Molecular mechanisms behind the generation of pro-oncogenic HIV-1 matrix protein p17 variants.

HIV-1 matrix protein p17 variants (vp17s), characterized by amino acid insertions at the COOH-terminal region of the viral protein, have been recently identified and studied for their biological activity. Different from their wild-type counterpart (refp17), vp17s display a potent B cell growth and clonogenic activity. Recent data have highlighted the higher prevalence of vp17s in people living with HIV-1 (PLWH) with lymphoma compared with those without lymphoma, suggesting that vp17s may play a key role in lymphomagenesis. Molecular mechanisms involved in vp17 development are still unknown. Here we assessed the efficiency of HIV-1 Reverse Transcriptase (RT) in processing this genomic region and highlighted the existence of hot spots of mutation in Gag, at the end of the matrix protein and close to the matrix-capsid junction. This is possibly due to the presence of inverted repeats and palindromic sequences together with a high content of Adenine in the 322-342 nucleotide portion, which constrain HIV-1 RT to pause on the template. To define the recombinogenic properties of hot spots of mutation in the matrix gene, we developed plasmid vectors expressing Gag and a minimally modified Gag variant, and measured homologous recombination following cell co-nucleofection by next-generation sequencing. Data obtained allowed us to show that a wide range of recombination events occur in concomitance with the identified hot spots of mutation and that imperfect events may account for vp17s generation.

Modelling evolutionary tempo and mode using formal morphological spaces and Markov chain principles.

Evolutionary tempo and mode summarize ancient and controversial subjects of theoretical biology such as gradualism, convergence, contingence, trends, and entrenchment. We employed an integrative methodological approach to explore the evolutionary tempo and mode of Lepidosaurian phalangeal formulae (PFs). This approach involves quantifying the frequencies of morphological changes along an evolutionary trajectory. The five meristic characters encoded by PFs are particularly valuable in revealing evolutionary patterns, owing to their discrete nature and extensive documentation in the literature. Based on a pre-existing dataset of PFs from 649 taxa (35 Lepidosauria families, including fossils), from which there exists a unique repertoire of 53 formulations, our approach simultaneously considers phenetic and phylogenetic data. This culminates in a diagram accounting for the phylogenetic dynamic of evolution traversing across different regions of morphospace. The method involves enumerating phenotypical options, reconstructing phenotypes across the phylogeny, projecting phenotypes onto a morphospace, and constructing a flow network from the frequency of evolutionary transitions between unique phenotypic conditions. This approach links Markovian chains and evolutionary trajectories to formally define parameters that describe the underlying transitions of morphological change. Among other results, we found that (a) PF evolution exhibits a clear trend towards reduction in the phalangeal count and that (b) evolutionary change tends to occur significantly between morphologically similar PFs. Notwithstanding, although minor but not trivial, transitions between distant formulas -jumps- occur. Our results support a pluralistic view including stasis, gradualism, and saltationism discriminating their prevalence in a target character evolution.

Evaluating virulence features of Acinetobacter baumannii resistant to polymyxin B.

The increasing resistance to polymyxins in Acinetobacter baumannii has made it even more urgent to develop new treatments. Anti-virulence compounds have been researched as a new solution. Here, we evaluated the modification of virulence features of A. baumannii after acquiring resistance to polymyxin B. The results showed lineages attaining unstable resistance to polymyxin B, except for Ab7 (A. baumannii polymyxin B resistant lineage), which showed stable resistance without an associated fitness cost. Analysis of virulence by a murine sepsis model indicated diminished virulence in Ab7 (A. baumannii polymyxin B resistant lineage) compared with Ab0 (A. baumannii polymyxin B susceptible lineage). Similarly, downregulation of virulence genes was observed by qPCR at 1 and 3 h of growth. However, an increase in bauE, abaI, and pgAB expression was observed after 6 h of growth. Comparison analysis of Ab0, Ab7, and Pseudomonas aeruginosa suggested no biofilm formation by Ab7. In general, although a decrease in virulence was observed in Ab7 when compared with Ab0, some virulence feature that enables infection could be maintained. In light of this, virulence genes bauE, abaI, and pgAB showed a potential relevance in the maintenance of virulence in polymyxin B-resistant strains, making them promising anti-virulence targets.

