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1.
PLoS One ; 15(12): e0239566, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33301503

RESUMEN

The ongoing pandemic of SARS-CoV-2 has brought tremendous crisis on global health care systems and industrial operations that dramatically affect the economic and social life of numerous individuals worldwide. Understanding anti-SARS-CoV-2 immune responses in population with different genetic backgrounds and tracking the viral evolution are crucial for successful vaccine design. In this study, we reported the generation of CD8 T cell epitopes by a total of 80 alleles of three major class I HLAs using NetMHC 4.0 algorithm for the SARS-CoV-2 spike protein, which can be targeted by both B cells and T cells. We found diverse capacities of S protein specific epitope presentation by different HLA alleles with very limited number of predicted epitopes for HLA-B*2705, HLA-B*4402 and HLA-B*4403 and as high as 132 epitopes for HLA-A*6601. Our analysis of 1000 S protein sequences from field isolates collected globally over the past few months identified three recurrent point mutations including L5F, D614G and G1124V. Differential effects of these mutations on CD8 T cell epitope generation by corresponding HLA alleles were observed. Finally, our multiple alignment analysis indicated the absence of seasonal CoV induced cross-reactive CD8 T cells to drive these mutations. Our findings suggested that individuals with certain HLA alleles, such as B*44 are more prone to SARS-CoV-2 infection. Studying anti-S protein specific CD8 T cell immunity in diverse genetic background is critical for better control and prevention of the SARS-CoV-2 pandemic.


Asunto(s)
Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Secuencia de Aminoácidos , Antígenos HLA/inmunología , Humanos , Mutación , Mutación Puntual , /genética , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética
2.
PLoS Genet ; 16(11): e1009090, 2020 11.
Artículo en Inglés | MEDLINE | ID: mdl-33147208

RESUMEN

Interferon ß (IFN-ß) is a cytokine that induces a global antiviral proteome, and regulates the adaptive immune response to infections and tumors. Its effects strongly depend on its level and timing of expression. Therefore, the transcription of its coding gene IFNB1 is strictly controlled. We have previously shown that in mice, the TRIM33 protein restrains Ifnb1 transcription in activated myeloid cells through an upstream inhibitory sequence called ICE. Here, we show that the deregulation of Ifnb1 expression observed in murine Trim33-/- macrophages correlates with abnormal looping of both ICE and the Ifnb1 gene to a 100 kb downstream region overlapping the Ptplad2/Hacd4 gene. This region is a predicted myeloid super-enhancer in which we could characterize 3 myeloid-specific active enhancers, one of which (E5) increases the response of the Ifnb1 promoter to activation. In humans, the orthologous region contains several single nucleotide polymorphisms (SNPs) known to be associated with decreased expression of IFNB1 in activated monocytes, and loops to the IFNB1 gene. The strongest association is found for the rs12553564 SNP, located in the E5 orthologous region. The minor allele of rs12553564 disrupts a conserved C/EBP-ß binding motif, prevents binding of C/EBP-ß, and abolishes the activation-induced enhancer activity of E5. Altogether, these results establish a link between a genetic variant preventing binding of a transcription factor and a higher order phenotype, and suggest that the frequent minor allele (around 30% worldwide) might be associated with phenotypes regulated by IFN-ß expression in myeloid cells.


Asunto(s)
Proteína beta Potenciadora de Unión a CCAAT/metabolismo , Elementos de Facilitación Genéticos , Regulación de la Expresión Génica/inmunología , Interferón beta/genética , Células Mieloides/metabolismo , Alelos , Animales , Capa Leucocitaria de la Sangre/citología , Células Cultivadas , Humanos , Interferón beta/inmunología , Lipopolisacáridos/inmunología , Ratones , Ratones Noqueados , Células Mieloides/inmunología , Mutación Puntual , Polimorfismo de Nucleótido Simple , Cultivo Primario de Células , Regiones Promotoras Genéticas , Sitios de Carácter Cuantitativo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
3.
PLoS One ; 15(11): e0241902, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33170892

