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Nuclear factor Y (NF-Y) gene family is an important transcription factor composed of three subfamilies of NF-YA, NF-YB and NF-YC, which is involved in plant growth, development and stress response. In this study, 63 tobacco NF-Y genes (NtNF-Ys) were identified in Nicotiana tabacum L., including 17 NtNF-YAs, 30 NtNF-YBs and 16 NtNF-YCs. Phylogenetic analysis revealed ten pairs of orthologues from tomato and tobacco and 25 pairs of paralogues from tobacco. The gene structure of NtNF-YAs exhibited similarities, whereas the gene structure of NtNF-YBs and NtNF-YCs displayed significant differences. The NtNF-Ys of the same subfamily exhibited a consistent distribution of motifs and protein 3D structure. The protein interaction network revealed that NtNF-YC12 and NtNF-YC5 exhibited the highest connectivity. Many cis-acting elements related to light, stress and hormone response were found in the promoter of NtNF-Ys. Transcriptome analysis showed that more than half of the NtNF-Y genes were expressed in all tissues, and NtNF-YB9/B14/B15/B16/B17/B29 were specifically expressed in roots. A total of 15, 12, 5, and 6 NtNF-Y genes were found to respond to cold, drought, salt, and alkali stresses, respectively. The results of this study will lay a foundation for further study of NF-Y genes in tobacco and other Solanaceae plants. [ABSTRACT FROM AUTHOR]

RAF-family kinases are activated by recruitment to the plasma membrane by GTP-bound RAS, whereupon they initiate signaling through the MAP kinase cascade. Prior structural studies of KRAS with RAF have focused on the isolated RAS-binding and cysteine-rich domains of RAF (RBD and CRD, respectively), which interact directly with RAS. Here we describe cryo-EM structures of a KRAS bound to intact BRAF in an autoinhibited state with MEK1 and a 14-3-3 dimer. Analysis of this KRAS/BRAF/MEK1/14-3-3 complex reveals KRAS bound to the RAS-binding domain of BRAF, captured in two orientations. Core autoinhibitory interactions in the complex are unperturbed by binding of KRAS and in vitro activation studies confirm that KRAS binding is insufficient to activate BRAF, absent membrane recruitment. These structures illustrate the separability of binding and activation of BRAF by RAS and suggest stabilization of this pre-activation intermediate as an alternative therapeutic strategy to blocking binding of KRAS. RAF kinases are autoinhibited in the cytosol and are activated by recruitment to the plasma membrane by GTP-bound RAS. Here, the authors describe cryoEM structures of a membrane-free KRAS/BRAF/MEK/14-3-3 complex that capture a pre-activation intermediate with BRAF still in an autoinhibited state. [ABSTRACT FROM AUTHOR]

The entry of SARS-CoV-2 into host cells depends on the refolding of the virus-encoded spike protein from a prefusion conformation, which is metastable after cleavage, to a lower-energy stable postfusion conformation1,2. This transition overcomes kinetic barriers for fusion of viral and target cell membranes3,4. Here we report a cryogenic electron microscopy (cryo-EM) structure of the intact postfusion spike in a lipid bilayer that represents the single-membrane product of the fusion reaction. The structure provides structural definition of the functionally critical membrane-interacting segments, including the fusion peptide and transmembrane anchor. The internal fusion peptide forms a hairpin-like wedge that spans almost the entire lipid bilayer and the transmembrane segment wraps around the fusion peptide at the last stage of membrane fusion. These results advance our understanding of the spike protein in a membrane environment and may guide development of intervention strategies.The SARS-CoV-2 spike internal fusion peptide forms a hairpin-like wedge that spans almost the entire lipid bilayer and the transmembrane segment wraps around the fusion peptide at the last stage of membrane fusion. [ABSTRACT FROM AUTHOR]

Inhibitors of triacylglycerol (TG) synthesis have been developed to treat metabolism-related diseases, but we know little about their mechanisms of action. Here, we report cryo-EM structures of the TG-synthesis enzyme acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1), a membrane bound O-acyltransferase (MBOAT), in complex with two different inhibitors, T863 and DGAT1IN1. Each inhibitor binds DGAT1's fatty acyl-CoA substrate binding tunnel that opens to the cytoplasmic side of the ER. T863 blocks access to the tunnel entrance, whereas DGAT1IN1 extends further into the enzyme, with an amide group interacting with more deeply buried catalytic residues. A survey of DGAT1 inhibitors revealed that this amide group may serve as a common pharmacophore for inhibition of MBOATs. The inhibitors were minimally active against the related MBOAT acyl-CoA:cholesterol acyltransferase 1 (ACAT1), yet a single-residue mutation sensitized ACAT1 for inhibition. Collectively, our studies provide a structural foundation for developing DGAT1 and other MBOAT inhibitors. Sui et al. elucidate the mechanisms of action for small molecule inhibitors of human triacylglycerol synthesis enzyme DGAT1 and provide insights into how these inhibitors achieve selectivity for DGAT1 rather than other closely related MBOAT enzymes. [ABSTRACT FROM AUTHOR]

