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AbstractPurposeMethodsResultsConclusionIschemic retinopathy is the major cause of vision-threatening conditions. Inflammation plays an important role in the pathogenesis of ischemic retinopathy. Formyl peptide receptor 1 (FPR1) has been reported to be implicated in the regulation of inflammatory disorders. However, the role of FPR1 in the progression of ischemic retinal injury has not been fully explained.The activation of FPR1 was measured by real-time PCR and western blotting in the retina of OIR. The effect of FPR1 on the expression of inflammatory cytokines and relevant pro-angiogenic factors was assessed between wild-type and FPR1-deficiency OIR mice. The impact of FPR1 on retinal angiogenesis was evaluated through quantifying retinal vaso-obliteration and neovascularization between FPR1+/+ and FPR1–/– OIR mice. At last, the neuronal effect of FPR1 on the ischemic retina was investigated by ERG between wild-type and FPR1-deficient OIR mice.The expression of FPR1 significantly increased in the retina of OIR. Furthermore, FPR1 deficiency downregulated pro-inflammatory and pro-angiogenic factors. Ablation of FPR1 suppressed the retinal pathological neovascularization and promoted reparative revascularization, ultimately improving retinal neural function after ischemic injury.In ischemic retinopathy, FPR1 aggravates inflammation and inhibits reparative angiogenesis to exacerbate neuronal dysfunction. [ABSTRACT FROM AUTHOR]

Retinal ischemia is the main reason for vision threatening. Inflammation and aberrant angiogenesis play an important role in the pathogenesis of ischemia. Annexin A1 (ANXA1) is an endogenous protein modulating anti-inflammatory processes, and its therapeutic potential has been reported in a range of inflammatory diseases. However, the effect of ANXA1 on ischemic retinal injury has not been examined. Expression of ANXA1 was assessed by real-time PCR and western blotting, and location of ANXA1 was evaluated by immunofluorescence staining in retina of OIR. The activation of ANXA1 was assayed in HRECs after hypoxia stimuli. The effect of ANXA1 on vascularization of OIR mouse through quantification vaso-obliteration (VO) and neovascularization (NV), as well as expression of relevant angiogenic factors and inflammatory cytokines was compared between wild type and ANXA1 deficiency mice. We also investigated the effect of ANXA1 on retinal neuronal degeneration as measured by electroretinography (ERG) and OCT. In retinas of OIR, the expression of ANXA1 significantly increased and located in inner retinal layers. ANXA1 was induced in HRECs after hypoxic stimuli. Furthermore, ANXA1 deficiency increased pro-angiogenic and pro-inflammatory cytokines. Ablation of ANXA1 suppressed aortic outgrowth and retinal reparative revascularization and promoted pathological NV to exacerbate retinal dysfunction after ischemia injury. ANXA1 inhibits angiogenic and inhibits pro-inflammatory cytokines and promotes reparative angiogenesis, thus exhibits neuronal protective function in ischemic retinopathy. [ABSTRACT FROM AUTHOR]

Polymerase sliding clamps are ring‐shaped proteins that encircle duplex DNA and hold polymerases to DNA for high processivity during synthesis. The crystal structure of clamp‐DNA complex reveals that the DNA is highly tilted through the clamp with extensive interaction with the clamp inner surface. In contrast to the tilted clamp‐DNA interaction without DNA polymerases, recent structures of replicative polymerases of bacteria, eukaryotes, and archaea that are bound to the clamp and DNA show that the polymerase positions DNA straight through the clamp without direct protein‐DNA contacts. Instead, the clamp‐to‐DNA interaction is mediated by one or two layers of water. Hence, clamps 'water skate' on DNA during function with replicative polymerases from all domains of life, providing a nearly frictionless bearing for fast and processive DNA synthesis. [ABSTRACT FROM AUTHOR]

Pestivirus nonstructural protein 3 (NS3) is a multifunctional protein with protease and helicase activities that are essential for virus replication. In this study, we used a combination of biochemical and genetic approaches to investigate the relationship between a positively charged patch on the protease module and NS3 function. The surface patch is composed of four basic residues, R50, K74 and K94 in the NS3 protease domain and H24 in the structurally integrated cofactor NS4APCS. Single-residue or simultaneous four-residue substitutions in the patch to alanine or aspartic acid had little effect on ATPase activity. However, single substitutions of R50, K94 or H24 or a simultaneous four-residue substitution resulted in apparent changes in the helicase activity and RNA-binding ability of NS3. When these mutations were introduced into a classical swine fever virus (CSFV) cDNA clone, a single substitution at K94 or a simultaneous four-residue substitution (Qua_A or Qua_D) impaired the production of infectious virus. Furthermore, the replication efficiency of the CSFV variants was partially correlated with the helicase activity of NS3 invitro. Our results suggest that the conserved positively charged patch on NS3 plays an important role in modulating the NS3 helicase activity invitro and CSFV production. [ABSTRACT FROM AUTHOR]

