Representative Member Research Articles

Newcastle disease virus activates diverse signaling pathways via Src to facilitate virus entry into host macrophages.

In general, it is known that the paramyxovirus gains access to host cells through direct penetration at the plasma membrane; however, emerging evidence suggests more complex entry mechanisms for paramyxoviruses. The endocytic entry of Newcastle disease virus (NDV), a representative member of the paramyxovirus family, into multiple types of cells has been recently reported. Herein, we demonstrate the binding of NDV to induce ganglioside-activated Src signaling, which is responsible for the endocytic entry of NDV through caveolae-mediated endocytosis. This process involved Src-dependent activation of the caveolae-associated Cav1 and Dyn2, as well as specific Rho GTPase and downstream effectors, thereby orchestrating the endocytic entry process of NDV. Our findings uncover a novel molecular mechanism of endocytic entry of NDV into host cells and provide novel insight into paramyxovirus mechanisms of entry.

Subcellular Colocalization Assay of Host Factors with Viral Replication Complex in the dsRNA Reporter Nicotiana benthamiana.

As obligate pathogens, plant viruses co-opt several host factors for viral replication. Double-stranded RNA (dsRNA) plays important roles in plants, including eliciting innate immune responses and RNA interference. dsRNA also represents the genetic entities of a number of viruses and is a marker of infection by positive-sense single-stranded RNA viruses. Previous detection methods for RNA viruses basically relied on immunological methods, but later researchers discovered that the dsRNA-binding domain of the Flock house virus B2 protein is a perfect alternative to the J2 mAb for sensitive and rapid detection of long dsRNA in vitro and in vivo, and developed B2:GFP transgenic Nicotiana benthamiana line. This method describes in detail how to visualize host factors in the viral replication complex in time and space with the help of B2:GFP transgenic plants, exemplified by Turnip mosaic virus (TuMV), a representative virus member of the Potyviruses.

Annexin gene family in Spirometra mansoni (Cestoda: Diphyllobothriidae) and its phylogenetic pattern among Platyhelminthes of medical interest.

The plerocercoid larvae of Spirometra mansoni are etiological agents of human and animal sparganosis. Annexins are proteins with important roles in parasites. However, our knowledge of annexins in S. mansoni is still inadequate. In this study, 18 new members of the Annexin (ANX) family were characterized in S. mansoni. The clustering analysis demonstrated that all the SmANXs were divided into two main classes, consistent with the patterns of conserved motif organization. The 18 SmANXs were detected at all developmental stages (plerocercoid, adult, and egg) and displayed ubiquitous but highly variable expression patterns in all tissues/organs studied. The representative member rSmANX18 was successfully cloned and expressed. The protein was immunolocalized in the tegument and parenchyma of the plerocercoid and in the tegument, parenchyma, uterus and egg shell of adult worms. The recombinant protein can bind phospholipids with high affinity in a Ca2+-dependent manner, shows high anticoagulant activity and combines with FITC to recognize apoptotic cells. Annexin gene polymorphism and conservative core motif permutation were found in both cestodes and trematodes. SmANXs also revealed high genetic diversity among Platyhelminthes of medical interest. Our findings lay a foundation for further studies on the biological functions of ANXs in S. mansoni as well as other taxa in which ANXs occur.

The Copper Efflux Regulator (CueR).

The copper efflux regulator (CueR) is a classical member of the MerR family of metalloregulators and is common in gram-negative bacteria. Through its C-terminal effector-binding domain, CueR senses cytoplasmic copper ions to regulate the transcription of genes contributing to copper homeostasis, an essential process for survival of all cells. In this chapter, we review the regulatory roles of CueR in the model organism Escherichia coli and the mechanisms for CueR in copper binding, DNA recognition, and interplay with RNA polymerase in regulating transcription. In light of biochemical and structural analyses, we provide molecular details for how CueR represses transcription in the absence of copper ions, how copper ions mediate CueR conformational change to form holo CueR, and how CueR bends and twists promoter DNA to activate transcription. We also characterize the functional domains and key residues involved in these processes. Since CueR is a representative member of the MerR family, elucidating its regulatory mechanisms could help to understand the CueR-like regulators in other organisms and facilitate the understanding of other metalloregulators in the same family.

