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The classification of the Uranoscopidae species is controversial and the Ichthyscopus pollicaris belonging to Uranoscopidae was first reported in 2019. In the present study, the whole genome sequence of I. pollicaris were generated by PacBio and Illumina platforms for the first time. After de novo assembly and correction of the high-quality PacBio data, a 527.25 Mb I. pollicaris genome with an N50 length of 11.25 Mb was finally generated. Meanwhile, 170.41 Mb repeating sequence, 21,263 genes, 784 miRNAs, 2,225 tRNAs, 3004 rRNAs, and 1422 snRNAs were annotated in I. pollicaris genome. Furthermore, 3,168 single-copy orthologous genes were applied to reconstructed the phylogenetic relationship between I. pollicaris and other 11 species. The draft genome sequences have been deposited in NCBI database with the accession number of PRJNA1071810.
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Protein arginylation is an essential posttranslational modification (PTM) catalyzed by arginyl-tRNA-protein transferase 1 (ATE1) in mammalian systems. Arginylation features a post-translational conjugation of an arginyl to a protein, making it extremely challenging to differentiate from translational arginine residues with the same mass in a protein sequence. Here we present a general activity-based arginylation profiling (ABAP) platform for the unbiased discovery of arginylation substrates and their precise modification sites. This method integrates isotopic arginine labeling into an ATE1 assay utilizing biological lysates (ex vivo) rather than live cells, thus eliminating translational bias derived from the ribosomal activity and enabling bona fide arginylation identification using isotopic features. ABAP has been successfully applied to an array of peptide, protein, cell, patient, and animal tissue samples using 20 µg sample input, with 229 unique arginylation sites revealed from human proteomes. Representative sites were validated and followed up for their biological functions. The developed platform is globally applicable to the aforementioned sample types and therefore paves the way for functional studies of this difficult-to-characterize protein modification.
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Glycans modify protein, lipid, and even RNA molecules to form the regulatory outer coat on cells called the glycocalyx. The changes in glycosylation have been linked to the initiation and progression of many diseases. Thus, while the significance of glycosylation is well established, a lack of accessible methods to characterize glycans has hindered the ability to understand their biological functions. Mass spectrometry (MS)-based methods have generally been at the core of most glycan profiling efforts; however, modern data-independent acquisition (DIA), which could increase sensitivity and simplify workflows, has not been benchmarked for analyzing glycans. Herein, we developed a DIA-based glycomic workflow, termed GlycanDIA, to identify and quantify glycans with high sensitivity and accuracy. The GlycanDIA workflow combined higher energy collisional dissociation (HCD)-MS/MS and staggered windows for glycomic analysis, which facilitates the sensitivity in identification and the accuracy in quantification compared to conventional data-dependent acquisition (DDA)-based glycomics. To facilitate its use, we also developed a generic search engine, GlycanDIA Finder, incorporating an iterative decoy searching for confident glycan identification and quantification from DIA data. The results showed that GlycanDIA can distinguish glycan composition and isomers from N-glycans, O-glycans, and human milk oligosaccharides (HMOs), while it also reveals information on low-abundant modified glycans. With the improved sensitivity, we performed experiments to profile N-glycans from RNA samples, which have been underrepresented due to their low abundance. Using this integrative workflow to unravel the N-glycan profile in cellular and tissue glycoRNA samples, we found that RNA-glycans have specific forms as compared to protein-glycans and are also tissue-specific differences, suggesting distinct functions in biological processes. Overall, GlycanDIA can provide comprehensive information for glycan identification and quantification, enabling researchers to obtain in-depth and refined details on the biological roles of glycosylation.
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Chemical proteomics utilizes small-molecule probes to covalently engage with their interacting proteins. Since chemical probes are tagged to the active or binding sites of functional proteins, chemical proteomics can be used to profile protein targets, reveal precise binding sites/mechanisms, and screen inhibitors competing with probes in a biological context. These capabilities of chemical proteomics have great potential to enable discoveries of both drug targets and lead compounds. However, chemical proteomics is limited by the time-consuming bottleneck of sample preparations, which are processed manually. With the advancement of robotics and artificial intelligence, it is now possible to automate workflows to make chemical proteomics sample preparation a high-throughput process. An automated robotic system represents a major technological opportunity to speed up advances in proteomics, open new frontiers in drug target discovery, and broaden the future of chemical biology.