Evolution toward beta common chain receptor usage links the matrix proteins of HIV-1 and its ancestors to human erythropoietin.

The HIV-1 matrix protein p17 (p17) is a pleiotropic molecule impacting on different cell types. Its interaction with many cellular proteins underlines the importance of the viral protein as a major determinant of human specific adaptation. We previously showed the proangiogenic capability of p17. Here, by integrating functional analysis and receptor binding, we identify a functional epitope that displays molecular mimicry with human erythropoietin (EPO) and promotes angiogenesis through common beta chain receptor (ßCR) activation. The functional EPO-like epitope was found to be present in the matrix protein of HIV-1 ancestors SIV originated in chimpanzees (SIVcpz) and gorillas (SIVgor) but not in that of HIV-2 and its ancestor SIVsmm from sooty mangabeys. According to biological data, evolution of the EPO-like epitope showed a clear differentiation between HIV-1/SIVcpz-gor and HIV-2/SIVsmm branches, thus highlighting this epitope on p17 as a divergent signature discriminating HIV-1 and HIV-2 ancestors. P17 is known to enhance HIV-1 replication. Similarly to other ßCR ligands, p17 is capable of attracting and activating HIV-1 target cells and promoting a proinflammatory microenvironment. Thus, it is tempting to speculate that acquisition of an epitope on the matrix proteins of HIV-1 ancestors capable of triggering ßCR may have represented a critical step to enhance viral aggressiveness and early human-to-human SIVcpz/gor dissemination. The hypothesis that the p17/ßCR interaction and ßCR abnormal stimulation may also play a role in sustaining chronic activation and inflammation, thus marking the difference between HIV-1 and HIV-2 in term of pathogenicity, needs further investigation.

Spatiotemporal evolution and driving factors of ecosystem service bundle based on multi-scenario simulation in Beibu Gulf urban agglomeration, China.

Rapid urbanization is profoundly impacting the ecological environment and landscape patterns, leading to a decline in ecosystem services (ES) and posing threats to both ecological security and human well-being. This study aimed to identify the spatial and temporal patterns of ecosystem service bundles (ESB) in the Beibu Gulf urban agglomeration from 2000 to 2030, analyze the trajectory of ESB evolution, and elucidate the drivers behind ESB formation and evolution. We utilized the Patch-generating Land Use Simulation (PLUS) model to establish baseline (BLS), carbon sequestration priority (CPS), and urbanization priority (UPS) scenarios for simulating land use patterns in 2030. Following the assessment of ecosystem service values (ESV) through the equivalent factor method, we identified the spatiotemporal distribution patterns of ESB using the K-means clustering algorithm. By employing stability mapping and landscape indices, we identified and analyzed various types of ESB evolutionary trajectories. Redundancy analysis (RDA) was employed to pinpoint the drivers of ESB formation and evolution. The results revealed that from 2000 to 2030, land use changes were primarily observed in cropland, forestland, and construction land. Between 2000 and 2020, 92.88% of the region did not experience shifts in ESB types. In UPS, the ESB pattern in the study area underwent significant changes, with only 76.68% of the region exhibiting stabilized trajectories, while the other two scenarios recorded percentages higher than 80%. Key drivers of ESB-type shifts included initial food provision services, elevation, slope, changes in the proportion of construction land, and population change. This multi-scenario simulation of ESB evolution due to land use changes aids in comprehending potential future development directions from diverse perspectives and serves as a valuable reference for formulating and changing ecological management policies and strategies.

Comprehensive single-cell transcriptomic and proteomic analysis reveals NK cell exhaustion and unique tumor cell evolutionary trajectory in non-keratinizing nasopharyngeal carcinoma.

