RESUMEN
Binding-activated optical sensors are powerful tools for imaging, diagnostics, and biomolecular sensing. However, biosensor discovery is slow and requires tedious steps in rational design, screening, and characterization. Here we report on a platform that streamlines biosensor discovery and unlocks directed nanosensor evolution through genetically encodable fluorogenic amino acids (FgAAs). Building on the classical knowledge-based semisynthetic approach, we engineer ~15 kDa nanosensors that recognize specific proteins, peptides, and small molecules with up to 100-fold fluorescence increases and subsecond kinetics, allowing real-time and wash-free target sensing and live-cell bioimaging. An optimized genetic code expansion chemistry with FgAAs further enables rapid (~3 h) ribosomal nanosensor discovery via the cell-free translation of hundreds of candidates in parallel and directed nanosensor evolution with improved variant-specific sensitivities (up to ~250-fold) for SARS-CoV-2 antigens. Altogether, this platform could accelerate the discovery of fluorogenic nanosensors and pave the way to modify proteins with other non-standard functionalities for diverse applications.
Asunto(s)
Aminoácidos , Técnicas Biosensibles , Colorantes Fluorescentes , SARS-CoV-2 , Técnicas Biosensibles/métodos , Colorantes Fluorescentes/química , Humanos , SARS-CoV-2/genética , COVID-19/virología , Nanotecnología/métodos , Péptidos/metabolismo , Péptidos/química , Péptidos/genéticaRESUMEN
Polyglutamine (PolyQ) disease is a group of hereditary neurodegenerative diseases. It has become clear that brain damage may occur decades before the onset of symptoms. As a sensitive marker for neuro-axonal damages, neurofilament light chain (NfL) has appeared as a promising biomarker for neurological diseases. it may be used as a preclinical and clinical marker for the neurodegeneration in polyQ diseases, and is closely correlated with disease severity and progression, in particular different disease stages. This article has provided a review for the value of NfL as a biomarker in polyQ disease and its future research directions.
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Enfermedades Neurodegenerativas , Proteínas de Neurofilamentos , Péptidos , Humanos , Péptidos/genética , Enfermedades Neurodegenerativas/genética , Enfermedades Neurodegenerativas/metabolismo , Proteínas de Neurofilamentos/genética , Biomarcadores , AnimalesRESUMEN
Recently, the mounting integration of probiotics into human health strategies has gathered considerable attention. Although the benefits of probiotics have been widely recognized in patients with gastrointestinal disorders, immune system modulation, and chronic-degenerative diseases, there is a growing need to evaluate their potential risks. In this context, new concerns have arisen regarding the safety of probiotics as some strains may have adverse effects in humans. Among these strains, Escherichia coli Nissle 1917 (EcN) exhibited traits of concern due to a pathogenic locus in its genome that produces potentially genotoxic metabolites. As the use of probiotics for therapeutic purposes is increasing, the effects of potentially harmful probiotics must be carefully evaluated. To this end, in this narrative review article, we reported the findings of the most relevant in vitro and in vivo studies investigating the expanding applications of probiotics and their impact on human well-being addressing concerns arising from the presence of antibiotic resistance and pathogenic elements, with a focus on the polyketide synthase (pks) pathogenic island of EcN. In this context, the literature data here discussed encourages a thorough profiling of probiotics to identify potential harmful elements as done for EcN where potential genotoxic effects of colibactin, a secondary metabolite, were observed. Specifically, while some studies suggest EcN is safe for gastrointestinal health, conflicting findings highlight the need for further research to clarify its safety and optimize its use in therapy. Overall, the data here presented suggest that a comprehensive assessment of the evolving landscape of probiotics is essential to make evidence-based decisions and ensure their correct use in humans.