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Inteligencia Artificial , Proteómica , Automatización , Proteínas/química , Descubrimiento de DrogasRESUMEN
The interest in MS-based analysis of modified nucleic acids is increasing due to the application of nucleic acids in therapeutics. However, there are few available integrated platforms for characterizing nucleic acid modifications. Herein, we report a general mass spectrometry-based SWATH platform to identify and quantify both RNA and DNA modifications, which we call SWATH analysis of modified nucleic acids (SWAMNA). SWAMNA incorporates the search engine, NuMo finder, enabling the analysis of modifications in native and permethylated form. SWAMNA will aid discoveries that provide new insights into nucleic acid modifications.
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Ácidos Nucleicos , ARN , Espectrometría de MasasRESUMEN
BACKGROUND: Hepatitis B virus (HBV) infection is one of the most common risk factors for intrahepatic cholangiocarcinoma (ICC). However, there is no direct evidence of a causal relationship between HBV infection and ICC. In this study, we attempted to prove that ICC may originate from hepatocytes through a pathological study involving ICC tissue-derived organoids. METHOD: The medical records and tumor tissue samples of 182 patients with ICC after hepatectomy were collected. The medical records of 182 patients with ICC were retrospectively analyzed to explore the prognostic factors. A microarray of 182 cases of ICC tumor tissue and 6 cases of normal liver tissue was made, and HBsAg was stained by immunohistochemistry (IHC) to explore the factors closely related to HBV infection. Fresh ICC tissues and corresponding adjacent tissues were collected to make paraffin sections and organoids. Immunofluorescence (IF) staining of factors including HBsAg, CK19, CK7, Hep-Par1 and Albumin (ALB) was performed on both fresh tissues and organoids. In addition, we collected adjacent nontumor tissues of 6 patients with HBV (+) ICC, from which biliary duct tissue and normal liver tissue were isolated and RNA was extracted respectively for quantitative PCR assay. In addition, the expression of HBV-DNA in organoid culture medium was detected by quantitative PCR and PCR electrophoresis. RESULTS: A total of 74 of 182 ICC patients were HBsAg positive (40.66%, 74/182). The disease-free survival (DFS) rate of HBsAg (+) ICC patients was significantly lower than that of HBsAg (-) ICC patients (p = 0.0137). IF and IHC showed that HBsAg staining was only visible in HBV (+) ICC fresh tissues and organoids, HBsAg expression was negative in bile duct cells in the portal area. Quantitative PCR assay has shown that the expression of HBs antigen and HBx in normal hepatocytes were significantly higher than that in bile duct epithelial cells. Combined with the IF and IHC staining, it was confirmed that HBV does not infect normal bile duct epithelial cells. In addition, IF also showed that the staining of bile duct markers CK19 and CK7 were only visible in ICC fresh tissue and organoids, and the staining of hepatocyte markers Hep-Par1 and ALB was only visible in normal liver tissue fresh tissue. Real-time PCR and WB had the same results. High levels of HBV-DNA were detected in the culture medium of HBV (+) organoids but not in the culture medium of HBV (-) organoids. CONCLUSION: HBV-related ICC might be derived from hepatocytes. HBV (+) ICC patients had shorter DFS than HBV (-) ICC patients.
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Neoplasias de los Conductos Biliares , Colangiocarcinoma , Hepatitis B , Humanos , Virus de la Hepatitis B/genética , Antígenos de Superficie de la Hepatitis B , Estudios Retrospectivos , ADN Viral , Receptor PAR-1 , Hepatitis B/complicaciones , Hepatocitos/patología , Conductos Biliares Intrahepáticos/patologíaRESUMEN
In this study, we determined the complete mitochondrial genome of Neolissochilus heterostomus. The genome is 16,585 bp in length, including 2 ribosomal RNA genes, 13 proteins-coding genes, 22 transfer RNA genes, and two non-coding control regions. Sequence analysis showed that the overall base composition of N. heterostomus is T 24.8%, C 27.7%, A 31.7%, and G 15.8%. The sequence is a slight A + T bias of 56.5%, which is similar to other fishes. We describe a phylogenetic analysis of 16 species of Cypriniformes based on the complete mitochondrial genome, and the result showed that N. stracheyi is most closely related to N. heterostomus. This mitogenome sequence data would play an important role in the investigation of phylogenetic relationship of the Cyprinidae.
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The terahertz (THz) metamaterial biosensor has great potential for label-free and rapid specificity testing. Here, we designed two highly sensitive structures to detect the carcinoembryonic antigen (CEA) of the cancer biomarker in early stages. There was about 29â GHz (500â ng/ml) resonance shift for CEA with an insert grate metamaterial, which was consistent with simulation results. Moreover, the concentration of CEA was gained through the relationship between the cancer marker concentration and frequency shift (Δƒ). Our design and detection methods may provide a potential route for the early warning stages of cancer.