Nat Methods. 2023 Apr 10. doi: 10.1038/s41592-023-01829-8. Online ahead of print.

ABSTRACT

Spatiotemporal regulation of the cellular transcriptome is crucial for proper protein expression and cellular function. However, the intricate subcellular dynamics of RNA remain obscured due to the limitations of existing transcriptomics methods. Here, we report TEMPOmap-a method that uncovers subcellular RNA profiles across time and space at the single-cell level. TEMPOmap integrates pulse-chase metabolic labeling with highly multiplexed three-dimensional in situ sequencing to simultaneously profile the age and location of individual RNA molecules. Using TEMPOmap, we constructed the subcellular RNA kinetic landscape in various human cells from transcription and translocation to degradation. Clustering analysis of RNA kinetic parameters across single cells revealed 'kinetic gene clusters' whose expression patterns were shaped by multistep kinetic sculpting. Importantly, these kinetic gene clusters are functionally segregated, suggesting that subcellular RNA kinetics are differentially regulated in a cell-state- and cell-type-dependent manner. Spatiotemporally resolved transcriptomics provides a gateway to uncovering new spatiotemporal gene regulation principles.

PMID:37038000 | DOI:10.1038/s41592-023-01829-8

Commun Biol. 2023 Apr 10;6(1):389. doi: 10.1038/s42003-023-04778-2.

ABSTRACT

Long-form collapsin response mediator protein-1 (LCRMP-1) belongs to the CRMP family which comprises brain-enriched proteins responsible for axon guidance. However, its role in spermatogenesis remains unclear. Here we find that LCRMP-1 is abundantly expressed in the testis. To characterize its physiological function, we generate LCRMP-1-deficient mice (Lcrmp-1-/-). These mice exhibit aberrant spermiation with apoptotic spermatids, oligospermia, and accumulation of immature testicular cells, contributing to reduced fertility. In the seminiferous epithelial cycle, LCRMP-1 expression pattern varies in a stage-dependent manner. LCRMP-1 is highly expressed in spermatids during spermatogenesis and especially localized to the spermiation machinery during spermiation. Mechanistically, LCRMP-1 deficiency causes disorganized F-actin due to unbalanced signaling of F-actin dynamics through upregulated PI3K-Akt-mTOR signaling. In conclusion, LCRMP-1 maintains spermatogenesis homeostasis by modulating cytoskeleton remodeling for spermatozoa release.

PMID:37037996 | DOI:10.1038/s42003-023-04778-2

Commun Biol. 2023 Apr 10;6(1):391. doi: 10.1038/s42003-023-04753-x.

ABSTRACT

Microbes live in diverse habitats (i.e. biomes), yet their species and genes were biome-specific, forming enrichment patterns. These enrichment patterns have mirrored the biome-species-function relationship, which is shaped by ecological and evolutionary principles. However, a grand picture of these enrichment patterns, as well as the roles of external and internal factors in driving these enrichment patterns, remain largely unexamined. In this work, we have examined the enrichment patterns based on 1705 microbiome samples from four representative biomes (Engineered, Gut, Freshwater, and Soil). Moreover, an "enrichment sphere" model was constructed to elucidate the regulatory principles behind these patterns. The driving factors for this model were revealed based on two case studies: (1) The copper-resistance genes were enriched in Soil biomes, owing to the copper contamination and horizontal gene transfer. (2) The flagellum-related genes were enriched in the Freshwater biome, due to high fluidity and vertical gene accumulation. Furthermore, this enrichment sphere model has valuable applications, such as in biome identification for metagenome samples, and in guiding 3D structure modeling of proteins. In summary, the enrichment sphere model aims towards creating a bluebook of the biome-species-function relationships and be applied in many fields.

PMID:37037946 | DOI:10.1038/s42003-023-04753-x

Nat Microbiol. 2023 Apr 10. doi: 10.1038/s41564-023-01346-6. Online ahead of print.