Huperzine alkaloids: forty years of total syntheses.

Covering: up to 2023Huperzine alkaloids are a group of natural products belonging to the Lycopodium alkaloids family. The representative member huperzine A has a unique structure and exhibits potent inhibitory activity against acetylcholine esterase (AChE). This subfamily of alkaloids provides a great opportunity for developing synthetic methodologies and asymmetric synthesis. The efforts towards the synthesis of huperzine A have cultivated dozens of total syntheses and a rich body of new chemistry. Impressive progress has also been made in the synthesis of other huperzine alkaloids. The total syntheses of huperzines B, U, O, Q and R, structure reassignment and total syntheses of huperzines K, M and N have been reported in the past decade. This review focuses on the synthetic organic chemistry and the biosynthesis and medicinal chemistry of huperzines are also covered briefly.

Quantifying the impact of assimilating sensitive observations on heavy precipitation forecast with ensemble-based forecast sensitivity to observations technique

In this study, the Ensemble-based Forecast Sensitivity to Observations (EFSO) technique was used to examine the impact of assimilated sensitive observations on the forecast of heavy rainfall through three data assimilation strategies, i.e., running without observation assimilation (CNTL); with assimilation of all observations (CNTL_DA); with assimilation of sensitive observations (SEN_DA). The Infrared Atmospheric Sounder Interferometer presented the largest positive impact on the forecast, followed by the Advanced Microwave Sounding Unit A and surface land reports. The top 10% of observations contributing the largest positive impact on the forecast were located in the upstream ocean of the low-level winds associated with precipitation maximum. Compared to ensemble CNTL and CNTL_DA experiments, ensemble SEN_DA exhibited better precipitation probability, in which the representative member in ensemble SEN_DA successfully predicted the precipitation magnitude and location. Based upon the representative experiments of the ensemble CNTL, CNTL_DA, and SEN_DA, synoptic conditions were examined to understand how the SEN_DA improves the forecast of heavy precipitation. The three representative experiments exhibited similar characteristics and variations in large-scale circulation, horizontal water vapor transport, and wind fields. However, compared to the representative experiments of CNTL and CNTL_DA, the advection and barotropic components of potential divergence contributed to the increase and persistence of convective instability throughout heavy rainfall in SEN_DA. Heavy precipitation was closely associated with the intensity, duration, and location of the maximum convective instability. Furthermore, the release of convective instability depended on the mesoscale ascent, which determined the maximum and location of the precipitation. These results help to understand the optimization of observational design and assimilation strategy.

Discovery of Potent and Selective Next-Generation Pan-BCR-ABL Inhibitors for the Treatment of CML and Ph+ ALL

