A mutation-sensitive, multiplexed and amplification-free detection of nucleic acids by stretching single-molecule tandem hairpin probes.

The detection of nucleic acid sequences in parallel with the discrimination of single nucleotide variations (SNVs) is critical for research and clinical applications. A few limitations make the detection technically challenging, such as too small variation in probe-hybridization energy caused by SNVs, the non-specific amplification of false nucleic acid fragments and the few options of dyes limited by spectral overlaps. To circumvent these limitations, we developed a single-molecule nucleic acid detection assay without amplification or fluorescence termed THREF (hybridization-induced tandem DNA hairpin refolding failure) based on multiplexed magnetic tweezers. THREF can detect DNA and RNA sequences at femtomolar concentrations within 30 min, monitor multiple probes in parallel, quantify the expression level of miR-122 in patient tissues, discriminate SNVs including the hard-to-detect G-U or T-G wobble mutations and reuse the probes to save the cost. In our demonstrative detections using mock clinic samples, we profiled the let-7 family microRNAs in serum and genotyped SARS-CoV-2 strains in saliva. Overall, the THREF assay can discriminate SNVs with the advantages of high sensitivity, ultra-specificity, multiplexing, reusability, sample hands-free and robustness.

Transcriptomic study of the role of MeFtsZ2-1 in pigment accumulation in cassava leaves.

MeFtsZ2-1 is a key gene for plant plastid division, but the mechanism by which MeFtsZ2-1 affects pigment accumulation in cassava (Manihot esculenta Crantz) through plastids remains unclear. We found that MeFtsZ2-1 overexpression in cassava (OE) exhibited darker colors of leaves, with increased levels of anthocyanins and carotenoids. Further observation via Transmission Electron Microscopy (TEM) revealed no apparent defects in chloroplast structure but an increase in the number of plastoglobule in OE leaves. RNA-seq results showed 1582 differentially expressed genes (DEGs) in leaves of OE. KEGG pathway analysis indicated that these DEGs were enriched in pathways related to flavonoid, anthocyanin, and carotenoid biosynthesis. This study reveals the role of MeFtsZ2-1 in cassava pigment accumulation from a physiological and transcriptomic perspective, providing a theoretical basis for improving cassava quality.

KSHV RTA antagonizes SMC5/6 complex-induced viral chromatin compaction by hijacking the ubiquitin-proteasome system.

Kaposi's sarcoma-associated herpesvirus (KSHV) is a double-stranded DNA virus with the capacity to establish life-long latent infection. During latent infection, the viral genome persists as a circular episome that associates with cellular histones and exists as a nonintegrated minichromosome in the nucleus of infected cells. Chromatin structure and epigenetic programming are required for the proper control of viral gene expression and stable maintenance of viral DNA. However, there is still limited knowledge regarding how the host regulates the chromatin structure and maintenance of episomal DNA. Here, we found that the cellular protein structural maintenance of chromosome (SMC) complex SMC5/6 recognizes and associates with the KSHV genome to inhibit its replication. The SMC5/6 complex can bind to the KSHV genome and suppress KSHV gene transcription by condensing the viral chromatin and creating a repressive chromatin structure. Correspondingly, KSHV employs an antagonistic strategy by utilizing the viral protein RTA to degrade the SMC5/6 complex and antagonize the inhibitory effect of this complex on viral gene transcription. Interestingly, this antagonistic mechanism of RTA is evolutionarily conserved among γ-herpesviruses. Our work suggests that the SMC5/6 complex is a new host factor that restricts KSHV replication.

CD226 knockout reduces the development of CD8+ T by impairing the TCR sensitivity of double-positive thymocytes.