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Escherichia coli , Péptidos , Policétidos , Probióticos , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Policétidos/metabolismo , Péptidos/metabolismo , Péptidos/genética , Animales , Mutágenos/metabolismo , Mutágenos/toxicidad , Daño del ADN , Sintasas Poliquetidas/genética , Sintasas Poliquetidas/metabolismoRESUMEN
Borosins are a class of ribosomally synthesized and post-translationally modified peptides (RiPPs) with α-N-methylated backbones. Although the first mature compound of borosin was reported in 1997, the biosynthetic pathway was elucidated 20 years later. Until this work, borosins have been able to be categorized into 11 types based on the features of their protein structure and core peptides. Type III borosins were reported only in fungi initially. In order to explore the sources and potential of type III borosins, a precise genome mining work of type III borosins was conducted in bacteria and KchMA's self-methylation activity was validated by biochemical experiment. Furthermore, a commercial protease and AI-assisted rational design was employed to engineer KchMA for the capacity to produce various N-methylated peptides. Our work demonstrates that type III borosins are abundant not only in eukaryotes but also in bacteria and have immense potential as a tool for synthetic biology.
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Genoma Bacteriano , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/química , Procesamiento Proteico-Postraduccional , Bacterias/genética , Bacterias/metabolismo , Péptidos/metabolismo , Péptidos/química , Péptidos/genética , Metilación , Secuencia de AminoácidosRESUMEN
This study presents the Maize PeptideAtlas resource (www.peptideatlas.org/builds/maize) to help solve questions about the maize proteome. Publicly available raw tandem mass spectrometry (MS/MS) data for maize collected from ProteomeXchange were reanalyzed through a uniform processing and metadata annotation pipeline. These data are from a wide range of genetic backgrounds and many sample types and experimental conditions. The protein search space included different maize genome annotations for the B73 inbred line from MaizeGDB, UniProtKB, NCBI RefSeq, and for the W22 inbred line. 445 million MS/MS spectra were searched, of which 120 million were matched to 0.37 million distinct peptides. Peptides were matched to 66.2% of proteins in the most recent B73 nuclear genome annotation. Furthermore, most conserved plastid- and mitochondrial-encoded proteins (NCBI RefSeq annotations) were identified. Peptides and proteins identified in the other B73 genome annotations will improve maize genome annotation. We also illustrate the high-confidence detection of unique W22 proteins. N-terminal acetylation, phosphorylation, ubiquitination, and three lysine acylations (K-acetyl, K-malonyl, and K-hydroxyisobutyryl) were identified and can be inspected through a PTM viewer in PeptideAtlas. All matched MS/MS-derived peptide data are linked to spectral, technical, and biological metadata. This new PeptideAtlas is integrated in MaizeGDB with a peptide track in JBrowse.
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Anotación de Secuencia Molecular , Proteínas de Plantas , Espectrometría de Masas en Tándem , Zea mays , Zea mays/genética , Zea mays/química , Proteínas de Plantas/genética , Bases de Datos de Proteínas , Péptidos/genética , Péptidos/química , Genoma de Planta , Proteoma/genética , Proteoma/análisis , Proteómica/métodosRESUMEN
Huntington's disease (HD) is caused by a polyglutamine expansion of the huntingtin protein, resulting in the formation of polyglutamine aggregates. The mechanisms of toxicity that result in the complex HD pathology remain only partially understood. Here, we show that nuclear polyglutamine aggregates induce nuclear envelope (NE) blebbing and ruptures that are often repaired incompletely. These ruptures coincide with disruptions of the nuclear lamina and lead to lamina scar formation. Expansion microscopy enabled resolving the ultrastructure of nuclear aggregates and revealed polyglutamine fibrils sticking into the cytosol at rupture sites, suggesting a mechanism for incomplete repair. Furthermore, we found that NE repair factors often accumulated near nuclear aggregates, consistent with stalled repair. These findings implicate nuclear polyQ aggregate-induced loss of NE integrity as a potential contributing factor to Huntington's disease and other polyglutamine diseases.