Introduction While 6 tyrosine kinase inhibitors (TKIs) are available for patients with chronic myeloid leukemia (CML) that is newly diagnosed (imatinib, dasatinib, nilotinib, bosutinib) or refractory to prior treatments (asciminib, ponatinib), only ponatinib has pan-BCR-ABL activity, i.e., maintains potent activity against all single resistance mutations, including the difficult to inhibit T315I and E255V mutants. BCR-ABL mutations are the major known mechanism of resistance not only in CML, but also in Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL), where use of ponatinib in newly diagnosed patients is yielding especially encouraging results, presumably due to its ability to suppress emergence of resistance mutations. However, toxicities observed with ponatinib often lead to dose reduction or termination that can limit the benefit of its pan-BCR-ABL profile. The suboptimal safety profile of ponatinib is consistent with its relative lack of kinome selectivity, and its potent activity against KDR, FLT3, and FGFRs (unique amongst the 6 approved BCR-ABL TKIs) could potentially contribute to its unique toxicities. We therefore set out to identify a selective, potent pan-BCR-ABL inhibitor. Methods Compound potency was determined by assessing effects on viability of Ba/F3 cell lines expressing BCR-ABL variants (wild type [WT] and 11 key mutant forms) and off-targets of interest. Preliminary assessment of broader kinome selectivity was performed by calculating the percentage of 88 kinases inhibited by >80% by 1 µM compound (S-score (80)). For efficacy studies, mice implanted with engineered Ba/F3 cell lines were dosed orally once daily. Results Ponatinib potently inhibited BCR-ABL WT, T315I, and E255V with IC50s of 2, 4, and 7 nM respectively, and all 9 other BCR-ABL mutants tested were inhibited with IC50s <4 nM. However, ponatinib also potently inhibited the activity of KDR, FLT3, and FGFR1, with IC50s of 5, 2, and 17 nM, respectively, and had a high kinome S-score (80) of 0.30. Cmpd A is a representative member of a series of compounds we have identified that have a pan-BCR-ABL inhibitory profile and substantially improved selectivity over ponatinib. Cmpd A, potently inhibited BCR-ABL WT, T315I, and E255V, with IC50s of 6, 21, and 55 nM respectively, and 8/9 other BCR-ABL mutants tested were inhibited with IC50s ≤13 nM (Y253H IC50 75 nM). But in contrast to ponatinib, Cmpd A had a high degree of selectivity over FLT3 and FGFR1 (IC50s 1898 and 3323 nM), good selectivity over KDR (IC50 420 nM), and a low kinome S-score (80) of 0.08. We next compared the efficacy and tolerability of ponatinib and Cmpd A in an efficacy model driven by E255V, a key clinical resistance mutant that both compounds inhibited less potently than T315I. At 10 mg/kg, ponatinib inhibited tumor growth by 64% compared to vehicle-treated mice and was well tolerated. At 25 mg/kg, while ponatinib induced strong tumor regression (by 82% compared to baseline), it was poorly tolerated (eg, keratinized skin in 4/6 mice). In contrast 120 mg/kg Cmpd A induced strong tumor regression, by 55%, and was well tolerated (eg, 0/6 mice had keratinized skin). Conclusions We have identified a series of next-generation pan-BCR-ABL inhibitors with substantially improved selectivity over ponatinib. In vitro and in vivo characterization of compounds with further improved selectivity profiles, and the potential to be developed for use in patients with refractory CML and newly diagnosed Ph+ ALL, will be presented.

Core-dependent post-translational modifications guide the biosynthesis of a new class of hypermodified peptides

The ribosomally synthesized and post-translationally modified peptide (RiPPs) class of natural products has undergone significant expansion due to the rapid growth in genome sequencing data. Using a bioinformatics approach, we identify the dehydrazoles, a novel class of hypermodified RiPPs that contain both side chain dehydration of Ser residues, and backbone heterocyclization at Ser, Thr, and Cys residues to the corresponding azol(in)es. Structure elucidation of the hypermodified peptide carnazolamide, a representative class member, shows that 18 post-translational modifications are installed by just five enzymes. Complete biosynthetic reconstitution demonstrates that dehydration is carried out by an unusual DUF4135 dehydration domain fused to a zinc-independent cyclase domain (CcaM). We demonstrate that CcaM only modifies Ser residues that precede an azole in the core peptide. As heterocyclization removes the carbonyl following the Ser residue, CcaM likely catalyzes dehydration without generating an enolate intermediate. Additionally, CcaM does not require the leader peptide, and this core-dependence effectively sets the order for the biosynthetic reactions. Biophysical studies demonstrate direct binding of azoles to CcaM consistent with this azole moiety-dependent dehydration. Bioinformatic analysis reveals more than 50 related biosynthetic gene clusters that contain additional catalysts that may produce structurally diverse scaffolds.

Physical Factors Affecting the Scale-Up of Vegetative Insecticidal Protein (Vip3A) Production by Bacillus thuringiensis Bt294