The costimulation molecule CD226 is widely involved in T cell differentiation, activation and immune functional regulation in peripheral immune tissues. CD226-deficient mice have impaired immune response capacity. The function of CD226 in regulating T cell development in the thymus, a central immune organ, is not yet fully understood. We investigated the development of thymocytes using CD226 knockout mice and single-cell sequencing techniques. CD226 began to be expressed in the second half of thymocyte development, with a gradual increase from the double-positive (DP) to single-positive (SP) phase and higher levels of CD226 on CD8+ T cells than on CD4+ T cells from the SP phase to mature T cells. In the thymus of CD226KO mice, the proportion of DPT at the quiescent phase (DPT-Q) increased, of which the Gzma+ cluster that tends to be CD8+ T cells and CD5+ cluster that is undergoing positive selection decreased dramatically. Afterward, the proportion of mature CD8+ T cells reduced dramatically. Depletion of CD226 impaired TCR activation signalling and diminished AKT/ERK/NF-κB/p38 phosphorylation levels. The diminished TCR responsiveness of DPT cells impeded their positive selection process and influenced the maturation of CD8+ T cells. In mechanism, CD226 knockout enhanced DPT cell apoptosis via impairing AKT phosphorylation. These results suggest that CD226 plays a significant role in T cell thymic development via modulation of TCR signalling, affecting CD8+ T cell maturation.

CD226 deficiency exacerbated intestinal immune dysregulation in mice with dinitrochlorobenzene-induced atopic dermatitis.

Intestinal mucosal immunity plays a pivotal role in host defence. In this study, we found that cluster of differentiation 226 (CD226) gene knockout (KO) led to more severe atopic dermatitis (AD)-related skin pathologies and bowel abnormalities in a 2,4-dinitrochlorobenzene (DNCB)-induced AD-like mouse model. Following DNCB administration, the expression of CD226 was elevated in intestinal mucosal tissues, including group 3 innate lymphoid cells (ILC3s) and CD4+ T cells of Peyer's patches (PPs). CD226 deficiency led to an overactive intestinal immune response in the AD-like mice, as evidenced by increased inflammation and Th1/Th2-related cytokine levels as well as increased Paneth cell numbers and antimicrobial peptide (AMP) expression, which was likely due to the higher interleukin (IL)-22 production in the lamina propria. Additionally, CD226 deficiency increased the production of IL-4 and IL-17 in mesenteric lymph nodes as well as the number of PPs and expression of immunoglobulin (Ig) A in B cells. Moreover, insufficient expression of CD226 affected the characterization of intraepithelial and lamina propria lymphocytes in the intestinal mucosa. Finally, the number of PPs was increased in CD4+ T cell-specific CD226 KO and regulatory T (Treg) cell-specific CD226 KO mice; thus, loss of CD226 in Treg cells resulted in impaired Treg cell-suppressive function. Therefore, our findings indicate that CD226 deficiency alters intestinal immune functionality in inflammatory diseases.

Host RAB11FIP5 protein inhibits the release of Kaposi's sarcoma-associated herpesvirus particles by promoting lysosomal degradation of ORF45.

Open reading frame (ORF) 45 is an outer tegument protein of Kaposi's sarcoma-associated herpesvirus (KSHV). Genetic analysis of an ORF45-null mutant revealed that ORF45 plays a key role in the events leading to the release of KSHV particles. ORF45 associates with lipid rafts (LRs), which is responsible for the colocalization of viral particles with the trans-Golgi network and facilitates their release. In this study, we identified a host protein, RAB11 family interacting protein 5 (RAB11FIP5), that interacts with ORF45 in vitro and in vivo. RAB11FIP5 encodes a RAB11 effector protein that regulates endosomal trafficking. Overexpression of RAB11FIP5 in KSHV-infected cells decreased the expression level of ORF45 and inhibited the release of KSHV particles, as reflected by the significant reduction in the number of extracellular virions. In contrast, silencing endogenous RAB11FIP5 increased ORF45 expression and promoted the release of KSHV particles. We further showed that RAB11FIP5 mediates lysosomal degradation of ORF45, which impairs its ability to target LRs in the Golgi apparatus and inhibits ORF45-mediated colocalization of viral particles with the trans-Golgi network. Collectively, our results suggest that RAB11FIP5 enhances lysosome-dependent degradation of ORF45, which inhibits the release of KSHV particles, and have potential implications for virology and antiviral design.

Plasma proteomic analysis reveals altered protein abundances in HIV-infected patients with or without non-Hodgkin lymphoma.

