GateView: A Multi-Omics Platform for Gene Feature Analysis of Virus Receptors within Human Normal Tissues and Tumors.

Viruses are obligate intracellular parasites that rely on cell surface receptor molecules to complete the first step of invading host cells. The experimental method for virus receptor screening is time-consuming, and receptor molecules have been identified for less than half of known viruses. This study collected known human viruses and their receptor molecules. Through bioinformatics analysis, common characteristics of virus receptor molecules (including sequence, expression, mutation, etc.) were obtained to study why these membrane proteins are more likely to become virus receptors. An in-depth analysis of the cataloged virus receptors revealed several noteworthy findings. Compared to other membrane proteins, human virus receptors generally exhibited higher expression levels and lower sequence conservation. These receptors were found in multiple tissues, with certain tissues and cell types displaying significantly higher expression levels. While most receptor molecules showed noticeable age-related variations in expression across different tissues, only a limited number of them exhibited gender-related differences in specific tissues. Interestingly, in contrast to normal tissues, virus receptors showed significant dysregulation in various types of tumors, particularly those associated with dsRNA and retrovirus receptors. Finally, GateView, a multi-omics platform, was established to analyze the gene features of virus receptors in human normal tissues and tumors. Serving as a valuable resource, it enables the exploration of common patterns among virus receptors and the investigation of virus tropism across different tissues, population preferences, virus pathogenicity, and oncolytic virus mechanisms.

Mechanism of complement inhibition by a mosquito protein revealed through cryo-EM.

Salivary complement inhibitors occur in many of the blood feeding arthropod species responsible for transmission of pathogens. During feeding, these inhibitors prevent the production of proinflammatory anaphylatoxins, which may interfere with feeding, and limit formation of the membrane attack complex which could damage arthropod gut tissues. Salivary inhibitors are, in many cases, novel proteins which may be pharmaceutically useful or display unusual mechanisms that could be exploited pharmaceutically. Albicin is a potent inhibitor of the alternative pathway of complement from the saliva of the malaria transmitting mosquito, Anopheles albimanus. Here we describe the cryo-EM structure of albicin bound to C3bBb, the alternative C3 convertase, a proteolytic complex that is responsible for cleavage of C3 and amplification of the complement response. Albicin is shown to induce dimerization of C3bBb, in a manner similar to the bacterial inhibitor SCIN, to form an inactive complex unable to bind the substrate C3. Size exclusion chromatography and structures determined after 30 minutes of incubation of C3b, factor B (FB), factor D (FD) and albicin indicate that FBb dissociates from the inhibited dimeric complex leaving a C3b-albicin dimeric complex which apparently decays more slowly.

A Novel Piecewise Cubic Hermite Interpolating Polynomial-Enhanced Convolutional Gated Recurrent Method under Multiple Sensor Feature Fusion for Tool Wear Prediction.

The monitoring of the lifetime of cutting tools often faces problems such as life data loss, drift, and distortion. The prediction of the lifetime in this situation is greatly compromised with respect to the accuracy. The recent rise of deep learning, such as Gated Recurrent Unit Units (GRUs), Hidden Markov Models (HMMs), Convolutional Neural Networks (CNNs), Recurrent Neural Networks (RNNs), Attention networks, and Transformers, has dramatically improved the data problems in tool lifetime prediction, substantially enhancing the accuracy of tool wear prediction. In this paper, we introduce a novel approach known as PCHIP-Enhanced ConvGRU (PECG), which leverages multiple-feature fusion for tool wear prediction. When compared to traditional models such as CNNs, the CNN Block, and GRUs, our method consistently outperformed them across all key performance metrics, with a primary focus on the accuracy. PECG addresses the challenge of missing tool wear measurement data in relation to sensor data. By employing PCHIP interpolation to fill in the gaps in the wear values, we have developed a model that combines the strengths of both CNNs and GRUs with data augmentation. The experimental results demonstrate that our proposed method achieved an exceptional relative accuracy of 0.8522, while also exhibiting a Pearson's Correlation Coefficient (PCC) exceeding 0.95. This innovative approach not only predicts tool wear with remarkable precision, but also offers enhanced stability.