RESUMEN

BACKGROUND: Glaucoma represents the second main cause of irreversible loss of eyesight worldwide. Progression of the disease is due to changes around the optic nerve, eye structure and optic nerve environment. Focusing on primary congenital glaucoma, which is not completely understood, we report an evaluation of an untested mutation (c.182G>A, p.Gly61Glu) within the CYP1B1 gene in the context of microglia, astrocytes and mesenchymal stem cells. We investigated the behaviours of these cells, which are needed to maintain eye homeostasis, in response to the CYP1B1 mutation. METHODS AND RESULTS: CRISPR technology was used to edit normal CYP1B1 genes within normal astrocytes, microglia and stem cells in vitro. Increased metabolic activities were found in microglia and astrocytes 24 hours after CYP1B1 manipulation. However, these activities dropped by 40% after 72 hrs. In addition, the nicotinamide adenine dinucleotide phosphate (NADP)/NADPH reducing equivalent process decreased by 50% on average after 72 hrs of manipulation. The cytokines measured in mutated microglia showed progressive activation leading to apoptosis, which was confirmed with annexin-V. The cytokines evaluated in mutant astrocytes were abnormal in comparison to those in the control. CONCLUSIONS: The results suggest a progressive inflammation that was induced by mutations (p.Gly61Glu) on CYP1B1. Furthermore, the mutations enhanced the microglia's loss of activity. We are the first to show the direct impact of the mutation on microglia. This progressive inflammation might be responsible for primary congenital glaucoma complications, which could be avoided via an anti-inflammatory regimen. This finding also reveals that progressive inflammation affects recovery failure after surgeries to relieve glaucoma. Moreover, microglia are important for the survival of ganglion cells, along with the clearing of pathogens and inflammation. The reduction of their activities may jeopardise homeostasis within the optic nerve environment and complicate the protection of optic nerve components (such as retinal ganglion and glial cells).


Asunto(s)
Astrocitos/citología , Citocromo P-450 CYP1B1/genética , Glaucoma/congénito , Células Madre Mesenquimatosas/citología , Microglía/citología , Mutación Puntual , Sustitución de Aminoácidos , Animales , Apoptosis , Astrocitos/metabolismo , Sistemas CRISPR-Cas , Proliferación Celular , Células Cultivadas , Citocinas/metabolismo , Glaucoma/genética , Humanos , Masculino , Células Madre Mesenquimatosas/metabolismo , Microglía/metabolismo , Modelos Animales , NADP/metabolismo , Ratas
4.
PLoS One ; 15(11): e0240932, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33141820

RESUMEN

Poly(ADP-ribose) Polymerase 2 (PARP2) is one of three DNA-dependent PARPs involved in the detection of DNA damage. Upon binding to DNA double-strand breaks, PARP2 uses nicotinamide adenine dinucleotide to synthesize poly(ADP-ribose) (PAR) onto itself and other proteins, including histones. PAR chains in turn promote the DNA damage response by recruiting downstream repair factors. These early steps of DNA damage signaling are relevant for understanding how genome integrity is maintained and how their failure leads to genome instability or cancer. There is no structural information on DNA double-strand break detection in the context of chromatin. Here we present a cryo-EM structure of two nucleosomes bridged by human PARP2 and confirm that PARP2 bridges DNA ends in the context of nucleosomes bearing short linker DNA. We demonstrate that the conformation of PARP2 bound to damaged chromatin provides a binding platform for the regulatory protein Histone PARylation Factor 1 (HPF1), and that the resulting HPF1•PARP2•nucleosome complex is enzymatically active. Our results contribute to a structural view of the early steps of the DNA damage response in chromatin.


Asunto(s)
Proteínas Portadoras/metabolismo , Roturas del ADN de Doble Cadena , Proteínas Nucleares/metabolismo , Poli(ADP-Ribosa) Polimerasas/metabolismo , Sustitución de Aminoácidos , Proteínas Portadoras/química , Proteínas Portadoras/genética , Microscopía por Crioelectrón , Humanos , Técnicas In Vitro , Modelos Moleculares , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas Nucleares/química , Proteínas Nucleares/genética , Nucleosomas/metabolismo , Nucleosomas/ultraestructura , Mutación Puntual , Poli(ADP-Ribosa) Polimerasas/química , Poli(ADP-Ribosa) Polimerasas/genética , Unión Proteica , Conformación Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
5.
PLoS One ; 15(11): e0241168, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33170884