The identification of circulating proteins associated with acquired immunodeficiency syndrome-related non-Hodgkin lymphoma (AIDS-NHL) may help in the development of promising biomarkers for screening, diagnosis, treatment, and prognosis. Here, we used quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify differentially expressed proteins (DEPs) in plasma collected from patients with AIDS-NHL and human immunodeficiency virus (HIV)-infected patients without NHL (HIV+ ). Proteins with a log2 (fold change) in abundance >0.26 and p < 0.05 were considered differentially abundant. In total, 84 DEPs were identified, among which 20 were further validated as potential biomarkers, with immunoglobulin and complement components being the most common proteins. Some of the proteins were further verified in a retrospective analysis of the medical records of patients in a larger cohort. These markedly altered proteins were found to mediate pathophysiological pathways that likely contribute to AIDS-NHL pathogenesis, such as the humoral immune response, complement activation, and complement and coagulation cascades. Our findings provide a new molecular understanding of AIDS-NHL pathogenesis and provide new evidence supporting the identification of these proteins as possible biomarkers in AIDS-NHL.

Interleukin-6 absence triggers intestinal microbiota dysbiosis and mucosal immunity in mice.

Interleukin-6 (IL-6) in mucosal immune cells is involved in post-injury intestinal regeneration, inflammation responses, and gastric homeostasis. However, the interaction between IL-6 and the dynamic balance of gut microbiota (GM) remains unexplored. Intestinal pathology was assessed by hematoxylin and eosin and periodic acid-Schiff staining in wild-type (WT) and IL-6 gene knockout (KO) C57BL/6J mice. GM profiles were established via high-throughput sequencing of the fecal bacterial 16S rRNA gene. Intestinal α- and ß-defensins were measured by quantitative real-time PCR; further, flow cytometry was performed to analyze isolated intraepithelial lymphocytes (IELs). Compared with the WT, IL-6 KO did not obviously change gut structures, but significantly reduced GM diversity, resulting in reduced metabolic pathways with decreased gram-positive but elevated gram-negative bacteria. More taxa alterations included differences at the phyla (e.g., increased Verrucomicrobia and decreased Firmicutes) and genera (e.g., increased Akkermansia and decreased Lactobacillus) levels. Absence of IL-6 also significantly increased intestinal expression of defensins α3 and α4 (Defa3 and Defa4) and the percentage of natural TCRγδ+ IELs, providing a molecular basis for triggering mucosal immune response. Therefore, IL-6 loss remodels GM composition and alters IEL maintenance, identifying IL-6 as a crucial cytokine for GM dysbiosis and mucosal immunity.

Discovery of aminothiazole derivatives as novel human enterovirus A71 capsid protein inhibitors.

Enterovirus A71 (EV-A71), one of the major pathogens that causes hand, foot and mouth disease (HFMD), has seriously threatened the health and safety of young children. In this study, aminothiazole derivatives were synthesized and screened against EV-A71 in Rhabdomyosarcoma (RD) cells. The best compound (12s), with a biphenyl group, showed activity against EV-A71 (EC50: 0.27 µM) but also against a series of different human enteroviruses without significant cytotoxicity (CC50 > 56.2 µM). Mechanistic studies including time-of-drug-addition assays, viral entry assays and microscale thermophoresis (MST) experiments, showed that 12s binds to EV-A71 capsid and blocks the binding between the viral protein VP1 and the relevant human scavenger receptor class B member 2 (hSCARB2).

Quality of life assessment after total knee arthroplasty in patients with Parkinson's disease.