Resolving phylogenetic relationships and taxonomic revision in the Pseudogastromyzon (Cypriniformes, Gastromyzonidae) genus: molecular and morphological evidence for a new genus, Labigastromyzon.

The Pseudogastromyzon genus, consisting of species predominantly distributed throughout southeastern China, has garnered increasing market attention in recent years due to its ornamental appeal. However, the overlapping diagnostic attributes render the commonly accepted criteria for interspecific identification unreliable, leaving the phylogenetic relationships among Pseudogastromyzon species unexplored. In the present study, we undertake molecular phylogenetic and morphological examinations of the Pseudogastromyzon genus. Our phylogenetic analysis of mitochondrial genes distinctly segregated Pseudogastromyzon species into two clades: the Pseudogastromyzon clade and the Labigastromyzon clade. A subsequent morphological assessment revealed that the primary dermal ridge (specifically, the second ridge) within the labial adhesive apparatus serves as an effective and precise interspecific diagnostic characteristic. Moreover, the distributional ranges of Pseudogastromyzon and Labigastromyzon are markedly distinct, exhibiting only a narrow area of overlap. Considering the morphological heterogeneity of the labial adhesive apparatus and the substantial division within the molecular phylogeny, we advocate for the elevation of the Labigastromyzon subgenus to the status of a separate genus. Consequently, we have ascertained the validity of the Pseudogastromyzon and Labigastromyzon species, yielding a total of six valid species. To facilitate future research, we present comprehensive descriptions of the redefined species and introduce novel identification keys.

Cryo-EM structures of anti-malarial antibody L9 with circumsporozoite protein reveal trimeric L9 association and complete 27-residue epitope.

Monoclonal antibody L9 recognizes the Plasmodium falciparum circumsporozoite protein (PfCSP) and is highly protective following controlled human malaria challenge. To gain insight into its function, we determined cryoelectron microscopy (cryo-EM) structures of L9 in complex with full-length PfCSP and assessed how this recognition influenced protection by wild-type and mutant L9s. Cryo-EM reconstructions at 3.6- and 3.7-Å resolution revealed L9 to recognize PfCSP as an atypical trimer. Each of the three L9s in the trimer directly recognized an Asn-Pro-Asn-Val (NPNV) tetrapeptide on PfCSP and interacted homotypically to facilitate L9-trimer assembly. We analyzed peptides containing different repeat tetrapeptides for binding to wild-type and mutant L9s to delineate epitope and homotypic components of L9 recognition; we found both components necessary for potent malaria protection. Last, we found the 27-residue stretch recognized by L9 to be highly conserved in P. falciparum isolates, suggesting the newly revealed complete L9 epitope to be an attractive vaccine target.

A bispecific inhibitor of complement and coagulation blocks activation in complementopathy models via a novel mechanism.

Inhibitors of complement and coagulation are present in the saliva of a variety of blood-feeding arthropods that transmit parasitic and viral pathogens. Here, we describe the structure and mechanism of action of the sand fly salivary protein lufaxin, which inhibits the formation of the central alternative C3 convertase (C3bBb) and inhibits coagulation factor Xa (fXa). Surface plasmon resonance experiments show that lufaxin stabilizes the binding of serine protease factor B (FB) to C3b but does not detectably bind either C3b or FB alone. The crystal structure of the inhibitor reveals a novel all ß-sheet fold containing 2 domains. A structure of the lufaxin-C3bB complex obtained via cryo-electron microscopy (EM) shows that lufaxin binds via its N-terminal domain at an interface containing elements of both C3b and FB. By occupying this spot, the inhibitor locks FB into a closed conformation in which proteolytic activation of FB by FD cannot occur. C3bB-bound lufaxin binds fXa at a separate site in its C-terminal domain. In the cryo-EM structure of a C3bB-lufaxin-fXa complex, the inhibitor binds to both targets simultaneously, and lufaxin inhibits fXa through substrate-like binding of a C-terminal peptide at the active site as well as other interactions in this region. Lufaxin inhibits complement activation in ex vivo models of atypical hemolytic uremic syndrome (aHUS) and paroxysmal nocturnal hemoglobinuria (PNH) as well as thrombin generation in plasma, providing a rationale for the development of a bispecific inhibitor to treat complement-related diseases in which thrombosis is a prominent manifestation.