BACKGROUND: Nonkeratinizing nasopharyngeal carcinoma (NK-NPC) has a strong association with Epstein-Barr virus (EBV) infection. The role of NK cells and the tumor cell evolutionary trajectory in NK-NPC remain unclear. In this study, we aim to investigate the function of NK cell and the evolutionary trajectory of tumor cells in NK-NPC by single-cell transcriptomic analysis, proteomics and immunohistochemistry. METHODS: NK-NPC (n = 3) and normal nasopharyngeal mucosa cases (n = 3) were collected for proteomic analysis. Single-cell transcriptomic data of NK-NPC (n = 10) and nasopharyngeal lymphatic hyperplasia (NLH, n = 3) were obtained from Gene Expression Omnibus (GSE162025 and GSE150825). Quality control, dimension reduction and clustering were based on Seurat software (v4.0.2) process and batch effects were removed by harmony (v0.1.1) software. Normal cells of nasopharyngeal mucosa and tumor cells of NK-NPC were identified using copykat software (v1.0.8). Cell-cell interactions were explored using CellChat software (v1.4.0). Tumor cell evolutionary trajectory analysis was performed using SCORPIUS software (v1.0.8). Protein and gene function enrichment analyses were performed using clusterProfiler software (v4.2.2). RESULTS: A total of 161 differentially expressed proteins were obtained between NK-NPC (n = 3) and normal nasopharyngeal mucosa (n = 3) by proteomics (log2 fold change > 0.5 and P value < 0.05). Most of proteins associated with the nature killer cell mediated cytotoxicity pathway were downregulated in the NK-NPC group. In single cell transcriptomics, we identified three NK cell subsets (NK1-3), among which NK cell exhaustion was identified in the NK3 subset with high ZNF683 expression (a signature of tissue-resident NK cell) in NK-NPC. We demonstrated the presence of this ZNF683 + NK cell subset in NK-NPC but not in NLH. We also performed immunohistochemical experiments with TIGIT and LAG3 to confirm NK cell exhaustion in NK-NPC. Moreover, the trajectory analysis revealed that the evolutionary trajectory of NK-NPC tumor cells was associated with the status of EBV infection (active or latent). The analysis of cell-cell interactions uncovered a complex network of cellular interactions in NK-NPC. CONCLUSIONS: This study revealed that the NK cell exhaustion might be induced by upregulation of inhibitory receptors on the surface of NK cells in NK-NPC. Treatments for the reversal of NK cell exhaustion may be a promising strategy for NK-NPC. Meanwhile, we identified a unique evolutionary trajectory of tumor cells with active status of EBV-infection in NK-NPC for the first time. Our study may provide new immunotherapeutic targets and new sight of evolutionary trajectory involving tumor genesis, development and metastasis in NK-NPC.

A full repertoire of Hemiptera genomes reveals a multi-step evolutionary trajectory of auto-RNA editing site in insect Adar gene.

Adenosine-to-inosine (A-to-I) RNA editing, mediated by metazoan ADAR enzymes, is a prevalent post-transcriptional modification that diversifies the proteome and promotes adaptive evolution of organisms. The Drosophila Adar gene has an auto-recoding site (termed S>G site) that forms a negative-feedback loop and stabilizes the global editing activity. However, the evolutionary trajectory of Adar S>G site in many other insects remains largely unknown, preventing us from a deeper understanding on the significance of this auto-editing mechanism. In this study, we retrieved the well-annotated genomes of 375 arthropod species including the five major insect orders (Lepidoptera, Diptera, Coleoptera, Hymenoptera and Hemiptera) and several outgroup species. We performed comparative genomic analysis on the Adar auto-recoding S>G site. We found that the ancestral state of insect S>G site was an uneditable serine codon (unSer) and that this state was largely maintained in Hymenoptera. The editable serine codon (edSer) appeared in the common ancestor of Lepidoptera, Diptera and Coleoptera and was almost fixed in the three orders. Interestingly, Hemiptera species possessed comparable numbers of unSer and edSer codons, and a few 'intermediate codons', demonstrating a multi-step evolutionary trace from unSer-to-edSer with non-synchronized mutations at three codon positions. We argue that the evolution of Adar S>G site is the best genomic evidence supporting the 'proteomic diversifying hypothesis' of RNA editing. Our work deepens our understanding on the evolutionary significance of Adar auto-recoding site which stabilizes the global editing activity and controls transcriptomic diversity.