BACKGROUND: The number of Parkinson's patients (PD) undergoing total knee arthroplasty (TKA) is increasing. The purpose of the study was to characterize quality of life (QOL) outcomes for patients with coexisting PD and knee osteoarthritis (KOA) following TKA. METHODS: Patients with coexisting PD and KOA undergoing TKA between June 2014 and June 2020 were included. These patients were matched to controls with KOA alone by age, gender, basic social background information and Knee society score (KSS). The primary measure was to assess the QOL by the absolute changes in the EuroQOL5-Dimensions (EQ-5D), Pain and Disability Questionnaire (PDQ), and Patient Health Questionnaire-9(PHQ-9) at the last follow-up (LFU). Secondary measures were changes in QOL that exceeded the minimum clinically important difference value (MCID). Data on the health status and QOL of all patients were collected. Simple and multivariate regression analysis was used to evaluate the impact of PD on their QOL. RESULTS: Twelve KOA patients with PD were compared with 48 controls. Control patients experienced QOL improvement across all three measures:EQ-5D index (0.545-0.717, P < 0.01), PDQ (81.1-52.3, P < 0.01) and PHQ-9(8.22-5.91, P < 0.01) were significantly improved at the LFU; while in patients with PD, only PDQ (91.0-81.4, P = 0.03) slightly improved. There were significant differences in the improvement of QOL between PD patients and the control group through EQ-5D (0.531 vs.0.717, P < 0.01) and PDQ (81.4vs.52.3, P < 0.01) at the LFU. CONCLUSION: TKA has no benefit of QOL beyond a slight improvement in pain-related disability in the KOA patients with PD.

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9-mediated kif15 mutations accelerate axonal outgrowth during neuronal development and regeneration in zebrafish.

KIF15, the vertebrate kinesin-12, is best known as a mitotic motor protein, but continues to be expressed in neurons. Like KIF11 (the vertebrate kinesin-5), KIF15 interacts with microtubules in the axon to limit their sliding relative to one another. Unlike KIF11, KIF15 also regulates interactions between microtubules and actin filaments at sites of axonal branch formation and in growth cones. Our original work on these motors was done on cultured rat neurons, but we are now using zebrafish to extend these studies to an in vivo model. We previously studied kif15 in zebrafish by injecting splice-blocking morpholinos injected into embryos. Consistent with the cell culture work, these studies demonstrated that axons grow faster and longer when KIF15 levels are reduced. In the present study, we applied CRISPR/Cas9-based knockout technology to create kif15 mutants and labeled neurons with Tg(mnx1:GFP) transgene or transient expression of elavl3:EGFP-alpha tubulin. We then compared by live imaging the homozygotic, heterozygotic mutants to their wildtype siblings to ascertain the effects of depletion of kif15 during Caudal primary motor neuron and Rohon-Beard (R-B) sensory neuron development. The results showed, compared to the kif15 wildtype, the number of branches was reduced while axon outgrowth was accelerated in kif15 homozygotic and heterozygotic mutants. In R-B sensory neurons, after laser irradiation, injured axons with loss of kif15 displayed significantly greater regenerative velocity. Given these results and the fact that kif15 drugs are currently under development, we posit kif15 as a novel target for therapeutically augmenting regeneration of injured axons.

NCOA2 promotes lytic reactivation of Kaposi's sarcoma-associated herpesvirus by enhancing the expression of the master switch protein RTA.

Reactivation of Kaposi's sarcoma-associated herpesvirus (KSHV) is important for persistent infection in the host as well as viral oncogenesis. The replication and transcription activator (RTA) encoded by KSHV ORF50 plays a central role in the switch from viral latency to lytic replication. Given that RTA is a transcriptional activator and RTA expression is sufficient to activate complete lytic replication, RTA must possess an elaborate mechanism for regulating its protein abundance. Previous studies have demonstrated that RTA could be degraded through the ubiquitin-proteasome pathway. A protein abundance regulatory signal (PARS), which consists of PARS I and PARS II, at the C-terminal region of RTA modulates its protein abundance. In the present study, we identified a host protein named Nuclear receptor coactivator 2 (NCOA2), which can interact with RTA in vitro and in vivo. We further showed that NCOA2 binds to the PARS II domain of RTA. We demonstrated that NCOA2 enhances RTA stability and prevents the proteasome-mediated degradation of RTA by competing with MDM2, an E3 ubiquitin ligase of RTA that interacts with the PARS II domain. Moreover, overexpression of NCOA2 in KSHV-infected cells significantly enhanced the expression level of RTA, which promotes the expression of RTA downstream viral lytic genes and lytic replication. In contrast, silencing of endogenous NCOA2 downregulated the expression of viral lytic genes and impaired viral lytic replication. Interestingly, we also found that RTA upregulates the expression of NCOA2 during lytic reactivation. Taken together, our data support the conclusion that NCOA2 is a novel RTA-binding protein that promotes RTA-driven lytic reactivation by increasing the stability of RTA, and the RTA-NCOA2 positive feedback regulatory loop plays an important role in KSHV reactivation.