tModBase: deciphering the landscape of tRNA modifications and their dynamic changes from epitranscriptome data.

tRNA molecules contain dense, abundant modifications that affect tRNA structure, stability, mRNA decoding and tsRNA formation. tRNA modifications and related enzymes are responsive to environmental cues and are associated with a range of physiological and pathological processes. However, there is a lack of resources that can be used to mine and analyse these dynamically changing tRNA modifications. In this study, we established tModBase (https://www.tmodbase.com/) for deciphering the landscape of tRNA modification profiles from epitranscriptome data. We analysed 103 datasets generated with second- and third-generation sequencing technologies and illustrated the misincorporation and termination signals of tRNA modification sites in ten species. We thus systematically demonstrate the modification profiles across different tissues/cell lines and summarize the characteristics of tRNA-associated human diseases. By integrating transcriptome data from 32 cancers, we developed novel tools for analysing the relationships between tRNA modifications and RNA modification enzymes, the expression of 1442 tRNA-derived small RNAs (tsRNAs), and 654 DNA variations. Our database will provide new insights into the features of tRNA modifications and the biological pathways in which they participate.

Alkaline State of the Domain-Swapped Dimer of Human Cytochrome c: A Conformational Switch for Apoptotic Peroxidase Activity.

A 2.08 Å structure of an alkaline conformer of the domain-swapped dimer of K72A human cytochrome c (Cytc) crystallized at pH 9.9 is presented. In the structure, Lys79 is ligated to the heme. All other domain-swapped dimer structures of Cytc have water bound to this coordination site. Part of Ω-loop D (residues 70-85) forms a flexible linker between the subunits in other Cytc domain-swapped dimer structures but instead converts to a helix in the alkaline conformer of the dimer combining with the C-terminal helix to form two 26-residue helices that bracket both sides of the dimer. The alkaline transition of the K72A human dimer monitored at both 625 nm (high spin heme) and 695 nm (Met80 ligation) yields midpoint pH values of 6.6 and 7.6, respectively, showing that the Met80 → Lys79 and high spin to low spin transitions are distinct. The dimer peroxidase activity increases rapidly below pH 7, suggesting that population of the high spin form of the heme is what promotes peroxidase activity. Comparison of the structures of the alkaline dimer and the neutral pH dimer shows that the neutral pH conformer has a better electrostatic surface for binding to a cardiolipin-containing membrane and provides better access for small molecules to the heme iron. Given that the pH of mitochondrial cristae ranges from 6.9 to 7.2, the alkaline transition of the Cytc dimer could provide a conformational switch to tune the peroxidase activity of Cytc that oxygenates cardiolipin in the early stages of apoptosis.

Effect on intrinsic peroxidase activity of substituting coevolved residues from Ω-loop C of human cytochrome c into yeast iso-1-cytochrome c.