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Enfermedad de Huntington , Membrana Nuclear , Péptidos , Membrana Nuclear/metabolismo , Membrana Nuclear/ultraestructura , Humanos , Péptidos/metabolismo , Péptidos/genética , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/patología , Enfermedad de Huntington/genética , Animales , Proteína Huntingtina/metabolismo , Proteína Huntingtina/genética , Agregado de Proteínas , Lámina Nuclear/metabolismo , Lámina Nuclear/ultraestructura , Núcleo Celular/metabolismoRESUMEN
Colibactin, is a cyclomodulin expressed from polyketide synthase (pk) genomic islands. These bacterial toxins interfere with the eukaryotic cell cycle and induce DNA damage. The aim of the present study was to investigate the prevalence of colibactin production among E. coli strains recovered from different infections, determine the similarity of clb nucleotide sequences, and identify genotype of isolates using multilocus sequence typing(MLST). This was a prospective, cross-sectional study conducted from January 2022 to February 2023. A total of 117 clinical isolates were obtained from various sample types collected from outpatients and inpatients recruited to the Department of Bacteriology Labs in different hospitals in Baghdad, Iraq. clbA/clbR, clbB and clbP/clbQ were detected via conventional PCR, and partial sequencing of amplicons was performed via Sanger sequencing. For select isolates, MLST genotyping was performed. The most common phylogenetic group was B2 (61/106; 57.54%). Among the E. coli strains, 27/106 (25.47%) were clb + ve, and the most common type was clbB (13/27; 48.14%). Analysis of the partial sequencing of clb among the strains revealed high molecular similarity. Genotyping of 37 selected E. coli strains via MLST revealed 28 different genotypes. There was a high prevalence of colibactin production in phylogroup B2, and it seems that the clb + ve strains had conserved molecular structures. There was high genetic diversity among the strains tested.
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Infecciones por Escherichia coli , Proteínas de Escherichia coli , Escherichia coli , Tipificación de Secuencias Multilocus , Péptidos , Policétidos , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Policétidos/metabolismo , Péptidos/metabolismo , Péptidos/genética , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/epidemiología , Filogenia , Estudios Transversales , Genotipo , Estudios Prospectivos , Masculino , Femenino , AdultoRESUMEN
BACKGROUND: As the leading cause of end-stage liver disease, nonalcoholic fatty liver disease (NAFLD) is mainly induced by lipid dyshomeostasis. The translation of endogenous circular RNAs (circRNAs) is closely related to the progression of various diseases, but the involvement of circRNAs in NAFLD has not been determined. METHODS: Combined high-throughput circRNA profiles were used to identify circRNAs with translational potential. The underlying molecular mechanisms were investigated by RNA sequencing, pull-down/MS and site-specific mutagenesis. RESULTS: In this study, we focused on circ-SLC9A6, an abnormally highly expressed circRNA in human and mouse liver tissue during NAFLD development that exacerbates metabolic dyshomeostasis in hepatocytes by encoding a novel peptide called SLC9A6-126aa in vivo and in vitro. YTHDF2-mediated degradation of m6A-modified circ-SLC9A6 was found to be essential for the regulation of SLC9A6-126aa expression. We further found that the phosphorylation of SLC9A6-126aa by AKT was crucial for its cytoplasmic localization and the maintenance of physiological homeostasis, whereas high-fat stress induced substantial translocation of unphosphorylated SLC9A6-126aa to the nucleus, resulting in a vicious cycle of lipid metabolic dysfunction. Nuclear SLC9A6-126aa promotes transcriptional activation of the target gene CD36 and enhances its occupancy of the CD36 promoter locus by regulating MOF-mediated histone H4K16 acetylation. Hepatic CD36 depletion significantly ameliorated hyperactivated MAPK signalling and lipid disturbance in SLC9A6-126aa transgenic mice. Clinically, increasing levels of SLC9A6-126aa were observed during NAFLD progression and were found to be positively correlated with the CD36 and MAPK cascades. CONCLUSION: This study revealed the role of circ-SLC9A6-derived SLC9A6-126aa in the epigenetic modification-mediated regulation of lipid metabolism. Our findings may provide promising therapeutic targets for NAFLD and new insights into the pathological mechanisms of metabolic diseases. HIGHLIGHTS: Under normal circumstances, driven by m6A modification, YTHDF2 directly recognizes and degrades circ-SLC9A6, thereby inhibiting the translation of SLC9A6-126aa. Additionally, AKT1 phosphorylates and inhibits the nuclear translocation of SLC9A6-126aa. In NAFLD, lipid overload leads to YTHDF2 and AKT1 deficiency, ultimately increasing the expression and nuclear import of SLC9A6-126aa. Nuclear SLC9A6-126aa binds directly to the CD36 promoter and initiates CD36 transcription, which induces lipid dyshomeostasis.