Dominant point mutation in a tetraspanin gene associated with field-evolved resistance of cotton bollworm to transgenic Bt cotton.

Extensive planting of crops genetically engineered to produce insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) has suppressed some major pests, reduced insecticide sprays, enhanced pest control by natural enemies, and increased grower profits. However, rapid evolution of resistance in pests is reducing these benefits. Better understanding of the genetic basis of resistance to Bt crops is urgently needed to monitor, delay, and counter pest resistance. We discovered that a point mutation in a previously unknown tetraspanin gene in the cotton bollworm (Helicoverpa armigera), a devastating global pest, confers dominant resistance to Cry1Ac, the sole Bt protein produced by transgenic cotton planted in China. We found the mutation using a genome-wide association study, followed by fine-scale genetic mapping and DNA sequence comparisons between resistant and susceptible strains. CRISPR/Cas9 knockout of the tetraspanin gene restored susceptibility to a resistant strain, whereas inserting the mutation conferred 125-fold resistance in a susceptible strain. DNA screening of moths captured from 23 field sites in six provinces of northern China revealed a 100-fold increase in the frequency of this mutation, from 0.001 in 2006 to 0.10 in 2016. The correspondence between the observed trajectory of the mutation and the trajectory predicted from simulation modeling shows that the dominance of the mutation accelerated adaptation. Proactive identification and tracking of the tetraspanin mutation demonstrate the potential for genomic analysis, gene editing, and molecular monitoring to improve management of resistance.

The mirid bug Apolygus lucorum deploys a glutathione peroxidase as a candidate effector to enhance plant susceptibility.

The mirid bug Apolygus lucorum has become a major agricultural pest since the large-scale cultivation of Bt-cotton. It was assumed that A. lucorum, similarly to other phloem sap insects, could secrete saliva that contains effector proteins into plant interfaces to perturb host cellular processes during feeding. However, the secreted effectors of A. lucorum are still uncharacterized and unstudied. In this study, 1878 putative secreted proteins were identified from the transcriptome of A. lucorum, which either had homology with published aphid effectors or shared common features with plant pathogens and insect effectors. One hundred and seventy-two candidate effectors were used for cell death-inducing/suppressing assays, and a putative salivary gland effector, Apolygus lucorum cell death inhibitor 6 (Al6), was characterized. The mRNAs of Al6 were enriched at feeding stages (nymph and adult) and, in particular, in salivary glands. Moreover, we revealed that the secreted Al6 encoded an active glutathione peroxidase that reduced reactive oxygen species (ROS) accumulation induced by INF1 or Flg22. Expression of the Al6 gene in planta altered insect feeding behavior and promoted plant pathogen infections. Inhibition of cell death and enhanced plant susceptibility to insect and pathogens are dependent on glutathione peroxidase activity of Al6. Thus, this study shows that a candidate salivary gland effector, Al6, functions as a glutathione peroxidase and suppresses ROS induced by pathogen-associated molecular pattern to inhibit pattern-triggered immunity (PTI)-induced cell death. The identification and molecular mechanism analysis of the Al6 candidate effector in A. lucorum will provide new insight into the molecular mechanisms of insect-plant interactions.

Reverse genetics reveals contrary effects of two Rdl-homologous GABA receptors of Helicoverpa armigera on the toxicity of cyclodiene insecticides.