Gasdermins (GSDMs) are pore-forming proteins that play critical roles in host defence through pyroptosis1,2. Among GSDMs, GSDMB is unique owing to its distinct lipid-binding profile and a lack of consensus on its pyroptotic potential3–7. Recently, GSDMB was shown to exhibit direct bactericidal activity through its pore-forming activity4. Shigella, an intracellular, human-adapted enteropathogen, evades this GSDMB-mediated host defence by secreting IpaH7.8, a virulence effector that triggers ubiquitination-dependent proteasomal degradation of GSDMB4. Here, we report the cryogenic electron microscopy structures of human GSDMB in complex with Shigella IpaH7.8 and the GSDMB pore. The structure of the GSDMB–IpaH7.8 complex identifies a motif of three negatively charged residues in GSDMB as the structural determinant recognized by IpaH7.8. Human, but not mouse, GSDMD contains this conserved motif, explaining the species specificity of IpaH7.8. The GSDMB pore structure shows the alternative splicing-regulated interdomain linker in GSDMB as a regulator of GSDMB pore formation. GSDMB isoforms with a canonical interdomain linker exhibit normal pyroptotic activity whereas other isoforms exhibit attenuated or no pyroptotic activity. Overall, this work sheds light on the molecular mechanisms of Shigella IpaH7.8 recognition and targeting of GSDMs and shows a structural determinant in GSDMB critical for its pyroptotic activity.The authors report the cryogenic electron microscopy structures of human GSDMB in complex with Shigella IpaH7.8 and the GSDMB pore, shedding light on the molecular mechanisms of Shigella IpaH7.8 recognition and targeting of GSDMs and GSDMB pore formation. [ABSTRACT FROM AUTHOR]

Lipoproteins are important for bacterial growth and antibiotic resistance. These proteins use lipid acyl chains attached to the N-terminal cysteine residue to anchor on the outer surface of cytoplasmic membrane. In Gram-negative bacteria, many lipoproteins are transported to the outer membrane (OM), a process dependent on the ATP-binding cassette (ABC) transporter LolCDE which extracts the OM-targeted lipoproteins from the cytoplasmic membrane. Lipid-anchored proteins pose a unique challenge for transport machinery as they have both hydrophobic lipid moieties and soluble protein component, and the underlying mechanism is poorly understood. Here we determined the cryo-EM structures of nanodisc-embedded LolCDE in the nucleotide-free and nucleotide-bound states at 3.8-Å and 3.5-Å resolution, respectively. The structural analyses, together with biochemical and mutagenesis studies, uncover how LolCDE recognizes its substrate by interacting with the lipid and N-terminal peptide moieties of the lipoprotein, and identify the amide-linked acyl chain as the key element for LolCDE interaction. Upon nucleotide binding, the transmembrane helices and the periplasmic domains of LolCDE undergo large-scale, asymmetric movements, resulting in extrusion of the captured lipoprotein. Comparison of LolCDE and MacB reveals the conserved mechanism of type VII ABC transporters and emphasizes the unique properties of LolCDE as a molecule extruder of triacylated lipoproteins. In Gram-negative bacteria, lipoproteins are transported from the inner membrane (IM) to the outer membrane (OM) by the ATP-binding cassette (ABC) transporter LolCDE. Here the authors present cryo-EM structures of nanodisc-embedded LolCDE in different states, providing mechanistic insight into the transport mechanism. [ABSTRACT FROM AUTHOR]

As an important material, titanium alloy is widely used in the manufacture of aircraft engine parts, and its processed surface quality is critical to the performance of aircraft engines. Abrasive belt grinding (ABG) is a kind of elastic grinding, which plays a significant role in improving titanium alloys' surface integrity. To validate the mathematical model's effectiveness from the grinding parameters to the surface residual stress after grinding, firstly, according to the molecular dynamics theory and ABG process, a physical model of titanium alloy ABG molecular system is proposed, and the embedded atom method is chosen as the interatomic potential of titanium alloy. Secondly, combined with the mathematical expression model of residual stress, the surface residual stress is characterized, and the heat correction coefficient is proposed to modify the mathematical model. Finally, based on molecular dynamics, the simulation of grinding residual stress and the grinding experiment is carried out for titanium alloy thin-walled parts. The results of simulation and experiment show that the trend of simulation results is similar to the experiment results. The simulation model can better represent the change rule of ABG surface residual stress for the titanium alloy material, but the average error rate is up to 15.01% due to the systematic error between the two. After correction, the average error rate between the simulation values and experiment values of residual stress on the surface decreases to 4.44%; the effectiveness of the mathematical model is verified. [ABSTRACT FROM AUTHOR]