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Antígenos CD36 , Enfermedad del Hígado Graso no Alcohólico , Animales , Humanos , Masculino , Ratones , Antígenos CD36/genética , Antígenos CD36/metabolismo , Homeostasis/genética , Metabolismo de los Lípidos/genética , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/genética , Péptidos/metabolismo , Péptidos/genética , ARN Circular/genética , ARN Circular/metabolismoRESUMEN
Prolines cause ribosomes to stall during translation due to their rigid structure. This phenomenon occurs in all domains of life and is exacerbated at polyproline motifs. Such stalling can be eased by the elongation factor P (EF-P) in bacteria. We discovered a potential connection between the loss of ancestral EF-P, the appearance of horizontally transferred EF-P variants, and genomic signs of EF-P dysfunction. Horizontal transfer of the efp gene has occurred several times among bacteria and is associated with the loss of highly conserved polyproline motifs. In this study, we pinpoint cases of horizontal EF-P transfer among a diverse set of bacteria and examine genomic features associated with these events in the phyla Thermotogota and Planctomycetes. In these phyla, horizontal EF-P transfer is also associated with the loss of entire polyproline motif-containing proteins, whose expression is likely dependent on EF-P. In particular, three proteases (Lon, ClpC, and FtsH) and three tRNA synthetases (ValS, IleS1, and IleS2) appear highly sensitive to EF-P transfer. The conserved polyproline motifs within these proteins all reside within close proximity to ATP-binding-regions, some of which are crucial for their function. Our work shows that an ancient EF-P dysfunction has left genomic traces that persist to this day, although it remains unclear whether this dysfunction was strictly due to loss of ancestral EF-P or was related to the appearance of an exogenous variant. The latter possibility would imply that the process of "domesticating" a horizontally transferred efp gene can perturb the overall function of EF-P.
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Evolución Molecular , Transferencia de Gen Horizontal , Factores de Elongación de Péptidos , Péptidos , Proteoma , Factores de Elongación de Péptidos/genética , Factores de Elongación de Péptidos/metabolismo , Péptidos/metabolismo , Péptidos/genética , Bacterias/genética , Bacterias/metabolismo , Filogenia , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismoRESUMEN
Follicular atresia in chickens seriously reduced the egg production and economic benefits of chickens. LncRNA plays a key role in the process of follicular atresia. In this study, RNA-seq and Ribo-seq were performed on normal and atretic follicles of Dahen broilers to screen out lncRNAs that may regulate follicle atresia, and to study the molecular mechanisms of their regulation. GRN granulin precursor (lncGRN, ID: 101748909) was highly expressed in atretic follicles with translational ability. A molecular regulatory network of lncGRN/miR-103-3p/FBXW7 was constructed through bioinformatics analysis and dual luciferase reporting. LncGRN promoted the expression of FBXW7 by adsorption of miR-103-3p, thereby inhibiting the proliferation of chicken granulosa cells (GCs), promoting apoptosis of chicken GCs and inhibiting steroid hormone synthesis thus induced follicular atresia. Meanwhile, we also found a micropeptide named GRN-122aa derived by lncGRN which can promote follicular atresia. In conclusion, our study found that lncGRN promoted follicular atresia through the lncGRN/miR-103-3p/FBXW7 axis and the translation micropeptide GRN-122aa. This study provided new insight into the post-transcriptional regulation mechanism of lncGRN suggesting that lncGRN may act as a potential to regulate chicken follicle development, and provided a theoretical argument for further improving the egg production of chickens through molecular breeding.