Characterization of the evolution trajectory and immune profiling of new histologic patterns in lung adenocarcinoma.

In lung adenocarcinoma (LUAD), the appearance of morphologically diverse tumor regions, termed histological patterns, is closely associated with disease progression and lymph node metastasis. However, the molecular characteristics of the histological patterns in LUAD and the underlying molecular evolutionary mechanisms between the histological patterns in primary tumors and lymph node metastases are poorly understood. Here, we re-analyzed the large TCGA-LUAD dataset and depicted a comprehensive profiling of the genome and transcriptome across the histological patterns in LUAD. Tumor phylogenetic trajectory analysis suggested that the complex glands is more apt to metastasize to the lymph node. Further deconvolution of the tumor microenvironment demonstrated that the complex glands had a higher infiltration of cancer-associated fibroblasts (CAFs). Single-cell transcriptome profiling of complex glands pattern identified a novel CAF subtype co-expressing fibroblast activation protein-alpha (FAP) and stimulator of interferon genes (STING). Moreover, our data demonstrated that FAP is an important downstream effector of STING in CAFs. In summary, our results provide the basis for the development of innovative therapeutic guidelines and intervention strategies for LUAD patients.

Molecular Epidemiology and Evolutionary Trajectory of Emerging Echovirus 30, Europe.

In 2018, an upsurge in echovirus 30 (E30) infections was reported in Europe. We conducted a large-scale epidemiologic and evolutionary study of 1,329 E30 strains collected in 22 countries in Europe during 2016-2018. Most E30 cases affected persons 0-4 years of age (29%) and 25-34 years of age (27%). Sequences were divided into 6 genetic clades (G1-G6). Most (53%) sequences belonged to G1, followed by G6 (23%), G2 (17%), G4 (4%), G3 (0.3%), and G5 (0.2%). Each clade encompassed unique individual recombinant forms; G1 and G4 displayed >2 unique recombinant forms. Rapid turnover of new clades and recombinant forms occurred over time. Clades G1 and G6 dominated in 2018, suggesting the E30 upsurge was caused by emergence of 2 distinct clades circulating in Europe. Investigation into the mechanisms behind the rapid turnover of E30 is crucial for clarifying the epidemiology and evolution of these enterovirus infections.

Landscape of cell heterogeneity and evolutionary trajectory in ulcerative colitis-associated colon cancer revealed by single-cell RNA sequencing.

OBJECTIVE: The goal of this study was to get preliminary insight on the intra-tumor heterogeneity in colitis-associated cancer (CAC) and to reveal a potential evolutionary trajectory from ulcerative colitis (UC) to CAC at the single-cell level. METHODS: Fresh samples of tumor tissues and adjacent UC tissues from a CAC patient with pT3N1M0 stage cancer were examined by single-cell RNA sequencing (scRNA-seq). Data from The Cancer Genome Atlas (TCGA) and The Human Protein Atlas were used to confirm the different expression levels in normal and tumor tissues and to determine their relationships with patient prognosis. RESULTS: Ultimately, 4,777 single-cell transcriptomes (1,220 genes per cell) were examined, of which 2,250 (47%) and 2,527 (53%) originated from tumor and adjacent UC tissues, respectively. We defined the composition of cancer-associated stromal cells and identified six cell clusters, including myeloid, T and B cells, fibroblasts, endothelial and epithelial cells. Notable pathways and transcription factors involved in these cell clusters were analyzed and described. Moreover, the precise cellular composition and developmental trajectory from UC to UC-associated colon cancer were graphed, and it was predicted that CD74, CLCA1, and DPEP1 played a potential role in disease progression. CONCLUSIONS: scRNA-seq technology revealed intra-tumor cell heterogeneity in UC-associated colon cancer, and might provide a promising direction to identify novel potential therapeutic targets in the evolution from UC to CAC.