RESUMEN

The SARS-CoV-2 virion responsible for the current world-wide pandemic COVID-19 has a characteristic Spike protein (S) on its surface that embellishes both a prefusion state and fusion state. The prefusion Spike protein (S) is a large trimeric protein where each protomer may be in a so-called Up state or Down state, depending on the configuration of its receptor binding domain (RBD) within its distal, prefusion S1 domain. The Up state is believed to allow binding of the virion to ACE-2 receptors on human epithelial cells, whereas the Down state is believed to be relatively inactive or reduced in its binding behavior. We have performed detailed all-atom, dominant energy landscape mappings for noncovalent interactions (charge, partial charge, and van der Waals) of the SARS-CoV-2 Spike protein in its static prefusion state based on two recent and independent experimental structure publications. We included both interchain interactions and intrachain (domain) interactions in our mappings in order to determine any telling differences (different so-called "glue" points) between residues in the Up and Down state protomers. The S2 proximal, fusion domain demonstrated no appreciable energetic differences between Up and Down protomers, including interchain as well as each protomer's intrachain, S1-S2 interactions. However, the S1 domain interactions across neighboring protomers, which include the RBD-NTD cross chain interactions, showed significant energetic differences between Up-Down and Down-Down neighboring protomers. This included, for example, a key RBD residue ARG357 in the Up-Down interaction and a three residue sequence ALA520-PRO521-ALA522, associated with a turn structure in the RBD of the Up state protomer, acting as a stabilizing interaction with the NTD of its neighbor protomer. Additionally, our intra chain dominant energy mappings within each protomer, identified a significant "glue" point or possible "latch" for the Down state protomer between the S1 subdomain, SD1, and the RBD domain of the same protomer that was completely missing in the Up state protomer analysis. Ironically, this dominant energetic interaction in the Down state protomer involved the backbone atoms of the same three residue sequence ALA520-PRO521-ALA522 of the RBD with the amino acid R-group of GLN564 in the SD1 domain. Thus, this same three residue sequence acts as a stabilizer of the RBD in the Up conformation through its interactions with its neighboring NTD chain and a kind of latch in the Down state conformation through its interactions with its own SD1 domain. The dominant interaction energy residues identified here are also conserved across reported variations of SARS-CoV-2, as well as the closely related virions SARS-Cov and the bat corona virus RatG13. We conducted preliminary molecular dynamics simulations across 0.1 µ seconds to see if this latch provided structural stability and indeed found that a single point mutation (Q564G) resulted in the latch releasing transforming the protomer from the Down to the Up state conformation. Full trimeric Spike protein studies of the same mutation across all protomers, however, did not exhibit latch release demonstrating the critical importance of interchain interactions across the S1 domain, including RBD-NTD neighboring chain interactions. Therapies aimed at disrupting these noncovalent interactions could be a viable route for the physico-chemical mitigation of this deadly virion.


Asunto(s)
Betacoronavirus/metabolismo , Glicoproteína de la Espiga del Coronavirus/metabolismo , Betacoronavirus/aislamiento & purificación , Infecciones por Coronavirus/patología , Infecciones por Coronavirus/virología , Humanos , Simulación de Dinámica Molecular , Pandemias , Peptidil-Dipeptidasa A/química , Peptidil-Dipeptidasa A/metabolismo , Neumonía Viral/patología , Neumonía Viral/virología , Mutación Puntual , Unión Proteica , Dominios Proteicos , Estabilidad Proteica , Subunidades de Proteína/química , Subunidades de Proteína/genética , Subunidades de Proteína/metabolismo , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Termodinámica
6.
Nucleic Acids Res ; 48(21): 12336-12347, 2020 12 02.
Artículo en Inglés | MEDLINE | ID: mdl-33231643

RESUMEN

In response to DNA damage, Escherichia coli cells activate the expression of the toxin gene tisB of the toxin-antitoxin system tisB-istR1. Of three isoforms, only the processed, highly structured +42 tisB mRNA is active. Translation requires a standby site, composed of two essential elements: a single-stranded region located 100 nucleotides upstream of the sequestered RBS, and a structure near the 5'-end of the active mRNA. Here, we propose that this 5'-structure is an RNA pseudoknot which is required for 30S and protein S1-alone binding to the mRNA. Point mutations that prevent formation of this pseudoknot inhibit formation of translation initiation complexes, impair S1 and 30S binding to the mRNA, and render the tisB mRNA non-toxic in vivo. A set of mutations created in either the left or right arm of stem 2 of the pseudoknot entailed loss of toxicity upon overexpression of the corresponding mRNA variants. Combining the matching right-left arm mutations entirely restored toxicity levels to that of the wild-type, active mRNA. Finally, since many pseudoknots have high affinity for S1, we predicted similar pseudoknots in non-homologous type I toxin-antitoxin systems that exhibit features similar to that of tisB-IstR1, suggesting a shared requirement for standby acting at great distances.