Naturally-occurring variants of human cytochrome c (Cytc) that induce thrombocytopenia IV occur within Ω-loop C (residues 40-57). These variants enhance the peroxidase activity of human Cytc apparently by facilitating access to the heme by destabilizing Ω-loops C and D (residues 70-85). Given the importance of peroxidase activity in the early stages of apoptosis, we identified three sites with the EVmutation algorithm in or near Ω-loop C that coevolve and differ between yeast iso-1-Cytc and human Cytc. We prepared iso-1-Cytc variants with all possible combinations of the S40T, V57I and N63T substitutions to determine if these residues decrease the peroxidase activity of iso-1-Cytc to that of human Cytc producing an effective off state for a peroxidase signaling switch. At pH 6 and above, all variants significantly decreased peroxidase activity. However, the correlation of peroxidase activity with local and global stability, expected if cooperative unfolding of Ω-loops C and D is required for peroxidase activity, was generally poor. The m-values derived from the guanidine hydrochloride dependence of the kinetics of imidazole binding to horse Cytc, which is well-characterized by native-state hydrogen exchange methods, and K72A/K73A/K79A iso-1-Cytc show that local structural fluctuations and not subglobal cooperative unfolding of Ω-loops C and D are sufficient to permit binding of a small molecule like peroxide to the heme. A 2.46 Å structure of N63T iso-1-Cytc identifies a change to a hydrogen bond network linking Ω-loops C and D that could modulate the local fluctuations needed for the intrinsic peroxidase activity of Cytc.

The large bat Helitron DNA transposase forms a compact monomeric assembly that buries and protects its covalently bound 5'-transposon end.

Helitrons are widespread eukaryotic DNA transposons that have significantly contributed to genome variability and evolution, in part because of their distinctive, replicative rolling-circle mechanism, which often mobilizes adjacent genes. Although most eukaryotic transposases form oligomers and use RNase H-like domains to break and rejoin double-stranded DNA (dsDNA), Helitron transposases contain a single-stranded DNA (ssDNA)-specific HUH endonuclease domain. Here, we report the cryo-electron microscopy structure of a Helitron transposase bound to the 5'-transposon end, providing insight into its multidomain architecture and function. The monomeric transposase forms a tightly packed assembly that buries the covalently attached cleaved end, protecting it until the second end becomes available. The structure reveals unexpected architectural similarity to TraI, a bacterial relaxase that also catalyzes ssDNA movement. The HUH active site suggests how two juxtaposed tyrosines, a feature of many replication initiators that use HUH nucleases, couple the conformational shift of an α-helix to control strand cleavage and ligation reactions.

Naturally Occurring A51V Variant of Human Cytochrome c Destabilizes the Native State and Enhances Peroxidase Activity.

The A51V variant of human cytochrome c is linked to thrombocytopenia 4 (THC4), a condition that causes decreased blood platelet counts. A 1.82 Å structure of the A51V variant shows only minor changes in tertiary structure relative to the wild-type (WT) protein. Guanidine hydrochloride denaturation demonstrates that the global stability of the A51V variant is 1.3 kcal/mol less than that of the WT protein. The midpoint pH, pH1/2, of the alkaline transition of the A51V variant is 1 unit less than that of the WT protein. Stopped-flow pH jump experiments show that the A51V substitution affects the triggering ionization for one of two kinetically distinguishable alkaline conformers and enhances the accessibility of a high-spin heme transient. The pH1/2 for acid unfolding of the A51V variant is 0.7 units higher than for that of the WT protein. Consistent with the greater accessibility of non-native conformers for the A51V variant, the kcat values for its peroxidase activity increase by 6- to 15-fold in the pH range of 5-8 versus those of the WT protein. These data along with previously reported data for the other THC4-linked variants, G41S and Y48H, underscore the role of Ω-loop C (residues 40-57) in modulating the peroxidase activity of cytochrome c early in apoptosis.

Effect of V83G and I81A Substitutions to Human Cytochrome c on Acid Unfolding and Peroxidase Activity below a Neutral pH.

Mitochondrial cytochrome c is a highly conserved protein in eukaryotes. Certain functions of cytochrome c have been tuned during evolution. For instance, the intrinsic peroxidase activity of human cytochrome c is much lower than that of the yeast counterpart. Structural studies on K72A yeast iso-1-cytochrome c [McClelland, L. J., et al. (2014) Proc. Natl. Acad. Sci. USA, 111, 6648-6653] revealed that residues 81 and 83 in Ω-loop D (residues 70-85) may be gatekeeper residues for the peroxidase activity linked to intrinsic apoptosis. Amino acids at both positions evolve to more sterically demanding amino acids. We hypothesized that residues 81 and 83 evolved such that steric constraints at these positions tune down the peroxidase activity of human cytochrome c. To test this hypothesis, I81A and V83G variants of human cytochrome c were prepared. Our results show that the I81A substitution significantly influences the thermodynamics and kinetics of access to alternate conformers of human cytochrome c, while the V83G substitution has a more modest effect on these properties. The I81A variant also shows a significant enhancement in peroxidase activity, particularly below pH 7, whereas the V83G variant has a similar peroxidase activity to wild-type human cytochrome c. However, neither variant increases the peroxidase activity of human cytochrome c to the level of yeast iso-1-cytochrome c, indicating that other substructures of cytochrome c are also involved in tuning the intrinsic peroxidase activity of mitochondrial cytochrome c.