Are developmental shifts the main driver of phenotypic evolution in Diplodus spp. (Perciformes: Sparidae)?

BACKGROUND: Sparid fishes of the genus Diplodus show a complex life history. Juveniles have adaptations well suited to life in the water column. When fishes recruit into the adult population, individuals develop a radically differentiated shape that reflects their adaptation to the new benthic environment typical of the adult. A comparative analysis of ontogenetic trajectories was performed to assess the presence of divergence in the developmental pattern. By using a geometric morphometric approach, we investigated the pattern of shape variation across ontogenetic stages that span from early settlement to the adult stage in four species of the genus Diplodus. Landmarks were collected on the whole body of fishes to quantify the phenotypic variation along two well defined life stages, i.e. juvenile and adult. A comparative analysis of ontogenetic trajectories was performed to assess the presence of divergence in the developmental pattern. Subsequently, we investigated the patterns of integration and modularity as proxy for the alteration of the developmental processes. This have allowed to give an insight in morphological developmental patterns across ecologically and ontogenetically differentiated life stages and to investigate the process leading to the adult shape. RESULT: Our results suggest that the origin of morphological novelties in Diplodus spp. arise from shifts of the ontogenetic trajectories during development. During the settlement phase, the juveniles' morphological shapes converge towards similar regions of the morphospace. When the four species approach the transition between settlement and recruitment, we observe the lowest level of inter- and intra-specific disparity. After this transition we detect an abrupt shift of ontogenetic trajectories, i.e. the path taken by species during development, that led to highly divergent adult phenotypes. DISCUSSION: We suggest that the evolution of new ecomorphologies, better suited to exploit different niches (pelagic vs. benthonic) and reduce inter-specific competition in Diplodus spp., are related to the shift in the ontogenetic trajectory that in turn is associated to changes in modularity and integration pattern.

Comprehensive analyses of the BES1 gene family in Brassica napus and examination of their evolutionary pattern in representative species.

BACKGROUND: The BES1 gene family, an important class of plant-specific transcription factors, play key roles in the BR signal pathway in plants, regulating various development processes. Until now, there has been no comprehensive analysis of the BES1 gene family in Brassica napus, and a cross-genome exploration of their origin, copy number changes, and functional innovation in plants was also not available. RESULTS: We identified 28 BES1 genes in B. napus from its two subgenomes (AA and CC). We found that 71.43% of them were duplicated in the tetraploidization, and their gene expression showed a prominent subgenome bias in the roots. Additionally, we identified 104 BES1 genes in another 18 representative angiosperms and performed a comparative analysis with B. napus, including evolutionary trajectory, gene duplication, positive selection, and expression pattern. Exploiting the available genome datasets, we performed a large-scale analysis across plants and algae suggested that the BES1 gene family could have originated from group F, expanding to form other groups (A to E) by duplicating or alternatively deleting some domains. We detected an additional domain containing M4 to M8 in exclusively groups F1 and F2. We found evidence that whole-genome duplication (WGD) contributed the most to the expansion of this gene family among examined dicots, while dispersed duplication contributed the most to expansion in certain monocots. Moreover, we inferred that positive selection might have occurred on major phylogenetic nodes during the evolution of plants. CONCLUSIONS: Grossly, a cross-genome comparative analysis of the BES1 genes in B. napus and other species sheds light on understanding its copy number expansion, natural selection, and functional innovation.