ABSTRACT

Previous urinary tract infections (UTIs) can predispose one to future infections; however, the underlying mechanisms affecting recurrence are poorly understood. We previously found that UTIs in mice cause differential bladder epithelial (urothelial) remodelling, depending on disease outcome, that impacts susceptibility to recurrent UTI. Here we compared urothelial stem cell (USC) lines isolated from mice with a history of either resolved or chronic uropathogenic Escherichia coli (UPEC) infection, elucidating evidence of molecular imprinting that involved epigenetic changes, including differences in chromatin accessibility, DNA methylation and histone modification. Epigenetic marks in USCs from chronically infected mice enhanced caspase-1-mediated cell death upon UPEC infection, promoting bacterial clearance. Increased Ptgs2os2 expression also occurred, potentially contributing to sustained cyclooxygenase-2 expression, bladder inflammation and mucosal wounding-responses associated with severe recurrent cystitis. Thus, UPEC infection acts as an epi-mutagen reprogramming the urothelial epigenome, leading to urothelial-intrinsic remodelling and training of the innate response to subsequent infection.

PMID:37037942 | DOI:10.1038/s41564-023-01346-6

Nat Biotechnol. 2023 Apr 10. doi: 10.1038/s41587-023-01733-8. Online ahead of print.

NO ABSTRACT

PMID:37037904 | DOI:10.1038/s41587-023-01733-8

Nat Biotechnol. 2023 Apr 10. doi: 10.1038/s41587-023-01749-0. Online ahead of print.

ABSTRACT

A kink-turn (K-turn) is a three-dimensional RNA structure that exists in all three primary phylogenetic domains. In this study, we developed the RIP-PEN-seq method to identify the full-length sequences of RNAs bound by the K-turn binding protein 15.5K and discovered a previously uncharacterized class of RNAs with backward K-turn motifs (bktRNAs) in humans and mice. All bktRNAs share two consensus sequence motifs at their fixed terminal position and have complex folding properties, expression and evolution patterns. We found that a highly conserved bktRNA1 guides the methyltransferase fibrillarin to install RNA methylation of U12 small nuclear RNA in humans. Depletion of bktRNA1 causes global splicing dysregulation of U12-type introns by impairing the recruitment of ZCRB1 to the minor spliceosome. Most bktRNAs regulate the splicing of local introns by interacting with the 15.5K protein. Taken together, our findings characterize a class of small RNAs and uncover another layer of gene expression regulation that involves crosstalk among bktRNAs, RNA splicing and RNA methylation.

PMID:37037902 | DOI:10.1038/s41587-023-01749-0

Nat Nanotechnol. 2023 Apr 10. doi: 10.1038/s41565-023-01328-z. Online ahead of print.

ABSTRACT

Expansion microscopy physically enlarges biological specimens to achieve nanoscale resolution using diffraction-limited microscopy systems1. However, optimal performance is usually reached using laser-based systems (for example, confocal microscopy), restricting its broad applicability in clinical pathology, as most centres have access only to light-emitting diode (LED)-based widefield systems. As a possible alternative, a computational method for image resolution enhancement, namely, super-resolution radial fluctuations (SRRF)2,3, has recently been developed. However, this method has not been explored in pathology specimens to date, because on its own, it does not achieve sufficient resolution for routine clinical use. Here, we report expansion-enhanced super-resolution radial fluctuations (ExSRRF), a simple, robust, scalable and accessible workflow that provides a resolution of up to 25 nm using LED-based widefield microscopy. ExSRRF enables molecular profiling of subcellular structures from archival formalin-fixed paraffin-embedded tissues in complex clinical and experimental specimens, including ischaemic, degenerative, neoplastic, genetic and immune-mediated disorders. Furthermore, as examples of its potential application to experimental and clinical pathology, we show that ExSRRF can be used to identify and quantify classical features of endoplasmic reticulum stress in the murine ischaemic kidney and diagnostic ultrastructural features in human kidney biopsies.

PMID:37037895 | DOI:10.1038/s41565-023-01328-z

NPJ Syst Biol Appl. 2023 Apr 10;9(1):12. doi: 10.1038/s41540-023-00272-x.