Vip3A (vegetative insecticidal protein) is a representative member of the Vip3 family, which is widely used for lepidopteran pest control. This Vip3A protein, a non-growth-associated protein, is an effective bioinsecticide against insect pests, but there is relatively little information about its production processes at large scales. Hence, the effects of environmental factors on Vip3A production by Bacillus thuringiensis Bt294 (antifoam agents, shaking speeds, agitation and aeration rates), as well as controlling physical conditions such as the lowest point of dissolved oxygen and controlling of culture pH, were observed in shaking flasks and bioreactors. The results showed that antifoam agents, flask types and shaking speeds had significant effects on Vip3A and biomass production. Cultivation without pH control and DO control in 5 L bioreactors at lower agitation and aeration rates, which was not favorable for biomass production, resulted in a high Vip3A protein production of 5645.67 mg/L. The scale-up studies of the Vip3A protein production in a pilot-scale 750 L bioreactor gave 3750.0 mg/L. Therefore, this study demonstrated the significant effects of agitation, aeration rates and culture pH on Vip3A production by B. thuringiensis Bt294. Balancing of physical conditions was necessary for obtaining the highest yield of Vip3A by slowing down the production rate of biomass. Moreover, this Vip3A protein has high potential as a bioinsecticide for lepidopteran pest control in organic crops. This information will be important for significantly increasing the Vip3A protein concentration by the bacterium and will be useful for field application at a lower cost.

Elucidating Multiple Electron-Transfer Pathways for Metavanadate Bioreduction by Actinomycetic Streptomyces microflavus.

While microbial reduction has gained widespread recognition for efficiently remediating environments polluted by toxic metavanadate [V(V)], the pool of identified V(V)-reducing strains remains rather limited, with the vast majority belonging to bacteria and fungi. This study is among the first to confirm the V(V) reduction capability of Streptomyces microflavus, a representative member of ubiquitous actinomycetes in environment. A V(V) removal efficiency of 91.0 ± 4.35% was achieved during 12 days of operation, with a maximum specific growth rate of 0.073 d-1. V(V) was bioreduced to insoluble V(IV) precipitates. V(V) reduction took place both intracellularly and extracellularly. Electron transfer was enhanced during V(V) bioreduction with increased electron transporters. The electron-transfer pathways were revealed through transcriptomic, proteomic, and metabolomic analyses. Electrons might flow either through the respiratory chain to reduce intracellular V(V) or to cytochrome c on the outer membrane for extracellular V(V) reduction. Soluble riboflavin and quinone also possibly mediated extracellular V(V) reduction. Glutathione might deliver electrons for intracellular V(V) reduction. Bioaugmentation of the aquifer sediment with S. microflavus accelerated V(V) reduction. The strain could successfully colonize the sediment and foster positive correlations with indigenous microorganisms. This study offers new microbial resources for V(V) bioremediation and improve the understanding of the involved molecular mechanisms.

Plastid phylogenomics provide evidence to accept a new genus Pseudopeucedanum (Apiaceae) separated from Peucedanum s.l.

Abstract The species Peucedanum ledebourielloides, a medicinally important herb, has previously been placed in Peucedanum s.l. based on dorsally compressed mericarps with slightly prominent dorsal ribs and narrowly winged lateral ribs. However, this genus has now been re-delimited in a narrow sense, namely Peucedanum s.s., further recognized by ternate leaves and glabrous plants, so that the taxonomic position of P. ledebourielloides is unknown. To investigate the taxonomic position of this species, we sequenced and assembled the complete plastomes for two samples of P. ledebourielloides. Phylogenetic analyses based on 79 protein-coding genes showed that the samples of P. ledebourielloides formed a separate clade and were distant from Peucedanum morisonii, a representative member of Peucedanum s.s., implying that P. ledebourielloides cannot not be placed in Peucedanum s.s. Additionally, plastome comparisons and chromosome studies indicated the clear specificity of P. ledebourielloides. In particular, pinnate leaves and dorsally compressed mericarps densely covered with forked scabrous bristles, slightly prominent dorsal ribs, narrowly winged lateral ribs, and one vitta in each furrow and two vittae on the commissure clearly distinguished P. ledebourielloides from Peucedanum s.s. and other genera of Apiaceae. Hence, a new genus, Pseudopeucedanum gen. nov., was established to accommodate this species. Our results improve the taxonomy of the family Apiaceae and provide guidance for the taxonomic revision of other taxa that are difficult to place systematically.