Evolution of cross-resistance to medical triazoles in Aspergillus fumigatus through selection pressure of environmental fungicides.

Resistance to medical triazoles in Aspergillus fumigatus is an emerging problem for patients at risk of aspergillus diseases. There are currently two presumed routes for medical triazole-resistance selection: (i) through selection pressure of medical triazoles when treating patients and (ii) through selection pressure from non-medical sterol-biosynthesis-inhibiting (SI) triazole fungicides which are used in the environment. Previous studies have suggested that SI fungicides can induce cross-resistance to medical triazoles. Therefore, to assess the potential of selection of resistance to medical triazoles in the environment, we assessed cross-resistance to three medical triazoles in lineages of A. fumigatus from previous work where we applied an experimental evolution approach with one of five different SI fungicides to select for resistance. In our evolved lines we found widespread cross-resistance indicating that resistance to medical triazoles rapidly arises through selection pressure of SI fungicides. All evolved lineages showed similar evolutionary dynamics to SI fungicides and medical triazoles, which suggests that the mutations inducing resistance to both SI fungicides and medical triazoles are likely to be the same. Whole-genome sequencing revealed that a variety of mutations were putatively involved in the resistance mechanism, some of which are in known target genes.

Arrested geomorphic trajectories and the long-term hidden potential for change.

Geomorphic systems often experience morphological changes that define a trajectory over decadal time periods. These trends can be halted by natural inhibitors such as vegetation, knickpoints, bed armor, or bank cohesion, or by anthropogenic inhibitors such as revetment, levees, or dams. Details about where and how channels and floodplains are stabilized are often poorly understood, which poses a risk that modern projects could unwittingly remove critical stabilizing elements (inhibitors) and unleash an episode of rapid change. The potential for destabilization is particularly keen for rivers that were severely altered by human activities but were stabilized by an inhibitor before readjustment was complete. This study uses aerial photographs to examine two cases of arrested geomorphic trajectories in the lower Yuba and Feather Rivers of northern California after 150 years of severe human disturbance. Channel adjustments were inhibited in distinctly different ways. First, channelization of the Feather River across a high-amplitude meander bend ∼4 km below the Yuba-Feather River confluence resulted in a knickpoint at Shanghai Shoals that retreated upstream at an average rate of 3.67 m/yr from 1963 to 2013 with two episodes of rapid retreat. Shanghai Shoals was breached in 2013. Second, numerous wing dams on the Yuba River constructed in the early nineteenth century limit floodplain widening and prevent return to an anastomosing channel planform. Their stabilizing role is important to preventing mobilization of mining sediment with high concentrations of mercury. These rivers exemplify how arrested geomorphic trajectories may impact sustainable river management, and how recognition of fluvial evolution is essential to sustainable river management.

Molecular evolution of anthocyanin pigmentation genes following losses of flower color.

BACKGROUND: Phenotypic transitions, such as trait gain or loss, are predicted to carry evolutionary consequences for the genes that control their development. For example, trait losses can result in molecular decay of the pathways underlying the trait. Focusing on the Iochrominae clade (Solanaceae), we examine how repeated losses of floral anthocyanin pigmentation associated with flower color transitions have affected the molecular evolution of three anthocyanin pathway genes (Chi, F3h, and Dfr). RESULTS: We recovered intact coding regions for the three genes in all of the lineages that have lost floral pigmentation, suggesting that molecular decay is not associated with these flower color transitions. However, two of the three genes (Chi, F3h) show significantly elevated dN/dS ratios in lineages without floral pigmentation. Maximum likelihood analyses suggest that this increase is due to relaxed constraint on anthocyanin genes in the unpigmented lineages as opposed to positive selection. Despite the increase, the values for dN/dS in both pigmented and unpigmented lineages were consistent overall with purifying selection acting on these loci. CONCLUSIONS: The broad conservation of anthocyanin pathway genes across lineages with and without floral anthocyanins is consistent with the growing consensus that losses of pigmentation are largely achieved by changes in gene expression as opposed to structural mutations. Moreover, this conservation maintains the potential for regain of flower color, and indicates that evolutionary losses of floral pigmentation may be readily reversible.