• The recombinant chimeric CSFV/bE2 harboring heterologous BVDV E2 was generated by genetic approaches and serial passages of transfected cells. • The novel functional residues at positions 259 and 111 in CSFV E2 was identified. • The conserved residue G259 played a critical role in CSFV production and replication. • The residues G259 and V111 in the E2 modulated synergistically infectious CSFV production and replication. The E2 glycoprotein of classical swine fever virus (CSFV) plays multiple roles in the viral life cycle. The chimeric live attenuated C strain with the E2 substitution of bovine viral diarrhea virus (BVDV) is a promising marker vaccine candidate. In this study, the recombinant chimeric CSFV/bE2 cDNA clone harboring heterologous E2 (bE2) of BVDV was constructed by genetic approaches. Recombinant infectious virus rCSFV/bE2 (P11) was recovered by 11 serial passages of transfected PK15 cells. Viral genome sequencing showed that a glutamic acid to glycine mutation (E260G) at position 260 of the bE2 was observed in rCSFV/bE2 P11. Alignment of amino acid sequences displayed that the glycine was one of three conserved residues in pestivirus E2. When the glutamic acid to glycine substitution (E260G) was introduced into chimeric CSFV/bE2 cDNA clone, the high-titer infectious rCSFV/bE2 E260G was rescued. The glycine to glutamic acid substitution at corresponding position in CSFV E2 resulted in significantly decreased rCSFV/E2 G259E production. We further identified that the conserved E2 residue G259 played a critical role in the release and binding activity of CSFV and that the E2 residues G259 and V111 modulated synergistically infectious virus production and replication. [ABSTRACT FROM AUTHOR]

Rice paddies and rivers draining agricultural watersheds have been documented as two major sources of atmospheric methane (CH 4), while parallel flux measurements have been rarely taken on CH 4 from rice paddies against the drainage rivers in agricultural watersheds. Moreover, the drainage events during the rice growing season deliver sufficient organic and inorganic nutrients to rivers, which makes there an increasing concerned source of atmospheric CH 4. Here, we compared CH 4 emissions from rice paddies and the associated rivers draining agricultural watersheds by a parallel field experiment in southeast China. Over the whole rice–growing season (June–November), CH 4 fluxes from drainage rivers averaged 25 mg m–2 h–1, which were 82 % higher than those from rice paddies (13.7 mg m–2 h–1). Besides the dependence of riverine CH 4 fluxes on water dissolved oxygen (DO), both ecosystem CH 4 fluxes were significantly correlated with water/soil dissolved organic carbon (DOC), oxidation-reduction potential (Eh) and the microbial functional genes that drive CH 4 production or uptake [methanogens (mcrA and methanogenic archaeal 16S rRNA) and methanotrophs (pmoA)]. Our results highlighted that the CH 4 mitigation potential benefited from the drainage practice in rice paddies might have been totally offset by its induced increases in CH 4 emissions from rivers draining agricultural watersheds at the regional scale. ● Rivers draining agricultural watersheds are an important source of atmospheric CH 4. ● CH 4 fluxes from drainage rivers have been rarely related to the abundance of microbes. ● CH 4 fluxes from drainage rivers were 82 % greater than those from rice paddies. ● The increase in CH 4 emissions in rivers may offset the mitigation benefit of CH 4 in rice paddies. [ABSTRACT FROM AUTHOR]

