The subgenomic flaviviral RNA suppresses RNA interference through competing with siRNAs for binding RISC components.

During the life cycle of mosquito-borne flaviviruses, substantial subgenomic flaviviral RNA (sfRNA) is produced via incomplete degradation of viral genomic RNA by host XRN1. Zika virus (ZIKV) sfRNA has been detected in mosquito and mammalian somatic cells. Human neural progenitor cells (hNPCs) in the developing brain are the major target cells of ZIKV, and antiviral RNA interference (RNAi) plays a critical role in hNPCs. However, whether ZIKV sfRNA was produced in ZIKV-infected hNPCs as well as its function remains not known. In this study, we demonstrate that abundant sfRNA was produced in ZIKV-infected hNPCs. RNA pulldown and mass spectrum assays showed ZIKV sfRNA interacted with host proteins RHA and PACT, both of which are RNA-induced silencing complex (RISC) components. Functionally, ZIKV sfRNA can antagonize RNAi by outcompeting small interfering RNAs (siRNAs) in binding to RHA and PACT. Furthermore, the 3' stem loop (3'SL) of sfRNA was responsible for RISC components binding and RNAi inhibition, and 3'SL can enhance the replication of a viral suppressor of RNAi (VSR)-deficient virus in a RHA- and PACT-dependent manner. More importantly, the ability of binding to RISC components is conversed among multiple flaviviral 3'SLs. Together, our results identified flavivirus 3'SL as a potent VSR in RNA format, highlighting the complexity in virus-host interaction during flavivirus infection.IMPORTANCEZika virus (ZIKV) infection mainly targets human neural progenitor cells (hNPCs) and induces cell death and dysregulated cell-cycle progression, leading to microcephaly and other central nervous system abnormalities. RNA interference (RNAi) plays critical roles during ZIKV infections in hNPCs, and ZIKV has evolved to encode specific viral proteins to antagonize RNAi. Herein, we first show that abundant sfRNA was produced in ZIKV-infected hNPCs in a similar pattern to that in other cells. Importantly, ZIKV sfRNA acts as a potent viral suppressor of RNAi (VSR) by competing with siRNAs for binding RISC components, RHA and PACT. The 3'SL of sfRNA is responsible for binding RISC components, which is a conserved feature among mosquito-borne flaviviruses. As most known VSRs are viral proteins, our findings highlight the importance of viral non-coding RNAs during the antagonism of host RNAi-based antiviral innate immunity.

Changes in functional connectivity and structural covariance between the fronto-parietal network and medial orbitofrontal cortex are associated with disinhibition in restrained eaters.

Disinhibition, characterized by a loss of dietary control, is a significant risk factor for diet failure and the onset of eating disorders in restrained eaters. This study employs resting-state functional connectivity and structural covariance network analyses to explore the neural associations underlying this behavior. By analyzing functional MRI data from 63 female college students, we found that increased disinhibition correlates with enhanced functional connectivity between the medial orbitofrontal cortex and key components of the inhibition system, particularly within the fronto-parietal network. Moreover, we observed a relationship between the structural covariance of the medial orbitofrontal cortex and the inferior parietal lobule and the severity of disinhibition. Importantly, the functional connectivity between the medial orbitofrontal cortex and the inferior parietal lobule predicts the severity of binge eating symptoms in these individuals. These findings indicate that imbalances in the interaction between the brain's reward and inhibition systems can lead to dietary failures and eating disorders, emphasizing the need for targeted interventions.

The glycosylation deficiency of flavivirus NS1 attenuates virus replication through interfering with the formation of viral replication compartments.