Asunto(s)
Toxinas Bacterianas/genética , Proteínas de Escherichia coli/genética , Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica , Biosíntesis de Proteínas , ARN Bacteriano/genética , ARN Mensajero/genética , Sistemas Toxina-Antitoxina/genética , Toxinas Bacterianas/metabolismo , Emparejamiento Base , Secuencia de Bases , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Conformación de Ácido Nucleico , Mutación Puntual , Unión Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , ARN Bacteriano/química , ARN Bacteriano/metabolismo , ARN Mensajero/química , ARN Mensajero/metabolismo , Proteínas Ribosómicas/genética , Proteínas Ribosómicas/metabolismo , Subunidades Ribosómicas Pequeñas Bacterianas/genética , Subunidades Ribosómicas Pequeñas Bacterianas/metabolismo
7.
PLoS Genet ; 16(11): e1009172, 2020 11.
Artículo en Inglés | MEDLINE | ID: mdl-33137101

RESUMEN

Mutations in the gene rhodopsin are one of the major causes of autosomal dominant retinitis pigmentosa (adRP). Mutant forms of Rhodopsin frequently accumulate in the endoplasmic reticulum (ER), cause ER stress, and trigger photoreceptor cell degeneration. Here, we performed a genome-wide screen to identify suppressors of retinal degeneration in a Drosophila model of adRP, carrying a point mutation in the major rhodopsin, Rh1 (Rh1G69D). We identified two novel E3 ubiquitin ligases SORDD1 and SORDD2 that effectively suppressed Rh1G69D-induced photoreceptor dysfunction and retinal degeneration. SORDD1/2 promoted the ubiquitination and degradation of Rh1G69D through VCP (valosin containing protein) and independent of processes reliant on the HRD1 (HMG-CoA reductase degradation protein 1)/HRD3 complex. We further demonstrate that SORDD1/2 and HRD1 function in parallel and in a redundant fashion to maintain rhodopsin homeostasis and integrity of photoreceptor cells. These findings identify a new ER-associated protein degradation (ERAD) pathway and suggest that facilitating SORDD1/2 function may be a therapeutic strategy to treat adRP.


Asunto(s)
Proteínas de Drosophila/metabolismo , Degradación Asociada con el Retículo Endoplásmico , Retinitis Pigmentosa/patología , Rodopsina/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Animales , Animales Modificados Genéticamente , Modelos Animales de Enfermedad , Proteínas de Drosophila/genética , Drosophila melanogaster , Electrorretinografía , Retículo Endoplásmico/metabolismo , Técnicas de Inactivación de Genes , Humanos , Larva , Microscopía Electrónica de Rastreo , Mutación Puntual , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteolisis , Retina/diagnóstico por imagen , Retina/patología , Retina/ultraestructura , Retinitis Pigmentosa/diagnóstico , Retinitis Pigmentosa/genética , Rodopsina/genética , Ubiquitina-Proteína Ligasas/genética , Ubiquitinación , Proteína que Contiene Valosina/metabolismo
8.
Nat Commun ; 11(1): 5973, 2020 11 25.
Artículo en Inglés | MEDLINE | ID: mdl-33239610

RESUMEN

The assembly of actin filaments into distinct cytoskeletal structures plays a critical role in cell physiology, but how proteins localize differentially to these structures within a shared cytoplasm remains unclear. Here, we show that the actin-binding domains of accessory proteins can be sensitive to filament conformational changes. Using a combination of live cell imaging and in vitro single molecule binding measurements, we show that tandem calponin homology domains (CH1-CH2) can be mutated to preferentially bind actin networks at the front or rear of motile cells. We demonstrate that the binding kinetics of CH1-CH2 domain mutants varies as actin filament conformation is altered by perturbations that include stabilizing drugs and other binding proteins. These findings suggest that conformational changes of actin filaments in cells could help to direct accessory binding proteins to different actin cytoskeletal structures through a biophysical feedback loop.