The resistance to dieldrin gene (Rdl) encodes a subunit of the insect γ-amino butyric acid (GABA) receptor, and the encoded Rdl subunit is a major target site for cyclodiene and phenylpyrazole insecticides. Since the substitution of a single amino acid (Ala to Ser/Gly at position 302) of the Drosophila melanogaster Rdl gene was first identified to confer high level resistance to dieldrin, mutations at the equivalent positions have been reported to confer resistance to dieldrin and/or fipronil in a wide range of different insects. In the cotton bollworm Helicoverpa armigera, there are two Rdl homologs (HaRdl-1 and HaRdl-2) in close proximity on the Z chromosome, which as wild-type sequences, encode alanine and serine respectively at amino acid position 302. In the present study, we used the CRISPR/Cas9 gene editing approach to knock out HaRdl-1 and HaRdl-2 and establish two homozygous knockout strains (ΔRdl-1 and ΔRdl-2). The ΔRdl-1 strain showed low levels of resistance (8.0- to 9.3-fold) to three cyclodiene insecticides (endosulfan, aldrin and dieldrin) compared with the background SCD strain. In contrast, toxicity of the three cyclodiene insecticides to the ΔRdl-2 strain increased significantly (3.6- to 6.3-fold) when compared with the SCD strain. Genetic analysis indicated the obtained resistance to endosulfan and dieldrin in the ΔRdl-1 strain was sex-linked, which is consistent with the fact that HaRdl-1 locus is located on the Z chromosome. The above results demonstrate that both HaRdl-1 and HaRdl-2 are important determinants for the susceptibility of H. armigera SCD strain to the three cyclodiene insecticides, but have opposite effects. It was also found that HaRdl-1 and HaRdl-2 are involved, to some extent, in mediating sensitivity of H. armigera to avermectin and fipronil respectively. We speculate that the HaRdl-1 and HaRdl-2 subunits have different pharmacological properties, which contribute to the differential sensitivities of H. armigera to the tested cyclodienes and other insecticides.

The genetic association between osteoprotegerin gene polymorphisms and fracture risk in Chinese Han population.

This article put the genetic association exploration of osteoprotegerin (OPG) gene polymorphisms in promoter region (A-163G, T-245G) and fracture risk first and hoped to explain the ethology of fracture. The genotyping of OPG gene polymorphisms was conducted with the method of polymerase chain reaction-restriction fragment length polymorphism in 125 fracture patients and 138 relative controls. The genotype frequencies of selected controls based on OPG gene polymorphisms were checked by the χ2 test whether conformed to Hardy-Weinberg equilibrium (HWE). The relative risk was represented with odds ratio (OR) and 95% confidence interval (95% CI) between gene polymorphism and disease. The linkage disequilibrium (LD) and haplotype were also analyzed. The genotypes distributions of selected controls in OPG polymorphisms conformed to HWE. The G allele of A-163G polymorphism carriers had the tendency to suffer from fracture in the same condition, compared with A allele carriers (OR = 1.63, 95% CI = 1.04-2.55). TG and TG/GG genotypes of OPG T-245G polymorphism also showed the increased risk of fracture development, but not TT genotype (OR = 2.22, 95% CI = 1.15-4.28; OR = 2.45, 95% CI = 1.28-4.68). Likely, the mutant allele G had an abnormally higher frequency in cases than controls (14.00% and 6.16%). These two polymorphisms existed the LD and the haplotype G -163 -G -245 obviously increased the risk of fracture. OPG A-163G, T-245G polymorphisms were associated with the onset of fracture and both the independent risk factors.

Colorimetric determination of As(III) based on 3-mercaptopropionic acid assisted active site and interlayer channel dual-masking of Fe-Co-layered double hydroxides with oxidase-like activity.

A colorimetric method is described for the determination of As(III). It is based on the use of 3-mercaptopropionic acid (3-MPA) assisted active site and interlayer channel dual-masking of oxidase-like Fe-Co-layered double hydroxides (Fe-Co-LDH). The Fe-Co-LDH acts as an oxidase-mimicking nanozyme with high activity. It catalyzes the oxidation of colorless 3,3'5,5'-tetramethylbenzidine (TMB) to form a blue product (oxTMB) with an absorption maximum at 652 nm. It is found that As(III) firmly anchors onto the Fe* sites of the 3-MPA-modified Fe-Co-LDH via forming a stable Fe─As(III)─3-MPA─As(III)─Fe structure. This results in masking the active sites and interlayer channels of the Fe-Co-LDH nanozyme. As a result, the presence of As(III) as well as 3-MPA specifically inhibit the LDH-catalyzed chromogenic reaction. Based on the above principle, a colorimetric assay was designed for the determination of As(III). It provided linear response in the 0.10~8.33 µM As(III) concentration range and a detection limit as low as 35 nM. The assay was applied to the quantitation of As(III), even in the presence of potential interferents including As(V), Hg(II) and Pb(II), in environmental and drinking water samples. Graphical abstractSchematic illustration of the As(III) sensing mechanism based on 3-mercaptopropionic acid (3-MPA) assisted active site and interlayer channel dual-masking of Fe-Co-layered double hydroxides (Fe-Co-LDH) with oxidase-like activity. 3-MPA with sulfhydryl and carboxyl groups can assist As(III) to firmly anchor onto the Fe* sites inside the interlayer channels of the Fe-Co-LDH via forming a Fe─As(III)─3-MPA─As(III)─Fe structure, thus selectively resulting in a significant suppression of the chromogenic reaction.