BACKGROUND: Flavivirus is a challenge all over the world. The replication of flavivirus takes place within membranous replication compartments (RCs) derived from endoplasmic reticulum (ER). Flavivirus NS1 proteins have been proven essential for the formation of viral RCs by remodeling the ER. The glycosylation of flavivirus NS1 proteins is important for viral replication, yet the underlying mechanism remains unclear. METHODS: HeLa cells were used to visualize the ER remodeling effects induced by NS1 expression. ZIKV replicon luciferase assay was performed with BHK-21 cells. rZIKV was generated from BHK-21 cells and the plaque assay was done with Vero Cells. Liposome co-floating assay was performed with purified NS1 proteins from 293T cells. RESULTS: We found that the glycosylation of flavivirus NS1 contributes to its ER remodeling activity. Glycosylation deficiency of NS1, either through N-glycosylation sites mutations or tunicamycin treatment, compromises its ER remodeling activity and interferes with viral RCs formation. Disruption of NS1 glycosylation results in abnormal aggregation of NS1, rather than reducing its membrane-binding activity. Consequently, deficiency in NS1 glycosylation impairs virus replication. CONCLUSIONS: In summary, our results highlight the significance of NS1 glycosylation in flavivirus replication and elucidate the underlying mechanism. This provides a new strategy for combating flavivirus infections.

Genetically encoded site-specific 19F unnatural amino acid incorporation in V. natriegens for in-cell NMR analysis.

Nuclear magnetic resonance (NMR) spectroscopy NMR is a well-established technique for probing protein structure, dynamics and conformational changes. Taking advantage of the high signal sensitivity and broad chemical shift range of 19F nuclei, 19F NMR has been applied to investigate protein function at atomic resolution. In this report, we extend the unnatural amino acid site-specific incorporation into V. natriegens, an alternate protein expression system. The unnatural amino acid L-4-trifluoromethylphenylalanine (tfmF) was site-specifically introduced into the mitogen-activated protein kinase MEKK3 in V. natriegens using genetically encoded technology, which will be an extensive method for in-cell protein structure and dynamic investigation.

Structural insights into somatostatin receptor 5 bound with cyclic peptides.

Somatostatin receptor 5 (SSTR5) is highly expressed in ACTH-secreting pituitary adenomas and is an important drug target for the treatment of Cushing's disease. Two cyclic SST analog peptides (pasireotide and octreotide) both can activate SSTR5 and SSTR2. Pasireotide is preferential binding to SSTR5 than octreotide, while octreotide is biased to SSTR2 than SSTR5. The lack of selectivity of both pasireotide and octreotide causes side effects, such as hyperglycemia, gastrointestinal disturbance, and abnormal glucose homeostasis. However, little is known about the binding and selectivity mechanisms of pasireotide and octreotide with SSTR5, limiting the development of subtype-selective SST analog drugs specifically targeting SSTR5. Here, we report two cryo-electron microscopy (cryo-EM) structures of SSTR5-Gi complexes activated by pasireotide and octreoitde at resolutions of 3.09 Å and 3.24 Å, respectively. In combination with structural analysis and functional experiments, our results reveal the molecular mechanisms of ligand recognition and receptor activation. We also demonstrate that pasireotide preferentially binds to SSTR5 through the interactions between Tyr(Bzl)/DTrp of pasireotide and SSTR5. Moreover, we find that the Q2.63, N6.55, F7.35 and ECL2 of SSTR2 play a crucial role in octreotide biased binding of SSTR2. Our results will provide structural insights and offer new opportunities for the drug discovery of better selective pharmaceuticals targeting specific SSTR subtypes.

Highly stretchable graphene kirigami with tunable mechanical properties.

In recent years, kirigami techniques have inspired the design of graphene-based nanodevices with exceptional stretchability and ductility. Based on an I-shaped cutting pattern, here we propose a graphene kirigami that exhibits remarkable stretchability and ductility in two independent planar directions along with negative Poisson's ratios. The deformation mechanism underlying the high stretchability of the structure is the flipping and rotation of its cutting ligaments during elongation. Molecular dynamics simulations show that the yield and fracture strains of graphene kirigami can be enhanced by factors of 6 and 10 in the two planar directions. In addition, the mechanical properties of the graphene kirigami can be tuned by altering the cutting geometric parameters as well as incorporating distinct cutting patterns in series. We develop a numerical algorithm to predict the stress-strain response of the series-connected graphene kirigami, and verify its accuracy using appropriate simulations. On this basis, the stress-strain response of the series-connected graphene kirigami can be tuned by altering its geometric parameters and the number of building blocks. This graphene kirigami could be applied to the design and development of next-generation flexible electronics such as stretchable electrodes and strain sensors.

Quercetin stimulates an accelerated burst of oviposition-based reproductive strategy in codling moth controlled by juvenile hormone signaling pathway.