Design of novel analogues of t-DPH1 with reduced cytotoxicity, taking the three conserved characteristics of the dermaseptin family as the feasible starting point

In recent years, there has been growing scientific interest in balancing the bioactivity and cytotoxicity of antimicrobial peptides (AMPs) for more comprehensive research and application. In this study, we focused on t-DPH1, a representative member of the dermaseptin family, as a template. We synthesized five analogues were synthesised to to assess the impact of three conserved features of the dermaseptin family (1. the tryptophan residue at 3rd position, 2. the consensus motif at mid-region, and 3. C-terminal amidation) on the bioactivity of t-DPH1 and their potential to reduce cytotoxicity. Our results demonstrated that analogues lacking an amino group at the C-terminus, specifically t-DPH1-NH2, exhibited potent antimicrobial activity against Gram-negative bacteria, including Escherichia coli (E. coli) and Klebsiella pneumoniae (K. pneumoniae). Furthermore, t-DPH1-NH2 retained its robust antiproliferative activity against the human non-small lung cancer cell line H157. Regarding cytotoxicity, t-DPH1-NH2 displayed lower levels of haemolytic activity and cytotoxicity against two normal cell lines. Notably, we employed C. elegans as an in vivo model to assess the cytotoxicity of peptides, and the results revealed no induction of in vivo toxicity by t-DPH1-NH2. Collectively, t-DPH1-NH2 provides valuable insights for further investigation as promising bioactive peptide templates in the development of antibiotic and anticancer prototype drugs. The modifications implemented based on the conserved structure of the peptide family offer a new direction for reducing the cytotoxicity of peptides.

High-fat diet alters N-glycosylation of PTPRJ in murine liver

Protein tyrosine phosphatases (PTPs) regulate multiple signaling pathways. Disruption of tyrosine phosphorylation through imbalanced action between protein tyrosine kinases (RTKs) and PTPs is a hallmark of metabolic disorders, including insulin resistance. A representative member of the receptor-type PTP family, PTPRJ (DEP-1), was previously identified as a negative regulator of insulin signaling and possesses post-translational glycosylation sites. In this regard, it seems of great importance to decipher the structure of PTPRJ's glycosylation, particularly in the context of metabolic disturbances, but this has not been done in detail. Thus, here we aimed at characterizing the glycosylation pattern of PTPRJ in liver. We show that N-glycosylation accounts for up to half of PTPRJ's molecular weight. Applying mass spectrometry, we detected increased levels of high-mannose structures in PTPRJ in liver tissue of obese mice compared to lean littermates. In addition, complex neutral structures without fucose were also elevated in PTPRJ of high-fat diet (HFD) mice. Conversely, complex fucosylated N-glycans as well as sialylated bi- and triantennary N-glycans, were significantly reduced in PTPRJ of HFD-derived liver tissue compared to LFD by ∼two fold (P≤.01, P≤.0001 and P≤.001, respectively). In congruence with these findings, the mannosidase MAN2A1, responsible for the conversion of high-mannose to complex N-glycans, was significantly downregulated under HFD conditions. Here we present for the first time that HFD-induced obesity impacts on the glycosylation pattern of the insulin signaling component PTPRJ in liver. These findings may inspire new research on the glycosylation of PTPs in metabolic diseases and may open up new therapeutic approaches.

State-dependent dynamics of extramembrane domains in the β2 -adrenergic receptor.

G protein-coupled receptors (GPCRs) are membrane-bound signaling proteins that play an essential role in cellular signaling processes. Due to their intrinsic function of transmitting internal signals in response to external cues, these receptors are adapted to be highly dynamic in nature. The β2 -adrenergic receptor (β2 AR) is a representative member of the family that has been extensively analyzed in terms of its structure and activation. Although the structure of the transmembrane domain has been characterized in the different functional states of the receptor, the conformational dynamics of the extramembrane domains, especially the intrinsically disordered regions are still emerging. In this study, we analyze the state-dependent dynamics of extramembrane domains of β2 AR using atomistic molecular dynamics simulations. We introduce a parameter, the residue excess dynamics that allows us to better quantify receptor dynamics. Using this measure, we show that the dynamics of the extramembrane domains are sensitive to the receptor state. Interestingly, the ligand-bound intermediate state shows the maximal dynamics compared to either the active R*G or inactive R states. Ligand binding appears to be correlated with high residue excess dynamics that are dampened upon G protein coupling. The intracellular loop-3 (ICL3) domain has a tendency to flip towards the membrane upon ligand binding, which could contribute to receptor "priming." We highlight an important ICL1-helix-8 interplay that is broken in the ligand-bound state but is retained in the active state. Overall, our study highlights the importance of characterizing the functional dynamics of the GPCR loop domains.