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Pollos , Atresia Folicular , MicroARNs , ARN Largo no Codificante , Animales , Pollos/genética , MicroARNs/genética , ARN Largo no Codificante/genética , Atresia Folicular/genética , Atresia Folicular/metabolismo , Femenino , Células de la Granulosa/metabolismo , RNA-Seq , Regulación de la Expresión Génica , Apoptosis/genética , Proliferación Celular/genética , Péptidos/genética , Perfilado de RibosomasRESUMEN
Intermediate-length repeat expansions in ATAXIN-2 (ATXN2) are the strongest genetic risk factor for amyotrophic lateral sclerosis (ALS). At the molecular level, ATXN2 intermediate expansions enhance TDP-43 toxicity and pathology. However, whether this triggers ALS pathogenesis at the cellular and functional level remains unknown. Here, we combine patient-derived and mouse models to dissect the effects of ATXN2 intermediate expansions in an ALS background. iPSC-derived motor neurons from ATXN2-ALS patients show altered stress granules, neurite damage and abnormal electrophysiological properties compared to healthy control and other familial ALS mutations. In TDP-43Tg-ALS mice, ATXN2-Q33 causes reduced motor function, NMJ alterations, neuron degeneration and altered in vitro stress granule dynamics. Furthermore, gene expression changes related to mitochondrial function and inflammatory response are detected and confirmed at the cellular level in mice and human neuron and organoid models. Together, these results define pathogenic defects underlying ATXN2-ALS and provide a framework for future research into ATXN2-dependent pathogenesis and therapy.
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Esclerosis Amiotrófica Lateral , Ataxina-2 , Modelos Animales de Enfermedad , Células Madre Pluripotentes Inducidas , Ratones Transgénicos , Neuronas Motoras , Péptidos , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Ataxina-2/genética , Ataxina-2/metabolismo , Humanos , Animales , Péptidos/metabolismo , Péptidos/genética , Ratones , Células Madre Pluripotentes Inducidas/metabolismo , Neuronas Motoras/metabolismo , Neuronas Motoras/patología , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Fenotipo , Masculino , Femenino , Mitocondrias/metabolismo , Neuritas/metabolismoRESUMEN
The aberrant fibrillization of huntingtin exon 1 (Httex1) characterized by an expanded polyglutamine (polyQ) tract is a defining feature of Huntington's disease, a neurodegenerative disorder. Recent investigations underscore the involvement of a small EDRK-rich factor 1a (SERF1a) in promoting Httex1 fibrillization through interactions with its N terminus. By establishing an integrated approach with size-exclusion-column-based small- and wide-angle X-ray scattering (SEC-SWAXS), NMR, and molecular simulations using Rosetta, the analysis here reveals a tight binding of two NT17 fragments of Httex1 (comprising the initial 17 amino acids at the N terminus) to the N-terminal region of SERF1a. In contrast, examination of the complex structure of SERF1a with a coiled NT17-polyQ peptide (33 amino acids in total) indicates sparse contacts of the NT17 and polyQ segments with the N-terminal side of SERF1a. Furthermore, the integrated SEC-SWAXS and molecular-simulation analysis suggests that the coiled NT17 segment can transform into a helical conformation when associated with a polyQ segment exhibiting high helical content. Intriguingly, NT17-polyQ peptides with enhanced secondary structures display diminished interactions with SERF1a. This insight into the conformation-dependent binding of NT17 provides clues to a catalytic association mechanism underlying SERF1a's facilitation of Httext1 fibrillization.
