed an artificial selection experiment, selecting flies with extreme diurnal or nocturnal preference. After 10 generations, we obtained highly diurnal and nocturnal strains. We used whole-genome expression analysis to identify differentially expressed genes in diurnal, nocturnal and crepuscular (control) flies. Other than one circadian clock gene (pdp1), most differentially expressed genes were associated with either clock output (pdf, to) or input (Rh3, Rh2, msn). This finding was congruent with behavioural experiments indicating that both light masking and the circadian pacemaker are involved in driving nocturnality. CONCLUSIONS: Our study demonstrates that genetic variation segregating in wild populations contributes to substantial variation in diurnal preference. We identified candidate genes associated with diurnality/nocturnality, while data emerging from our expression analysis and behavioural experiments suggest that both clock and clock-independent pathways are involved in shaping diurnal preference. The diurnal and nocturnal selection strains provide us with a unique opportunity to understand the genetic architecture of diurnal preference