Evolutionary trajectory of diverse SARS-CoV-2 variants at the beginning of COVID-19 outbreak.

Despite extensive scientific efforts directed toward the evolutionary trajectory of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in humans at the beginning of the COVID-19 epidemic, it remains unclear how the virus jumped into and evolved in humans so far. Herein, we recruited almost all adult coronavirus disease 2019 (COVID-19) cases appeared locally or imported from abroad during the first 8 months of the outbreak in Shanghai. From these patients, SARS-CoV-2 genomes occupying the important phylogenetic positions in the virus phylogeny were recovered. Phylogenetic and mutational landscape analyses of viral genomes recovered here and those collected in and outside of China revealed that all known SARS-CoV-2 variants exhibited the evolutionary continuity despite the co-circulation of multiple lineages during the early period of the epidemic. Various mutations have driven the rapid SARS-CoV-2 diversification, and some of them favor its better adaptation and circulation in humans, which may have determined the waxing and waning of various lineages.

Exploring the evolutionary trajectory and functional landscape of cannabinoid receptors: A comprehensive bioinformatic analysis.

The bioinformatic analysis of cannabinoid receptors (CBRs) CB1 and CB2 reveals a detailed picture of their structure, evolution, and physiological significance within the endocannabinoid system (ECS). The study highlights the evolutionary conservation of these receptors evidenced by sequence alignments across diverse species including humans, amphibians, and fish. Both CBRs share a structural hallmark of seven transmembrane (TM) helices, characteristic of class A G-protein-coupled receptors (GPCRs), which are critical for their signalling functions. The study reports a similarity of 44.58 % between both CBR sequences, which suggests that while their evolutionary paths and physiological roles may differ, there is considerable conservation in their structures. Pathway databases like KEGG, Reactome, and WikiPathways were employed to determine the involvement of the receptors in various signalling pathways. The pathway analyses integrated within this study offer a detailed view of the CBRs interactions within a complex network of cannabinoid-related signalling pathways. High-resolution crystal structures (PDB ID: 5U09 for CB1 and 5ZTY for CB2) provided accurate structural information, showing the binding pocket volume and surface area of the receptors, essential for ligand interaction. The comparison between these receptors' natural sequences and their engineered pseudo-CBRs (p-CBRs) showed a high degree of sequence identity, confirming the validity of using p-CBRs in receptor-ligand interaction studies. This comprehensive analysis enhances the understanding of the structural and functional dynamics of cannabinoid receptors, highlighting their physiological roles and their potential as therapeutic targets within the ECS.

Contrasting macroevolutionary patterns in pelagic tetrapods across the Triassic-Jurassic transition.

The iconic marine raptorial predators Ichthyosauria and Eosauropterygia co-existed in the same ecosystems throughout most of the Mesozoic Era, facing similar evolutionary pressures and environmental perturbations. Both groups seemingly went through a massive macroevolutionary bottleneck across the Triassic-Jurassic (T/J) transition that greatly reduced their morphological diversity, leaving pelagic lineages as the only survivors. However, analyses of marine reptile disparity across the T/J transition have usually employed coarse morphological and temporal data. We comprehensively compare the evolution of ichthyosaurian and eosauropterygian morphology and body size across the Middle Triassic to Early Jurassic interval and find contrasting macroevolutionary patterns. The ecomorphospace of eosauropterygians predominantly reflects a strong phylogenetic signal, resulting in the clustering of three clades with clearly distinct craniodental phenotypes, suggesting 'leaps' towards novel feeding ecologies. Ichthyosaurian diversification lacks a discernible evolutionary trend, as we find evidence for a wide overlap of craniodental morphologies between Triassic and Early Jurassic forms. The temporal evolution of ecomorphological disparity, fin shape and body size of eosauropterygians and ichthyosaurians during the Late Triassic does not support the hypothesis of an abrupt macroevolutionary bottleneck near the T/J transition. Rather, an important turnover event should be sought earlier, during times of rapid sea level falls.