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Proteína Huntingtina , Péptidos , Proteína Huntingtina/genética , Proteína Huntingtina/química , Proteína Huntingtina/metabolismo , Péptidos/química , Péptidos/metabolismo , Péptidos/genética , Humanos , Exones/genética , Unión Proteica , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Simulación de Dinámica Molecular , Espectroscopía de Resonancia Magnética , Difracción de Rayos XRESUMEN
Advances in mass spectrometry (MS) have enabled high-throughput analysis of proteomes in biological systems. The state-of-the-art MS data analysis relies on database search algorithms to quantify proteins by identifying peptide-spectrum matches (PSMs), which convert mass spectra to peptide sequences. Different database search algorithms use distinct search strategies and thus may identify unique PSMs. However, no existing approaches can aggregate all user-specified database search algorithms with a guaranteed increase in the number of identified peptides and a control on the false discovery rate (FDR). To fill in this gap, we proposed a statistical framework, Aggregation of Peptide Identification Results (APIR), that is universally compatible with all database search algorithms. Notably, under an FDR threshold, APIR is guaranteed to identify at least as many, if not more, peptides as individual database search algorithms do. Evaluation of APIR on a complex proteomics standard dataset showed that APIR outpowers individual database search algorithms and empirically controls the FDR. Real data studies showed that APIR can identify disease-related proteins and post-translational modifications missed by some individual database search algorithms. The APIR framework is easily extendable to aggregating discoveries made by multiple algorithms in other high-throughput biomedical data analysis, e.g., differential gene expression analysis on RNA sequencing data. The APIR R package is available at https://github.com/yiling0210/APIR.
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Algoritmos , Bases de Datos de Proteínas , Péptidos , Proteómica , Proteómica/métodos , Péptidos/metabolismo , Péptidos/genética , Humanos , Programas InformáticosRESUMEN
Ganoderma has received much attention for its medicinal value, but the manipulation of multiple genes remains a challenge, hindering the genetic engineering of this species for the development of cell factories. Here, we first showed that the presence of an intron is necessary for the efficient expression of the endogenous cDNA of carboxin-resistant gene (cbx) in G. lucidum. Then, the self-cleaving function of 2â¯A peptide was investigated in G. lucidum by linking cbx cDNA to the codon-optimized hygromycin B-resistant gene (ophph) using the 2A-peptide sequence. The results showed that cbx cDNA and ophph can be successfully expressed in G. lucidum in a bicistronic manner from a single transcript. Moreover, the expression of both genes was not affected by the order within the 2â¯A cassette. In addition, simultaneous expression of cbx cDNA, ophph, and codon-optimized yellow fluorescent protein gene (opyfp) was conducted for the first time in G. lucidum using the 2â¯A peptide-based approach. The developed method was successfully applied to express both cDNA of the 3-hydroxy-3-methylglutaryl coenzyme A reductase (hmgr) and squalene epoxidase gene (se) for enhanced production of ganoderic acids (GAs) in G. lucidum. The engineered strain produced the maximum content of GA-Mk, GA-T, GA-S, and GA-Me were 26.56±3.53,39.58±3.75, 16.54±2.16, and 19.1±1.87 µg/100â¯mg dry weight, respectively. These values were 3.85-, 4.74-, 3.65-, and 3.23-fold higher than those produced by the control strain. The developed method will be useful for the manipulation of complex metabolic or regulatory pathways involving multiple genes in Ganoderma.