ABSTRACT

The immune system has to fight off hundreds of microbial invaders every day, such as the human-pathogenic fungus Aspergillus fumigatus. The fungal conidia can reach the lower respiratory tract, swell and form hyphae within six hours causing life-threatening invasive aspergillosis. Invading pathogens are continuously recognized and eliminated by alveolar macrophages (AM). Their number plays an essential role, but remains controversial with measurements varying by a factor greater than ten for the human lung. We here investigate the impact of the AM number on the clearance of A. fumigatus conidia in humans and mice using analytical and numerical modeling approaches. A three-dimensional to-scale hybrid agent-based model (hABM) of the human and murine alveolus allowed us to simulate millions of virtual infection scenarios, and to gain quantitative insights into the infection dynamics for varying AM numbers and infection doses. Since hABM simulations are computationally expensive, we derived and trained an analytical surrogate infection model on the large dataset of numerical simulations. This enables reducing the number of hABM simulations while still providing (i) accurate and immediate predictions on infection progression, (ii) quantitative hypotheses on the infection dynamics under healthy and immunocompromised conditions, and (iii) optimal AM numbers for combating A. fumigatus infections in humans and mice.

PMID:37037824 | DOI:10.1038/s41540-023-00272-x

Nat Commun. 2023 Apr 11;14(1):2021. doi: 10.1038/s41467-023-37730-3.

ABSTRACT

Chrysanthemum (Chrysanthemum morifolium Ramat.) is a globally important ornamental plant with great economic, cultural, and symbolic value. However, research on chrysanthemum is challenging due to its complex genetic background. Here, we report a near-complete assembly and annotation for C. morifolium comprising 27 pseudochromosomes (8.15 Gb; scaffold N50 of 303.69 Mb). Comparative and evolutionary analyses reveal a whole-genome triplication (WGT) event shared by Chrysanthemum species approximately 6 million years ago (Mya) and the possible lineage-specific polyploidization of C. morifolium approximately 3 Mya. Multilevel evidence suggests that C. morifolium is likely a segmental allopolyploid. Furthermore, a combination of genomics and transcriptomics approaches demonstrate the C. morifolium genome can be used to identify genes underlying key ornamental traits. Phylogenetic analysis of CmCCD4a traces the flower colour breeding history of cultivated chrysanthemum. Genomic resources generated from this study could help to accelerate chrysanthemum genetic improvement.

PMID:37037808 | DOI:10.1038/s41467-023-37730-3

Emerg Microbes Infect. 2023 Apr 10:2202263. doi: 10.1080/22221751.2023.2202263. Online ahead of print.

ABSTRACT

The adaptive immunity against SARS-CoV-2 prototype strain and Omicron sublineages induced by BA.1 breakthrough infection in vaccinees of inactivated COVID-19 vaccines have not been well characterized. Here, we report that BA.1 breakthrough infection induced mucosal sIgA and resulted in higher IgG titers against SARS-CoV-2 prototype strain and Omicron sublineages in vaccinees than in vaccine naïve-infected individuals. BA.1 breakthrough infection also boosted antibody-dependent cellular cytotoxicity and antibody-dependent cellular phagocytosis to SARS-CoV-2 prototype strain and BA.1, BA.1.1, BA.2, BA.2.12.1, and BA.2.75 but not to BA.4/5 and induced neutralizing antibody (NAb) responses against the prototype strain and BA.1, BA.1.1, BA.2, BA.2.12.1, BA.2.75, and BA.4/5 but not against BF.7, BQ.1, and XBB. In total, BA.1 breakthrough infection individuals produced less extensive sIgA, plasma IgG and NAb responses against Omicron sublineages compared with those against the prototype strain. Further, BA.1 breakthrough infection induced recall B cell response to the prototype strain and the Omicron variant, primarily targeting memory B cells producing conserved epitopes. Memory T cell responses against Omicron is largely preserved. Individuals with vaccine booster did not induce more beneficial immune responses to Omicron sublineages upon BA.1 breakthrough infection than those with primary vaccine doses only. The breakthrough infection individuals produced stronger antibody and T cell responses than those of inactivated vaccine-healthy individuals. These data have important implications for understanding the vaccine effectiveness and adapted immunity to breakthrough infection in individuals fully immunized with inactivated vaccines. The Omicron sublineages, especially for those emerged after BA.4/5 strain, evade the NAb responses induced by BA.1 breakthrough infection. It is urgent to optimize the vaccine immunogen design and formulations to SARS-CoV-2 variants.

PMID:37037791 | DOI:10.1080/22221751.2023.2202263

Genomics Inform. 2023 Mar;21(1):e6. doi: 10.5808/gi.22070. Epub 2023 Mar 31.