Cryo-EM structure of cyanophage P-SCSP1u offers insights into DNA gating and evolution of T7-like viruses

Cyanophages, together with their host cyanobacteria, play important roles in marine biogeochemical cycles and control of marine food webs. The recently identified MPP-C (Marine Picocyanobacteria Podovirus clade C) cyanophages, belonging to the T7-like podoviruses, contain the smallest genomes among cyanopodoviruses and exhibit distinct infection kinetics. However, understanding of the MPP-C cyanophage infection process is hindered by the lack of high-resolution structural information. Here, we report the cryo-EM structure of the cyanophage P-SCSP1u, a representative member of the MPP-C phages, in its native form at near-atomic resolution, which reveals the assembly mechanism of the capsid and molecular interaction of the portal-tail complex. Structural comparison of the capsid proteins of P-SCSP1u and other podoviruses with known structures provides insights into the evolution of T7-like viruses. Furthermore, our study provides the near-atomic resolution structure of portal-tail complex for T7-like viruses. On the basis of previously reported structures of phage T7, we identify an additional valve and gate to explain the DNA gating mechanism for the T7-like viruses.

Inclusion/energy transfer/release of L-ascorbate in the interlayer gallery of layered yttrium hydroxide

L-ascorbate (AS, Vitamin C) was intercalated into the interlayer gallery of layered yttrium hydroxide (LYH), a representative member of the layered rare earth hydroxide series (LRHs), through an ion-exchange reaction in an aqueous solution of sodium ascorbate. Based on the expansion of the interlayer space of the resulting AS-LYH hybrid, it is suggested that AS anions vertically oriented to hydroxide layers would be arranged to form a partially interdigitated bilayer in the interlayer space of LYH with a high loading capacity and density of AS. This arrangement is different from the antiparallel monolayer configuration of AS anions that are horizontally or vertically oriented to the layer of classical layered double hydroxides (LDHs). The durability of AS against light and oxidation was sufficiently enhanced in the interlayer gallery of LYH so that no degradation occurred after exposure to simulated sunlight or heating at 60 °C for a long time. Practically no release of AS from AS-LYH was observed in a saline solution and simulated sea water, whereas a sustained release of AS occurred selectively in a phosphate buffer solution, with a release process best described by the first-order model. When LYH was doped with Tb3+ activator to give it photoluminescence ability, the UV light energy absorbed for the π → π* transition of AS was efficiently transferred to Tb3+, leading to strong green emission attributed to its 5D4 → 7F5 transition.

More Is Better: Dual-Acceptor Engineering for Constructing Second Near-Infrared Aggregation-Induced Emission Luminogens to Boost Multimodal Phototheranostics.

The manipulation of electron donor/acceptor (D/A) shows an endless impetus for innovating optical materials. Currently, there is booming development in electron donor design, while research on electron acceptor engineering has received limited attention. Inspired by the philosophical idea of "more is different", two systems with D'-D-A-D-D' (1A system) and D'-D-A-A-D-D' (2A system) structures based on acceptor engineering were designed and studied. It was demonstrated that the 1A system presented a weak aggregation-induced emission (AIE) to aggregation-caused quenching (ACQ) phenomenon, along with the increased acceptor electrophilicity and planarity. In sharp contrast, the 2A system with one more acceptor exhibited an opposite ACQ-to-AIE transformation. Interestingly, the fluorophore with a more electron-deficient A-A moiety in the 2A system displayed superior AIE activity. More importantly, all compounds in the 2A system showed significantly higher molar absorptivity (ε) in comparison to their counterparts in the 1A system. Thanks to the highest ε, near-infrared-II (NIR-II, 1000-1700 nm) emission, desirable AIE property, favorable reactive oxygen species (ROS) generation, and high photothermal conversion efficiency, a representative member of the 2A system handily performed in fluorescence-photoacoustic-photothermal multimodal imaging-guided photodynamic-photothermal collaborative therapy for efficient tumor elimination. Meanwhile, the NIR-II fluorescence imaging of blood vessels and lymph nodes in living mice was also accomplished. This study provides the first evidence that the dual-connected acceptor tactic could be a new molecular design direction for the AIE effect, resulting in high ε, aggregation-intensified NIR-II fluorescence emission, and improved ROS and heat generation capacities of phototheranostic agents.