Asunto(s)
Citoesqueleto de Actina/metabolismo , Multimerización de Proteína , Utrofina/metabolismo , Citoesqueleto de Actina/química , Factores Despolimerizantes de la Actina/metabolismo , Actinas , Animales , Citoplasma/metabolismo , Células HEK293 , Células HeLa , Humanos , Microscopía Intravital , Cinética , Microscopía Fluorescente , Mutación , Neuropéptidos/metabolismo , Mutación Puntual , Unión Proteica/genética , Dominios Proteicos/genética , Conejos , Imagen Individual de Molécula , Utrofina/química , Utrofina/genética
9.
Yonsei Med J ; 61(12): 1064-1067, 2020 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-33251782

RESUMEN

Hemoglobin M (HbM) is a group of abnormal hemoglobin variants that form methemoglobin, which leads to cyanosis and hemolytic anemia. HbM-Milwaukee-2 is a rare variant caused by the point mutation CAC>TAC on codon 93 of the hemoglobin subunit beta (HBB) gene, resulting in the replacement of histidine by tyrosine. We here report the first Korean family with HbM-Milwaukee-2, whose diagnosis was confirmed by gene sequencing. A high index of suspicion for this rare Hb variant is necessary in a patient presenting with cyanosis since childhood, along with methemoglobinemia and a family history of cyanosis.


Asunto(s)
Cianosis/etiología , Globinas/genética , Hemoglobina M , Metahemoglobinemia/congénito , Adolescente , Niño , Cianosis/genética , Femenino , Globinas/química , Hemoglobina M/genética , Hemoglobinas Anormales/genética , Humanos , Masculino , Metahemoglobina/análisis , Metahemoglobina/genética , Metahemoglobinemia/diagnóstico , Metahemoglobinemia/genética , Mutación Puntual , República de Corea , Análisis de Secuencia de ADN
10.
Int J Mol Sci ; 21(20)2020 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-33076559

RESUMEN

Among the natural bases, guanine is the most oxidizable base. The damage caused by oxidation of guanine, commonly referred to as oxidative guanine damage, results in the formation of several products, including 2,5-diamino-4H-imidazol-4-one (Iz), 2,2,4-triamino-5(2H)-oxazolone (Oz), guanidinoformimine (Gf), guanidinohydantoin/iminoallantoin (Gh/Ia), spiroiminodihydantoin (Sp), 5-carboxamido-5-formamido-2-iminohydantoin (2Ih), urea (Ua), 5-guanidino-4-nitroimidazole (NI), spirodi(iminohydantoin) (5-Si and 8-Si), triazine, the M+7 product, other products by peroxynitrite, alkylated guanines, and 8,5'-cyclo-2'-deoxyguanosine (cG). Herein, we summarize the present knowledge about base pairs containing the products of oxidative guanine damage and guanine. Of these products, Iz is involved in G-C transversions. Oz, Gh/Ia, and Sp form preferably Oz:G, Gh/Ia:G, and Sp:G base pairs in some cases. An involvement of Gf, 2Ih, Ua, 5-Si, 8-Si, triazine, the M+7 product, and 4-hydroxy-2,5-dioxo-imidazolidine-4-carboxylic acid (HICA) in G-C transversions requires further experiments. In addition, we describe base pairs that target the RNA-dependent RNA polymerase (RdRp) of RNA viruses and describe implications for the 2019 novel coronavirus (SARS-CoV-2): When products of oxidative guanine damage are adapted for the ribonucleoside analogs, mimics of oxidative guanine damages, which can form base pairs, may become antiviral agents for SARS-CoV-2.