ABSTRACT

Glaucoma is the second leading cause of irreversible blindness, and primary open-angle glaucoma (POAG) is the most common type. Due to inadequate diagnosis, treatment is often not administered until symptoms occur. Hence, approaches enabling earlier prediction or diagnosis of POAG are necessary. We aimed to identify novel drugs for glaucoma through bioinformatics and network analysis. Data from 36 samples, obtained from the trabecular meshwork of healthy individuals and patients with POAG, were acquired from a dataset. Next, differentially expressed genes (DEGs) were identified to construct a protein-protein interaction (PPI) network. In both stages, the genes were enriched by studying the critical biological processes and pathways related to POAG. Finally, a drug-gene network was constructed, and novel drugs for POAG treatment were proposed. Genes with p < 0.01 and |log fold change| > 0.3 (1,350 genes) were considered DEGs and utilized to construct a PPI network. Enrichment analysis yielded several key pathways that were upregulated or downregulated. For example, extracellular matrix organization, the immune system, neutrophil degranulation, and cytokine signaling were upregulated among immune pathways, while signal transduction, the immune system, extracellular matrix organization, and receptor tyrosine kinase signaling were downregulated. Finally, novel drugs including metformin hydrochloride, ixazomib citrate, and cisplatin warrant further analysis of their potential roles in POAG treatment. The candidate drugs identified in this computational analysis require in vitro and in vivo validation to confirm their effectiveness in POAG treatment. This may pave the way for understanding life-threatening disorders such as cancer.

PMID:37037464 | DOI:10.5808/gi.22070

Clin Chim Acta. 2023 Apr 8:117335. doi: 10.1016/j.cca.2023.117335. Online ahead of print.

ABSTRACT

BACKGROUND: Muscle damage leads to increased serum creatine kinase (CK) levels in diseases such as acute myocardial infarction. Still, many individuals have abnormal serum CK activities lacking muscle-related diagnoses. The current study hypothesized that failed or overactivated CK clearance by non-muscle organs/tissues might be responsible for increased or decreased CK activities in blood.

METHODS: We analyzing 37,081 independent CK test results in 36 human diseases during the past 5 y.

RESULTS: We found that 33 out of 36 diseases were associated with decreased median CK activities compared to healthy controls. Besides muscle damage-related conditions, the highest mean CK activities were observed in hepatitis and cirrhosis. In contrast, 6 blood cell-related illnesses had the lowest mean CK values. ROC analysis showed that CK activities were the best biomarkers (AUC: 0.80-0.94) for the 6 blood-related diseases, especially myeloproliferative disorders. The principal component analysis revealed that the same category of diseases, such as liver-, blood -, kidney-, cancers, and vascular-related diseases, had clustered CK distributions.

CONCLUSIONS: We proposed that the liver and blood cells were mainly responsible for CK clearance in blood circulation based on overall results. The testable mechanisms were presented and discussed.

PMID:37037296 | DOI:10.1016/j.cca.2023.117335

Am J Pathol. 2023 Apr 8:S0002-9440(23)00123-2. doi: 10.1016/j.ajpath.2023.02.020. Online ahead of print.

ABSTRACT

Over 150,000 Americans are diagnosed with colorectal cancer (CRC) every year, and annually over 50,000 individuals will die from CRC, necessitating improvements in screening, prognostication, disease management, and therapeutic options. CRC tumors are removed en bloc with surrounding vasculature and lymphatics. Examination of regional lymph nodes at the time of surgical resection is essential for prognostication. Developing alternative approaches to indirectly assess recurrence risk would have utility in cases where lymph node yield is incomplete or inadequate. Spatially dependent, immune cell-specific (e.g., Tumor Infiltrating Lymphocytes- TILs), proteomic, and transcriptomic expression patterns inside and around the tumor - the tumor immune microenvironment (TIME) - can predict nodal/distant metastasis and probe the coordinated immune response from the primary tumor site. The comprehensive characterization of TILs and other immune infiltrates is possible using highly multiplexed spatial omics technologies, such as the GeoMX Digital Spatial Profiler (DSP). In this study, machine learning and differential co-expression analyses helped identify biomarkers from DSP-assayed protein expression patterns inside, at the invasive margin, and away from the tumor, associated with extracellular matrix remodeling (e.g., GZMB, fibronectin), immune suppression (e.g., FOXP3), exhaustion and cytotoxicity (e.g., CD8), PD-L1 expressing dendritic cells, neutrophil proliferation, amongst other concomitant alterations. Further investigation of these biomarkers may reveal independent risk factors of CRC metastasis that can be formulated into low-cost, widely available assays.

