The outbreak caused by the novel coronavirus SARS-CoV-2 has been declared a global health emergency. G-quadruplex structures in genomes have long been considered essential for regulating a number of biological processes in a plethora of organisms. We have analyzed and identified 25 four contiguous GG runs (G2NxG2NyG2NzG2) in the SARS-CoV-2 RNA genome, suggesting putative G-quadruplex-forming sequences (PQSs). Detailed analysis of SARS-CoV-2 PQSs revealed their locations in the open reading frames of ORF1 ab, spike (S), ORF3a, membrane (M) and nucleocapsid (N) genes. Identical PQSs were also found in the other members of the Coronaviridae family. The top-ranked PQSs at positions 13385 and 24268 were confirmed to form RNA G-quadruplex structures in vitro by multiple spectroscopic assays. Furthermore, their direct interactions with viral helicase (nsp13) were determined by microscale thermophoresis. Molecular docking model suggests that nsp13 distorts the G-quadruplex structure by allowing the guanine bases to be flipped away from the guanine quartet planes. Targeting viral helicase and G-quadruplex structure represents an attractive approach for potentially inhibiting the SARS-CoV-2 virus.

A 'binge' is defined by National Institute on Alcohol Abuse and Alcoholism as an excessive pattern of alcohol drinking that produces blood-alcohol levels (BALs) greater than 0.08 g% within a 2-h period and may or may not be associated with dependence. The purpose of this investigation was to explore the effects of several neuropharmacological agents in an animal model in which outbred rats voluntarily and orally self-administer pharmacologically meaningful alcohol doses that produce BALs ≥ 0.08 g% in daily limited access two-bottle choice and operant drinking sessions. Rats were trained to self-administer either 10% (w/v) alcohol solution sweetened with 'supersac' (3% glucose + 0.125% saccharin) or supersac alone versus water in a two-bottle choice or operant situation during 30-min daily sessions. Rats were then injected systemically with multiple doses of duloxetine, naltrexone, and the corticotropin-releasing factor antagonist, MPZP, in Latin-square designs. Alcohol binge drinkers reliably consumed amounts of alcohol sufficient to produce BALs ≥ 0.08 g%. Duloxetine dose-dependently suppressed two-bottle choice alcohol binge drinking and operant alcohol responding as well as operant supersac responding, but did not affect two-bottle choice supersac drinking. Naltrexone-suppressed alcohol binge drinking at very low doses and suppressed supersac drinking at moderate-to-high doses. MPZP did not affect alcohol or supersac consumption. Different profiles for drugs that suppress binge-like alcohol drinking compared with dependenceinduced drinking provide a heuristic foundation for future medications development.

The highly conserved cellular degradation pathway, macroautophagy, regulates the homeostasis of organelles and promotes the survival of T lymphocytes. Previous results indicate that Atg3-, Atg5-, or Pik3c3/Vps34-deficient T cells cannot proliferate efficiently. Here we demonstrate that the proliferation of Atg7-deficient T cells is defective. By using an adoptive transfer and Listeria monocytogenes (LM) mouse infection model, we found that the primary immune response against LM is intrinsically impaired in autophagy-deficient CD8 + T cells because the cell population cannot expand after infection. Autophagy-deficient T cells fail to enter into S-phase after TCR stimulation. The major negative regulator of the cell cycle in T lymphocytes, CDKN1B, is accumulated in autophagy-deficient na€ ıve T cells and CDKN1B cannot be degraded after TCR stimulation. Furthermore, our results indicate that genetic deletion of one allele of CDKN1B in autophagy-deficient T cells restores proliferative capability and the cells can enter into S-phase after TCR stimulation. Finally, we found that natural CDKN1B forms polymers and is physiologically associated with the autophagy receptor protein SQSTM1/p62 (sequestosome 1). Collectively, autophagy is required for maintaining the expression level of CDKN1B in na€ ıve T cells and selectively degrades CDKN1B after TCR stimulation.