Synthesis and biological evaluation of water-soluble derivatives of chiral gossypol as HIV fusion inhibitors targeting gp41.

A series of novel or known water-soluble derivatives of chiral gossypol were synthesized and screened in vitro for their anti-HIV-1 activity. (-)-gossypol derivative was more active against HIV-1 than the corresponding (+)-gossypol derivative, respectively. Among these derivatives, d-glucosamine derivative of (-)-gossypol, oligopeptide derivative of (-)-gossypol and taurine derivative of (-)-gossypol, such as compounds 1a, 3a and 14a, showed significant inhibitory activities against HIV-1 replication, HIV-1 mediated cell-cell fusion and HIV gp41 6-helix bundle formation as some amino acid derivatives of (-)-gossypol.

DYRK2 Negatively Regulates Type I Interferon Induction by Promoting TBK1 Degradation via Ser527 Phosphorylation.

Viral infection activates the transcription factors NF-κB and IRF3, which contribute to the induction of type I interferons (IFNs) and cellular antiviral responses. Protein kinases play a critical role in various signaling pathways by phosphorylating their substrates. Here, we identified dual-specificity tyrosine-(Y)-phosphorylation-regulated kinase 2 (DYRK2) as a negative regulator of virus-triggered type I IFN induction. DYRK2 inhibited the virus-triggered induction of type I IFNs and promoted the K48-linked ubiquitination and degradation of TANK-binding kinase 1 (TBK1) in a kinase-activity-dependent manner. We further found that DYRK2 phosphorylated Ser527 of TBK1, which is essential for the recruitment of NLRP4 and for the E3 ubiquitin ligase DTX4 to degrade TBK1. These findings suggest that DYRK2 negatively regulates virus-triggered signaling by targeting TBK1 for phosphorylation and priming it for degradation, and these data provide new insights into the molecular mechanisms that dictate the cellular antiviral response.

Dominant Inheritance of Field-Evolved Resistance to Fipronil in Plutella xylostella (Lepidoptera: Plutellidae).

A field-collected strain (HF) of Plutella xylostella (L.) showed 420-fold resistance to fipronil compared with a susceptible laboratory strain (Roth). The HF-R strain, derived from the HF strain by 25 generations of successive selection with fipronil in the laboratory, developed 2,200-fold resistance to fipronil relative to the Roth strain. The F(1) progeny of the reciprocal crosses between HF-R and Roth showed 640-fold (R♀ × S♂) and 1,380-fold (R♂ × S♀) resistance to fipronil, indicating resistance is inherited as an incompletely dominant trait. Analysis of progeny from a backcross (F1♂ × S♀) suggests that resistance is controlled by one major locus. The LC(50) of the R♂ × S♀ cross F(1) progeny is slightly but significantly higher than that of the R♀ × S♂ cross F(1) progeny, suggesting a minor resistance gene on the Z chromosome. Sequence analysis of PxGABARα1 (an Rdl-homologous GABA receptor gene of P. xylostella) from the HF-R strain identified two mutations A282S and A282G (corresponding to the A302S mutation of the Drosophila melanogaster Rdl gene), which have been previously implicated in fipronil resistance in several insect species including P. xylostella. PxGABARα1 was previously mapped to the Z chromosome of P. xylostella. In conclusion, fipronil resistance in the HF-R strain of P. xylostella was incompletely dominant, and controlled by a major autosomal locus and a sex-linked minor gene (PxGABARα1) on the Z chromosome.