PMID:37037284 | DOI:10.1016/j.ajpath.2023.02.020

J Med Chem. 2023 Apr 10. doi: 10.1021/acs.jmedchem.2c01864. Online ahead of print.

ABSTRACT

Heterobifunctional degraders, known as proteolysis targeting chimeras (PROTACs), theoretically possess a catalytic mode-of-action, yet few studies have either confirmed or exploited this potential advantage of event-driven pharmacology. Degraders of oncogenic EML4-ALK fusions were developed by conjugating ALK inhibitors to cereblon ligands. Simultaneous optimization of pharmacology and compound properties using ternary complex modeling and physicochemical considerations yielded multiple catalytic degraders that were more resilient to clinically relevant ATP-binding site mutations than kinase inhibitor drugs. Our strategy culminated in the design of the orally bioavailable derivative CPD-1224 that avoided hemolysis (a feature of detergent-like PROTACs), degraded the otherwise recalcitrant mutant L1196M/G1202R in vivo, and commensurately slowed tumor growth, while the third generation ALK inhibitor drug lorlatinib had no effect. These results validate our original therapeutic hypothesis by exemplifying opportunities for catalytic degraders to proactively address binding site resistant mutations in cancer.

PMID:37036171 | DOI:10.1021/acs.jmedchem.2c01864

Plant J. 2023 Apr 10. doi: 10.1111/tpj.16237. Online ahead of print.

ABSTRACT

Chemical inhibitors are often implemented for the functional characterization of genes to overcome the limitations associated with genetic approaches. Although it is well established that the specificity of the compound is key to success of a pharmacological approach, off-target effects are often overlooked or simply neglected in a complex biological setting. Here we illustrate the cause and implications of such secondary effects by focusing on piperonylic acid (PA), an inhibitor of CINNAMATE-4-HYDROXYLASE (C4H) that is frequently used to investigate the involvement of lignin during plant growth and development. When supplied to plants, we found that PA is recognized as a substrate by GRETCHEN HAGEN 3.6 (GH3.6), an amido synthetase involved in the formation of the indole-3-acetic acid (IAA) conjugate IAA-Asp. By competing for the same enzyme, PA interferes with IAA conjugation, resulting in an increase in IAA concentrations in the plant. In line with the broad substrate specificity of the GH3 family of enzymes, treatment with PA increased not only IAA levels but also those of other GH3-conjugated phytohormones, namely jasmonic and salicylic acid. Finally, we found that interference with the endogenous function of GH3s potentially contributes to phenotypes previously observed upon PA-treatment. We conclude that deregulation of phytohormone homeostasis by surrogate occupation of the conjugation machinery in the plant is likely a general phenomenon when using chemical inhibitors. Our results hereby provide a novel and important basis for future reference in studies using chemical inhibitors.

PMID:37036146 | DOI:10.1111/tpj.16237

Adv Sci (Weinh). 2023 Apr 10:e2206789. doi: 10.1002/advs.202206789. Online ahead of print.

ABSTRACT

Acetaminophen (APAP) overdose is a leading cause of drug-induced liver injury and acute liver failure, while the detection, prognosis prediction, and therapy for APAP-induced liver injury (AILI) remain improved. Here, it is determined that the temporal pattern of circulating cell-free DNA (cfDNA) is strongly associated with damage and inflammation parameters in AILI. CfDNA is comparable to alanine aminotransferase (ALT) in predicting mortality and outperformed ALT when combined with ALT in AILI. The depletion of cfDNA or neutrophils alleviates liver damage, while the addition of cfDNA or adoptive transfer of neutrophils exacerbates the damage. The combination of DNase I and N-acetylcysteine attenuates AILI significantly. This study establishes that cfDNA is a mechanistic biomarker to predict mortality in AILI mice. The combination of scavenging cfDNA and reducing oxidative damage provides a promising treatment for AILI.