The S2 Pocket Governs the Genus-Specific Substrate Selectivity of Coronavirus 3C-Like Protease.

Coronavirus 3C-like protease (CoV 3CLpro) is essential for viral replication, providing an attractive target for monitoring the evolution of CoV and developing anti-CoV drugs. Here, the substrate-binding modes of 3CLpros from four CoV genera are analyzed and found that the S2 pocket in 3CLpro is highly conserved within each genus but differs between genera. Functionally, the S2 pocket, in conjunction with S4 and S1' pockets, governs the genus-specific substrate selectivity of 3CLpro. Resurrected ancestral 3CLpros from four CoV genera validate the genus-specific divergence of S2 pocket. Drawing upon the genus-specific S2 pocket as evolutionary marker, eight newly identified 3CLpros uncover the ancestral state of modern 3CLpro and elucidate the possible evolutionary process for CoV. It is also demonstrated that the S2 pocket is highly correlated with the genus-specific inhibitory potency of PF-07321332 (an FDA-approved drug against COVID-19) on different CoV 3CLpros. This study on 3CLpro provides novel insights to inform evolutionary mechanisms for CoV and develop genera-specific or broad-spectrum drugs against CoVs.

Phage-antibiotic synergy suppresses resistance emergence of Klebsiella pneumoniae by altering the evolutionary fitness.

Phage-antibiotic synergy (PAS) represents a superior treatment strategy for pathogen infections with less probability of resistance development. Here, we aim to understand the molecular mechanism by which PAS suppresses resistance in terms of population evolution. A novel hypervirulent Klebsiella pneumoniae (KP) phage H5 was genetically and structurally characterized. The combination of H5 and ceftazidime (CAZ) showed a robust synergistic effect in suppressing resistance emergence. Single-cell Raman analysis showed that the phage-CAZ combination suppressed bacterial metabolic activities, contrasting with the upregulation observed with phage alone. The altered population evolutionary trajectory was found to be responsible for the contrasting metabolic activities under different selective pressures, resulting in pleiotropic effects. A pre-existing wcaJ point mutation (wcaJG949A) was exclusively selected by H5, conferring a fitness advantage and up-regulated activity of carbohydrate metabolism, but also causing a trade-off between phage resistance and collateral sensitivity to CAZ. The wcaJ point mutation was counter-selected by H5-CAZ, inducing various mutations in galU that imposed evolutionary disadvantages with higher fitness costs, and suppressed carbohydrate metabolic activity. H5 and H5-CAZ treatments resulted in opposite effects on the transcriptional activity of the phosphotransferase system and the ascorbate and aldarate metabolism pathway, suggesting potential targets for phage resistance suppression. Our study reveals a novel mechanism of resistance suppression by PAS, highlighting how the complexity of bacterial adaptation to selective pressures drives treatment outcomes. IMPORTANCE: Phage-antibiotic synergy (PAS) has been recently proposed as a superior strategy for the treatment of multidrug-resistant pathogens to effectively reduce bacterial load and slow down both phage and antibiotic resistance. However, the underlying mechanisms of resistance suppression by PAS have been poorly and rarely been studied. In this study, we tried to understand how PAS suppresses the emergence of resistance using a hypervirulent Klebsiella pneumoniae (KP) strain and a novel phage H5 in combination with ceftazidime (CAZ) as a model. Our study reveals a novel mechanism by which PAS drives altered evolutionary trajectory of bacterial populations, leading to suppressed emergence of resistance. The findings advance our understanding of how PAS suppresses the emergence of resistance, and are imperative for optimizing the efficacy of phage-antibiotic therapy to further improve clinical outcomes.