Pestivirus nonstructural protein 2 (NS2) is a multifunctional, hydrophobic protein with an important but poorly understood role in viral RNA replication and infectious virus production. In the present study, based on sequence analysis, we mutated several representative conserved residues within the N-terminus of NS2 of classical swine fever virus (CSFV) and investigated how these mutations affected viral RNA replication and infectious virus production. Our results demonstrated that the mutation of two aspartic acids, NS2/D60A or NS2/D60K and NS2/D78K, in the N-terminus of NS2 abolished infectious virus production and that the substitution of arginine for alanine at position 100 (NS2/R100A) resulted in significantly decreased viral titer. The serial passage of cells containing viral genomic RNA molecules generated the revertants NS2/A60D, NS2/K60D and NS2/K78D, leading to the recovery of infectious virus. In the context of the NS2/R100A mutant, the NS2/I90L mutation compensated for infectious virus production. The regulatory roles of the indicated amino acid residues were identified to occur at the viral RNA replication level. These results revealed a novel function for the NS2 N-terminus of CSFV in modulating viral RNA replication. [ABSTRACT FROM AUTHOR]

Biochar amendment has been shown to reduce nitrous oxide (N 2 O) emissions from acidic soils in tea plantations. Given that both soil bacterial and fungal denitrifications can produce N 2 O, their relative contributions to the decrease in N 2 O emissions following biochar amendment remain unclear. Here, we examined soils sampled from a subtropical tea plantation that had received 2 years of biochar amendment. Measurements of the relative contributions of fungi and bacteria to N 2 O production were taken by the substrate-induced respiration method implemented with the addition of selective inhibitors. The abundances of total fungi, bacteria, and key N 2 O-related bacterial genes were quantified by q-PCR, and the composition of the fungal community was analyzed by 18S rRNA amplicon sequencing. The results showed that the contribution of fungi to N 2 O production (52%) was greater than that of bacteria (18%) for the N-applied acidic soils. Biochar amendment significantly decreased the fungal abundances and the fungal contribution to N 2 O production (by 28%). In contrast, biochar amendment significantly increased the abundances of N 2 O-related bacteria (e.g., ammonia-oxidizing bacteria (AOB), nirS , nosZ). Structural equation models (SEMs) revealed that biochar decreased the fungal contribution to N 2 O production through enhancing the soil pH and shifting the fungal community composition. Our results highlight that the decreased N 2 O emissions could be ascribed to the stimulated N 2 O consumption process (driven by N 2 O-consuming bacteria encoded by the nosZ gene) and suppressed fungal dominance in acidic soils from tea plantations. This study presents relatively comprehensive insights into the regulatory roles of biochar on soil microbe-mediated N 2 O production processes. • Fungi played a more important role than bacteria in N 2 O production from acidic soils. • Biochar shifted the fungal community on genus level from acidic soils. • Biochar decreased the fungal contribution to N 2 O production. [ABSTRACT FROM AUTHOR]

Freshwater aquaculture ponds constitute one of the important anthropogenic sources of atmospheric methane (CH 4). Nevertheless, estimates of global CH 4 emissions from freshwater aquaculture have large uncertainties due to a lack of data from different aquaculture types. Furthermore, despite that ebullition is a major pathway of CH 4 in aquatic systems, the quantification of ebullitive CH 4 fluxes from typical freshwater aquaculture ponds has been poorly represented. Here, field measurements of CH 4 fluxes over two years were taken to quantify ebullitive CH 4 fluxes from inland freshwater fish and crab aquaculture ponds in subtropical China. Ebullitive CH 4 fluxes averaged 15.97 ± 1.57 and 11.22 ± 1.26 mg m−2 d−1 in the fish and crab ponds in the first experimental year, respectively, and were 22.86 ± 2.30 and 21.95 ± 2.19 mg m−2 d−1 in the second year. During aquaculture period, ebullition dominated the emission pathways of CH 4 , accounting for 83% and 98% of the total CH 4 emissions in the fish and crab ponds, respectively. Ebullitive CH 4 fluxes exhibited considerable spatial variations, with the lowest flux rates captured at the aeration area due to aerator-use in both the fish and crab ponds. Dissolved oxygen and dissolved organic carbon were the two primary factors that drove ebullitive CH 4 fluxes in both aquaculture ponds. By incorporating global measurement data, we further assessed the CH 4 mitigation potential of aerator use in freshwater aquaculture and revealed the dominant role of ebullition in this mitigation contribution. Together with the rice-based aquaculture, aerator use could reduce CH 4 emissions from freshwater aquaculture ponds globally by 71% and in China by 63%. • Ebullition dominates the pathways of CH 4 emissions from inland aquaculture. • Ebullitive CH 4 fluxes differ with bioturbation to sediment in aquaculture ponds. • Aerator use could reduce global inland aquaculture CH 4 emissions by 71%. • Specific area aerator use benefits the whole aquaculture pond CH 4 mitigation. [ABSTRACT FROM AUTHOR]