PMID:37035952 | DOI:10.1002/advs.202206789

Comput Struct Biotechnol J. 2023 Mar 23;21:2276-2285. doi: 10.1016/j.csbj.2023.03.035. eCollection 2023.

ABSTRACT

The RNA binding protein (RBP) and non-coding RNA (ncRNA) interacting networks are increasingly recognized as the main mechanism in gene regulation, and are tightly associated with cellular malfunction and disease. Here, we present fRNC, a systems biology tool to uncover the dynamic spectrum of RBP-ncRNA circuits (RNC) by integrating transcriptomics, interactomics and proteomics data. fRNC constructs the RBP-ncRNA network derived from CLIP-seq or PARE experiments. Given scoring on nodes and edges according to differential analysis of expression data, it finds an RNC containing global maximum significant RBPs and ncRNAs. Alternatively, it can also capture the locally maximum scoring RNC according to user-defined starting nodes with the greedy search. When compared with existing tools, fRNC can detect more accurate and robust sub-network with scalability. As shown in the cases of esophageal carcinoma, breast cancer and Alzheimer's disease, fRNC enables users to analyze the collective behaviors between RBP and the interacting ncRNAs, and reveal novel insights into the disease-associated processes. The fRNC R package is available at https://github.com/BioinformaticsSTU/fRNC.

PMID:37035550 | PMC:PMC10073992 | DOI:10.1016/j.csbj.2023.03.035

Front Bioinform. 2023 Mar 23;3:1125949. doi: 10.3389/fbinf.2023.1125949. eCollection 2023.

ABSTRACT

Cytoscape is an open-source bioinformatics environment for the analysis, integration, visualization, and query of biological networks. In this perspective piece, we describe our project to bring the Cytoscape desktop application to the web while explaining our strategy in ways relevant to others in the bioinformatics community. We examine opportunities and challenges in developing bioinformatics software that spans both the desktop and web, and we describe our ongoing efforts to build a Cytoscape web application, highlighting the principles that guide our development.

PMID:37035036 | PMC:PMC10076771 | DOI:10.3389/fbinf.2023.1125949

Food Energy Secur. 2022 Dec 2;12(1):e435. doi: 10.1002/fes3.435. eCollection 2023 Jan.

ABSTRACT

The growing world population and global increases in the standard of living both result in an increasing demand for food, feed and other plant-derived products. In the coming years, plant-based research will be among the major drivers ensuring food security and the expansion of the bio-based economy. Crop productivity is determined by several factors, including the available physical and agricultural resources, crop management, and the resource use efficiency, quality and intrinsic yield potential of the chosen crop. This review focuses on intrinsic yield potential, since understanding its determinants and their biological basis will allow to maximize the plant's potential in food and energy production. Yield potential is determined by a variety of complex traits that integrate strictly regulated processes and their underlying gene regulatory networks. Due to this inherent complexity, numerous potential targets have been identified that could be exploited to increase crop yield. These encompass diverse metabolic and physical processes at the cellular, organ and canopy level. We present an overview of some of the distinct biological processes considered to be crucial for yield determination that could further be exploited to improve future crop productivity.

PMID:37035025 | PMC:PMC10078444 | DOI:10.1002/fes3.435

iScience. 2023 Mar 13;26(4):106379. doi: 10.1016/j.isci.2023.106379. eCollection 2023 Apr 21.

ABSTRACT

Designing new functional motifs with unique properties is an important objective in the realm of synthetic biology. We uncover emergent properties of positive feedback loops (PFLs) under dual input signaling using pseudo potential energy-based high-throughput bifurcation analysis. We show that under dual signaling a single PFL generates a variety of noncanonical bistable switches, with one or more bistable regions, due to fusion of multiple canonical bistable switches. Regulatory signs of the dual signaling must be coherent for mutual inhibition loop and incoherent for mutual activation loop of the PFL. Occurrence probabilities show that some of the noncanonical switches, such as isola and mushroom, are highly recurrent under random parameterization. Phase diagrams of the noncanonical switches reveal that feedback strengths of the PFL control the transition from one switch to another. Our calculations decipher the design principles of noncanonical bistable switches that originate from synthetically feasible simple PFL motifs under dual signaling.

PMID:37034993 | PMC:PMC10074143 | DOI:10.1016/j.isci.2023.106379