The advantageous characteristics of invasive pests, particularly their ability to reproduce and adapt to the environment, have been observed. However, it remains unclear what specific inherent superiority enables fruit pests to successfully invade and dominate in interactions with other species. In this study, we report that Cydia pomonella (Linnaeus), a notorious invasive pest of pome fruits and walnuts globally, employs unique reproductive strategies in response to quercetin, a plant compound in host fruits. By monitoring adult dynamics and fruit infestation rates, we observed a competitive relationship between C. pomonella and the native species Grapholita molesta (Busck). C. pomonella was able to occupy vacant niches to ensure its population growth. We also found that quercetin had different effects on the reproductive capacity and population growth of C. pomonella and G. molesta. While quercetin stimulated the fecundity and population growth of G. molesta, it inhibited C. pomonella. However, C. pomonella was able to rapidly increase its population after exposure to quercetin by adopting an 'accelerated burst' of oviposition strategy, with each individual making a greater reproductive contribution compared to the control. We further demonstrated that the effect of quercetin on oviposition is regulated by the juvenile hormone (JH) signaling pathway in C. pomonella, allowing it to prioritize survival. The enhanced reproductive fitness of G. molesta in response to quercetin is attributed to the regulation of JH titers and key genes such as Met and Kr-h1, which in turn up-regulate reproduction-related genes Vg and VgR. In contrast, C. pomonella is inhibited. These findings shed light on the mechanisms interspecific competition and help to improve our understanding of the global spread of C. pomonella, which can be attributed to its inherent superiority in terms of reproductive strategy.

Secondary metabolites in host pears defense against two fruit borers and cytochrome-P450-mediated counter-defense.

Herbivorous insects have evolved metabolic strategies to survive the challenges posed by plant secondary metabolites (SMs). This study reports an exploration of SMs present in pears, which serve as a defense against invasive Cydia pomonella and native Grapholita molesta and their counter-defense response. The feeding preferences of fruit borers are influenced by the softening of two pear varieties as they ripen. The content of SMs, such as quercetin and rutin, increases due to feeding by fruit borers. Notably, quercetin levels only increase after C. pomonella feeding. The consumption of SMs affects the growth of fruit borer population differently, potentially due to the activation of P450 genes by SMs. These two fruit borers are equipped with specific P450 enzymes that specialize in metabolizing quercetin and rutin, enabling them to adapt to these SMs in their host fruits. These findings provide valuable insights into the coevolution of plants and herbivorous insects.

Different conformational responses of the ß2-adrenergic receptor-Gs complex upon binding of the partial agonist salbutamol or the full agonist isoprenaline.

G protein-coupled receptors (GPCRs) are responsible for most cytoplasmic signaling in response to extracellular ligands with different efficacy profiles. Various spectroscopic techniques have identified that agonists exhibiting varying efficacies can selectively stabilize a specific conformation of the receptor. However, the structural basis for activation of the GPCR-G protein complex by ligands with different efficacies is incompletely understood. To better understand the structural basis underlying the mechanisms by which ligands with varying efficacies differentially regulate the conformations of receptors and G proteins, we determined the structures of ß2AR-Gαs[Formula: see text]γ bound with partial agonist salbutamol or bound with full agonist isoprenaline using single-particle cryo-electron microscopy at resolutions of 3.26 Å and 3.80 Å, respectively. Structural comparisons between the ß2AR-Gs-salbutamol and ß2AR-Gs-isoprenaline complexes demonstrated that the decreased binding affinity and efficacy of salbutamol compared with those of isoprenaline might be attributed to weakened hydrogen bonding interactions, attenuated hydrophobic interactions in the orthosteric binding pocket and different conformational changes in the rotamer toggle switch in TM6. Moreover, the observed stronger interactions between the intracellular loop 2 or 3 (ICL2 or ICL3) of ß2AR and Gαs with binding of salbutamol versus isoprenaline might decrease phosphorylation in the salbutamol-activated ß2AR-Gs complex. From the observed structural differences between these complexes of ß2AR, a mechanism of ß2AR activation by partial and full agonists is proposed to provide structural insights into ß2AR desensitization.