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Reishi , Triterpenos , Reishi/genética , Reishi/metabolismo , Triterpenos/metabolismo , Péptidos/genética , Péptidos/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Hidroximetilglutaril-CoA Reductasas/genética , Hidroximetilglutaril-CoA Reductasas/metabolismoRESUMEN
HLA class II antigen presentation is modulated by the activity of the peptide editor HLA-DM and its antagonist HLA-DO, with their interplay controlling the peptide repertoires presented by normal and malignant cells. The role of these molecules in allogeneic hematopoietic cell transplantation (alloHCT) is poorly investigated. Balanced expression of HLA-DM and HLA-DO can influence the presentation of leukemia-associated antigens and peptides targeted by alloreactive T cells, therefore affecting both anti-leukemia immunity and the potential onset of Graft versus Host Disease. We leveraged on a large collection of bulk and single cell RNA sequencing data, available at different repositories, to comprehensively review the level and distribution of HLA-DM and HLA-DO in different cell types and tissues of the human body. The resulting expression atlas will help future investigations aiming to dissect the dual role of HLA class II peptide editing in alloHCT, and their potential impact on its clinical outcome.
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Antígenos HLA-D , Leucemia , Humanos , Leucemia/terapia , Leucemia/inmunología , Leucemia/genética , Antígenos HLA-D/genética , Antígenos HLA-D/inmunología , Trasplante de Células Madre Hematopoyéticas , Presentación de Antígeno , Péptidos/inmunología , Péptidos/genética , AloinjertosRESUMEN
It was previously thought that ncRNA could not encode polypeptides, but recent reports have challenged this notion. As research into ncRNA progresses, it is increasingly clear that it serves roles beyond traditional mechanisms, playing significant regulatory roles in various diseases, notably cancer, which is responsible for 70% of human deaths. Numerous studies have highlighted the diverse regulatory mechanisms of ncRNA that are pivotal in cancer initiation and progression. The role of ncRNA-encoded polypeptides in cancer regulation has gained prominence. This article explores the newly identified regulatory functions of these polypeptides in three types of ncRNA-lncRNA, pri-miRNA, and circRNA. These polypeptides can interact with proteins, influence signaling pathways, enhance miRNA stability, and regulate cancer progression, malignancy, resistance, and other clinical challenges. Furthermore, we discuss the evolutionary significance of these polypeptides in the transition from RNA to protein, examining their emergence and conservation throughout evolution.
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Neoplasias , Péptidos , ARN no Traducido , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Péptidos/genética , Péptidos/metabolismo , ARN no Traducido/genética , ARN no Traducido/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Regulación Neoplásica de la Expresión Génica , ARN Circular/genética , ARN Circular/metabolismo , Animales , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , Transducción de SeñalRESUMEN
Ribosome profiling experiments support the translation of a range of novel human open reading frames. By contrast, most peptides from large-scale proteomics experiments derive from just one source, 5' untranslated regions. Across the human genome we find evidence for 192 translated upstream regions, most of which would produce protein isoforms with extended N-terminal ends. Almost all of these N-terminal extensions are from highly abundant genes, which suggests that the novel regions we detect are just the tip of the iceberg. These upstream regions have characteristics that are not typical of coding exons. Their GC-content is remarkably high, even higher than 5' regions in other genes, and a large majority have non-canonical start codons. Although some novel upstream regions have cross-species conservation - five have orthologues in invertebrates for example - the reading frames of two thirds are not conserved beyond simians. These non-conserved regions also have no evidence of purifying selection, which suggests that much of this translation is not functional. In addition, non-conserved upstream regions have significantly more peptides in cancer cell lines than would be expected, a strong indication that an aberrant or noisy translation initiation process may play an important role in translation from upstream regions.