The abrasive belt grinding process is a highly nonlinear dynamic change process. The coupling action mechanism between mechanical field and thermal field during grinding is complex and variable. In this paper, firstly, the abrasive belt grinding experiment of titanium alloy was carried out to measure the change of temperature and force during the grinding process, and the Savitzky-Golay filter analysis was carried out on force signal. Secondly, the generation of grinding heat and force was described in detail, the equilibrium process of thermal-mechanical coupling was analyzed, and the finite element equation of thermal-mechanical coupling was deduced for abrasive belt grinding. Thirdly, the finite element simulation model of abrasive belt grinding is established to comprehensively analyze the change rule of thermal-mechanical coupling in the grinding process. Finally, the experimental results and simulation results of belt grinding are discussed and analyzed. The experimental results found that the grinding force changes are divided into three different stages, and the maximum surface temperature decreases to a certain extent with the grinding process. The simulation results show that the plastic deformation zone and "dead metal zone" are consistent with the theory in the grinding process. This study can provide a reference for relevant machining and further improve the grinding workpiece's surface quality through thermal-mechanical optimization. [ABSTRACT FROM AUTHOR]

Objectives: This study aimed to analyse the clinical and laboratory characteristics of different pathologic classifications of lupus nephritis (LN) patients in terms of age at systemic lupus erythematosus (SLE) diagnosis and nephritis onset. Method: Clinical, laboratory, and pathological data of 710 LN patients diagnosed by renal biopsy at our institution between 2000 and 2018 were retrospectively analysed. Patients were divided into the different pathological classification groups; childhood-, adult- and elderly-onset SLE groups and early- and late-onset LN groups. Results: Class IV occurred most frequently and had the lowest complement C3 level. There was an obvious increase in active index in class IV and class V + IV. Patients with class VI showed some clinical characteristics similar to end-stage renal disease. Patients with proliferative nephritis were younger at SLE diagnosis and had higher blood pressure, higher frequency of proteinuria and urinary erythrocyte and lower haemoglobin and complement C3. Pathologic classification between childhood-, adult- and elderly-onset SLE patients or between early- and late-onset LN patients was not significantly different. Elderly-onset SLE patients had the highest chronic index (CI), IgA, IgG and Sjögren's syndrome A antibodies and Sjögren's syndrome B antibodies rates, whereas late-onset LN patients showed significantly higher CI, haemoglobin, complement C3 and C4 but lower uric acid, IgM and IgG. Conclusions: LN patients present with different clinical and laboratory characteristics according to pathological classification, age at SLE diagnosis and nephritis onset. These results might be valuable for estimating the pathology and guiding treatment and prognosis. Key Points • Patients with proliferative nephritis have more severe immune disorders, worse renal function and stronger inflammatory state. • The elderly-onset SLE patients showed a poorer condition. • The late-onset LN patients might have a more stable status. [ABSTRACT FROM AUTHOR]

These authors contributed equally: Eunyoung Park, Shaun Rawson. The correct version of the final column of the 2nd row states '21.2 (ref. 24) 356 (ref. 17)' instead of the original, incorrect '21.2 (ref. 24, 356 (ref. 17)'; the correct version of the final column of the 3rd row states '190.5 (ref. 24) 152 (ref. 17)' instead of the original, incorrect '190.5 (ref. 24, 152 (ref. 17)'.This has been corrected in both the PDF and HTML versions of the Article. Correction to: I Nature Communications i https://doi.org/10.1038/s41467-023-40299-6, published online 29 July 2023 The original version of this Article contained errors in Table 1. [Extracted from the article]

The generic Meanshift is susceptible to interference of background pixels with the target pixels in the kernel of the reference model, which compromises the tracking performance. In this paper, we enhance the target color feature by attenuating the background color within the kernel through enlarging the pixel weightings which map to the pixels on the target. This way, the background pixel interference is largely suppressed in the color histogram in the course of constructing the target reference model. In addition, the proposed method also reduces the number of Meanshift iterations, which speeds up the algorithmic convergence. The two tests validate the proposed approach with improved tracking robustness on real-world video sequences. [ABSTRACT FROM AUTHOR]