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Guanine (G)-quadruplexes (G4s) are unique nucleic acid structures that are formed by stacked G-tetrads in G-rich DNA or RNA sequences. G4s have been reported to play significant roles in various cellular events in both macro- and micro-organisms. The identification and characterization of G4s can help to understand their different biological roles and potential applications in diagnosis and therapy. In addition to biophysical and biochemical methods to interrogate G4 formation, G4 fluorescent turn-on ligands can be used to target and visualize G4 formation both in vitro and in cells. Here, we review several representative classes of G4 fluorescent turn-on ligands in terms of their interaction mechanism and application perspectives. Interestingly, G4 structures are commonly identified in DNA and RNA aptamers against targets that include proteins and small molecules, which can be utilized as G4 tools for diverse applications. We therefore also summarize the recent development of G4-containing aptamers and highlight their applications in biosensing, bioimaging, and therapy. Moreover, we discuss the current challenges and future perspectives of G4 fluorescent turn-on ligands and G4-containing aptamers.

Studies from our laboratory indicated that local perfusion of the ventral tegmental area (VTA) with a serotonin-3 (5-HT 3 ) receptor agonist increased dopamine (DA) neuronal activity and that the self-infusion of ethanol (EtOH) and cocaine into the posterior VTA could be inhibited with coadministration of a 5-HT 3 receptor antagonist. The study tested the hypothesis that activating 5-HT 3 receptors within the VTA produces reinforcing effects. The study also examined whether there were differences between Wistar rats and a line of rats selectively bred for high alcohol consumption with regard to the self-infusion of a 5-HT 3 receptor agonist within the VTA. Adult female alcohol-preferring (P) and Wistar rats were allowed to self-infuse the 5-HT 3 receptor agonist 1-(m-chlorophenyl)-biguanide (CPBG) into the posterior or anterior VTA. Furthermore, experiments examined the effects of coinfusion of the 5-HT 3 antagonist ICS 205,930 (ICS), and the DA D 2,3 agonist quinpirole on the self-infusion of CPBG. Both Wistar and P rats readily self-administered CPBG into the posterior, but not anterior, VTA. P rats self-infused lower concentrations of CPBG (0.10 M) than did Wistar rats (1.0 M). Coinfusion of either ICS or quinpirole reduced CPBG self-infusion into the posterior VTA. The results of this study suggest that activation of 5-HT 3 receptors within the posterior VTA produces reinforcing effects and that these reinforcing effects are mediated through activation of DA neurons. Furthermore, the data suggest that selective breeding for alcoholpreference results in the posterior VTA being more sensitive to the reinforcing effects of CPBG.An in vivo microdialysis study (Campbell et al., 1996) demonstrated that local administration of a serotonin-3 (5-HT 3 ) receptor agonist increased somatodendritic dopamine (DA) release in the ventral tegmental area (VTA), suggesting that 5-HT 3 receptors are involved in regulating DA neuronal activity within the VTA. In support of this interpretation, electrophysiological studies indicated that systemic administration of 5-HT 3 receptor antagonists decreased the number of spontaneously active VTA DA neurons in rodents (Minabe et al., 1991;Rasmussen et al., 1991).The results of several studies suggested a role for the involvement of 5-HT 3 receptors in mediating the excitatory actions of ethanol (EtOH). EtOH can enhance 5-HT 3 receptor-mediated ion currents (Lovinger and White 1991); 5-HT 3 receptor antagonists inhibited the increase in extracellular DA levels in the nucleus accumbens elicited by EtOH when the antagonist was given systemically (Carboni et al., 1989;Wozniak et al., 1990) or locally (Campbell and McBride, 1995). Moreover, the in vivo microdialysis study of Campbell et al. (1996) indicated that local perfusion with a 5-HT 3 receptor antagonist blocked EtOH-stimulated somatodendritic DA release within the VTA, suggesting that the activating effects of EtOH on VTA DA neurons were mediated in part through 5-HT 3 receptors.The intracranial self-administration (ICSA) tec...

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