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Regiones no Traducidas 5' , Biosíntesis de Proteínas , Humanos , Codón Iniciador/genética , Composición de Base , Genoma Humano , Animales , Sistemas de Lectura Abierta/genética , Secuencia Conservada , Péptidos/genética , Péptidos/metabolismoRESUMEN
Butelase-1, the fastest known Asn/Asp-specific peptide ligase capable of catalyzing peptide ligation and cyclization, holds promising application prospects in the fields of food and biology. However, limited research exists on its recombinant expression and potential applications in peptide drugs. In this study, the activity of recombinantly-produced butelase-1 was enhanced by co-expressing it with a molecular chaperone in the SHuffle T7 strain. By introducing single or multiple synonymous rare codons at the beginning of the coding regions of beta-strand or alpha-helix, in combination with ribosomal binding site engineering, the activity of butelase-1 could be further improved. Consequently, the butelase-1 with a specific activity of 386.93 U/mg and a catalytic efficiency of 11,048 M-1 s-1 was successfully prepared in E. coli, resulting in a total activity of 8183.54 U/L and a yield of about 100 mg/L. This optimized butelase-1 was then used to efficiently cyclize the redesigned anti-cancer peptide lunasin, leading to enhanced bioavailability and anti-cancer effects. Overall, this study not only provided valuable biotechnology strategies for improving the recombinant expression of butelase-1 but also demonstrated a successful application for enhancing the biological efficacy of anti-cancer peptides.
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Antineoplásicos , Escherichia coli , Humanos , Antineoplásicos/farmacología , Antineoplásicos/química , Escherichia coli/genética , Escherichia coli/metabolismo , Expresión Génica , Péptidos/química , Péptidos/metabolismo , Péptidos/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/químicaRESUMEN
The prokaryotic translation elongation factor P (EF-P) and the eukaryotic/archaeal counterparts eIF5A/aIF5A are proteins that serve a crucial role in mitigating ribosomal stalling during the translation of specific sequences, notably those containing consecutive proline residues (1,2). Although mitochondrial DNA-encoded proteins synthesized by mitochondrial ribosomes also contain polyproline stretches, an EF-P/eIF5A mitochondrial counterpart remains unidentified. Here, we show that the missing factor is TACO1, a protein causative of a juvenile form of neurodegenerative Leigh's syndrome associated with cytochrome c oxidase deficiency, until now believed to be a translational activator of COX1 mRNA. By using a combination of metabolic labeling, puromycin release and mitoribosome profiling experiments, we show that TACO1 is required for the rapid synthesis of the polyproline-rich COX1 and COX3 cytochrome c oxidase subunits, while its requirement is negligible for other mitochondrial DNA-encoded proteins. In agreement with a role in translation efficiency regulation, we show that TACO1 cooperates with the N-terminal extension of the large ribosomal subunit bL27m to provide stability to the peptidyl-transferase center during elongation. This study illuminates the translation elongation dynamics within human mitochondria, a TACO1-mediated biological mechanism in place to mitigate mitoribosome stalling at polyproline stretches during protein synthesis, and the pathological implications of its malfunction.
Asunto(s)
Complejo IV de Transporte de Electrones , Proteínas Mitocondriales , Ribosomas Mitocondriales , Péptidos , Biosíntesis de Proteínas , Humanos , Complejo IV de Transporte de Electrones/metabolismo , Complejo IV de Transporte de Electrones/genética , Proteínas Mitocondriales/metabolismo , Proteínas Mitocondriales/genética , Péptidos/metabolismo , Péptidos/genética , Ribosomas Mitocondriales/metabolismo , Mitocondrias/metabolismo , Mitocondrias/genética , Factores de Iniciación de Péptidos/metabolismo , Factores de Iniciación de Péptidos/genética , Factores de Elongación de Péptidos/metabolismo , Factores de Elongación de Péptidos/genética , Células HEK293 , Ciclooxigenasa 1RESUMEN
Virus-like particles (VLPs) of the adeno-associated virus (AAV) can be produced using the baculovirus expression vector system. Insertion of small peptides on the surface of the AAV or AAV VLPs has been used to redirect the AAV to different target tissues and for vaccine development. Usually, the VLPs self-assemble intracellularly, and an extraction step must be performed before purification. Here, we describe the method we have used to extract AAV VLPs from insect cells successfully with peptide insertions on their surface.