Unfolding of an RNA G-quadruplex motif in the negative strand genome of porcine reproductive and respiratory syndrome virus by host and viral helicases to promote viral replication.

G-quadruplex (G4) is a unique secondary structure formed by guanine-rich nucleic acid sequences. Growing studies reported that the genomes of some viruses harbor G4 structures associated with viral replication, opening up a new field to dissect viral infection. Porcine reproductive and respiratory syndrome virus (PRRSV), a representative member of Arteriviridae, is an economically significant pathogen that has devastated the swine industry worldwide for over 30 years. In this study, we identified a highly conserved G-rich sequence with parallel-type G4 structure (named PRRSV-G4) in the negative strand genome RNA of PRRSV. Pyridostatin (PDS), a well-known G4-binding ligand, stabilized the PRRSV-G4 structure and inhibited viral replication. By screening the proteins interacting with PRRSV-G4 in PRRSV-infected cells and single-molecule magnetic tweezers analysis, we found that two helicases, host DDX18 and viral nsp10, interact with and efficiently unwound the PRRSV-G4 structure, thereby facilitating viral replication. Using a PRRSV reverse genetics system, we confirmed that recombinant PRRSV with a G4-disruptive mutation exhibited resistance to PDS treatment, thereby displaying higher replication than wild-type PRRSV. Collectively, these results demonstrate that the PRRSV-G4 structure plays a crucial regulatory role in viral replication, and targeting this structure represents a promising strategy for antiviral therapies.

Blocking ADP-ribosylation expands the anti-mycobacterial spectrum of rifamycins.

The clinical utility of rifamycins against non-tuberculous mycobacterial (NTM) disease is limited by intrinsic drug resistance achieved by ADP-ribosyltransferase Arr. By blocking the site of ribosylation, we recently optimized a series of analogs with substantially improved potency against Mycobacterium abscessus. Here, we show that a representative member of this series is significantly more potent than rifabutin against major NTM pathogens expressing Arr, providing a powerful medicinal chemistry approach to expand the antimycobacterial spectrum of rifamycins. IMPORTANCE Lung disease caused by a range of different species of non-tuberculous mycobacteria (NTM) is difficult to cure. The rifamycins are very active against Mycobacterium tuberculosis, which causes tuberculosis (TB), but inactive against many NTM species. Previously, we showed that the natural resistance of the NTM Mycobacterium abscessus to rifamycins is due to enzymatic inactivation of the drug by the bacterium. We generated chemically modified versions of rifamycins that prevent inactivation by the bacterium and thus become highly active against M. abscessus. Here, we show that such a chemically modified rifamycin is also highly active against several additional NTM species that harbor the rifamycin inactivating enzyme found in M. abscessus, including M. chelonae, M. fortuitum, and M. simiae. This finding expands the potential therapeutic utility of our novel rifamycins to include several currently difficult-to-cure NTM lung disease pathogens beyond M. abscessus.

Using Taxes to Attract the Creative Class in the Presence of a Region-Specific Rent

We analyze interregional competition between two regions A and B that use taxes to attract a representative creative class member (the entrepreneur). This entrepreneur establishes a firm in either region A or B, which guarantees her profit. However, if the entrepreneur locates in region A, she also obtains a stochastic, location-specific rent that is either high with positive probability or low with positive complementary probability. In this setting, we accomplish three tasks. First, given the values of the two tax rates, we determine the payoff to the entrepreneur in the two regions for the two possible values of the location-specific rent in A. Second, we ascertain when the entrepreneur will locate in A for both the rent values and when she will locate in B. Finally, we compute the tax rate that B will set and then specify a condition which ensures that the entrepreneur locates in B.