Humanlike substitutions to Ω-loop D of yeast iso-1-cytochrome c only modestly affect dynamics and peroxidase activity.

Structural studies of yeast iso-1-cytochrome c (L.J. McClelland, T.-C. Mou, M.E. Jeakins-Cooley, S.R. Sprang, B.E. Bowler, Proc. Natl. Acad. Sci. U.S.A. 111 (2014) 6648-6653) show that modest movement of Ω-loop D (residues 70-85, average RMSD versus the native structure: 0.81 Å) permits loss of Met80-heme ligation creating an available coordination site to catalyze the peroxidase activity mediated by cytochrome c early in apoptosis. However, Ala81 and Gly83 move significantly (RMSDs of 2.18 and 1.26 Å, respectively). Ala81 and Gly83 evolve to Ile and Val, respectively, in human cytochrome c and peroxidase activity decreases 25-fold relative to the yeast protein at pH 7. To test the hypothesis that these residues evolved to restrict the peroxidase activity of cytochrome c, A81I and G83V variants of yeast iso-1-cytochrome c were prepared. For both variants, the apparent pKa of the alkaline transition increases by 0.2 to 0.3 relative to the wild type (WT) protein and the rate of opening the heme crevice is slowed. The cooperativity of acid unfolding is decreased for the G83V variant. At pH 7 and 8, the catalytic rate constant, kcat, for the peroxidase activity of both variants decreases relative to WT, consistent with the effects on alkaline isomerization. Below pH 7, the loss in the cooperativity of acid unfolding causes kcat for peroxidase activity to increase for the G83V variant relative to WT. Neither variant decreases kcat to the level of the human protein, indicating that other residues also contribute to the low peroxidase activity of human cytochrome c.

Effect of a K72A Mutation on the Structure, Stability, Dynamics, and Peroxidase Activity of Human Cytochrome c.

We test the hypothesis that Lys72 suppresses the intrinsic peroxidase activity of human cytochrome c, as observed previously for yeast iso-1-cytochrome c [McClelland, L. J., et al. (2014) Proc. Natl. Acad. Sci. U. S. A. 111, 6648-6653]. A 1.25 Å X-ray structure of K72A human cytochrome c shows that the mutation minimally affects structure. Guanidine hydrochloride denaturation demonstrates that the K72A mutation increases global stability by 0.5 kcal/mol. The K72A mutation also increases the apparent pKa of the alkaline transition, a measure of the stability of the heme crevice, by 0.5 unit. Consistent with the increase in the apparent pKa, the rate of formation of the dominant alkaline conformer decreases, and this conformer is no longer stabilized by proline isomerization. Peroxidase activity measurements show that the K72A mutation increases kcat by 1.6-4-fold at pH 7-10, an effect larger than that seen for the yeast protein. X-ray structures of wild type and K72A human cytochrome c indicate that direct interactions of Lys72 with the far side of Ω-loop D, which are seen in X-ray structures of horse and yeast cytochrome c and could suppress peroxidase activity, are lacking. Instead, we propose that the stronger effect of the K72A mutation on the peroxidase activity of human versus yeast cytochrome c results from relief of steric interactions between the side chains at positions 72 and 81 (Ile in human vs Ala in yeast), which suppress the dynamics of Ω-loop D necessary for the intrinsic peroxidase activity of cytochrome c.