Asunto(s)
Emparejamiento Base , Guanina/análogos & derivados , Mutación Puntual , Animales , Betacoronavirus/genética , Daño del ADN , Guanina/metabolismo , Humanos , Oxidación-Reducción
11.
Arch Virol ; 165(12): 2937-2944, 2020 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-33025199

RESUMEN

SARS-CoV-2 was first detected in the city of Wuhan, Hubei Province, China. In this study, we identified 11 unique mutations in viral SARS-COV-2 isolates from Turkey. Nine of them cause structural alterations in the S protein, nsp2, nsp3, nsp4 and nsp12 regions. The mutations identified here might have significant functional implications that need to be addressed in future studies in the context of vaccine engineering and therapeutic interventions. Moreover, transmission and phylogenetic analysis revealed multiple independent sources of introductions of SARS-CoV-2 into Turkey and a close relationship to the isolates from Saudi Arabia.


Asunto(s)
Betacoronavirus/genética , Infecciones por Coronavirus/diagnóstico , Infecciones por Coronavirus/epidemiología , Pandemias , Neumonía Viral/diagnóstico , Neumonía Viral/epidemiología , Mutación Puntual , Glicoproteína de la Espiga del Coronavirus/genética , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Betacoronavirus/clasificación , Betacoronavirus/aislamiento & purificación , Betacoronavirus/patogenicidad , Técnicas de Laboratorio Clínico/métodos , Infecciones por Coronavirus/transmisión , Infecciones por Coronavirus/virología , Humanos , Filogenia , Neumonía Viral/transmisión , Neumonía Viral/virología , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Turquia/epidemiología
12.
Sci Rep ; 10(1): 17766, 2020 10 20.
Artículo en Inglés | MEDLINE | ID: mdl-33082451

RESUMEN

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection induces severe pneumonia and is the cause of a worldwide pandemic. Coronaviruses, including SARS-CoV-2, have RNA proofreading enzymes in their genomes, resulting in fewer gene mutations than other RNA viruses. Nevertheless, variants of SARS-CoV-2 exist and may induce different symptoms; however, the factors and the impacts of these mutations are not well understood. We found that there is a bias to the mutations occurring in SARS-CoV-2 variants, with disproportionate mutation to uracil (U). These point mutations to U are mainly derived from cytosine (C), which is consistent with the substrate specificity of host RNA editing enzymes, APOBECs. We also found the point mutations which are consistent with other RNA editing enzymes, ADARs. For the C-to-U mutations, the context of the upstream uracil and downstream guanine from mutated position was found to be most prevalent. Further, the degree of increase of U in SARS-CoV-2 variants correlates with enhanced production of cytokines, such as TNF-α and IL-6, in cell lines when compared with stimulation by the ssRNA sequence of the isolated virus in Wuhan. Therefore, RNA editing is a factor for mutation bias in SARS-CoV-2 variants, which affects host inflammatory cytokines production.


Asunto(s)
Betacoronavirus/inmunología , Infecciones por Coronavirus/patología , Neumonía Viral/patología , Desaminasas APOBEC/metabolismo , Adenosina Desaminasa/metabolismo , Betacoronavirus/clasificación , Betacoronavirus/genética , Betacoronavirus/aislamiento & purificación , Línea Celular Tumoral , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/virología , Humanos , Interleucina-6/metabolismo , Pandemias , Filogenia , Neumonía Viral/inmunología , Neumonía Viral/virología , Mutación Puntual , Edición de ARN , Factor de Necrosis Tumoral alfa/metabolismo , Uracilo/metabolismo
13.
Commun Biol ; 3(1): 641, 2020 10 27.
Artículo en Inglés | MEDLINE | ID: mdl-33110195

RESUMEN

The emergence of SARS-CoV-2 has caused over a million human deaths and massive global disruption. The viral infection may also represent a threat to our closest living relatives, nonhuman primates. The contact surface of the host cell receptor, ACE2, displays amino acid residues that are critical for virus recognition, and variations at these critical residues modulate infection susceptibility. Infection studies have shown that some primate species develop COVID-19-like symptoms; however, the susceptibility of most primates is unknown. Here, we show that all apes and African and Asian monkeys (catarrhines), exhibit the same set of twelve key amino acid residues as human ACE2. Monkeys in the Americas, and some tarsiers, lemurs and lorisoids, differ at critical contact residues, and protein modeling predicts that these differences should greatly reduce SARS-CoV-2 binding affinity. Other lemurs are predicted to be closer to catarrhines in their susceptibility. Our study suggests that apes and African and Asian monkeys, and some lemurs, are likely to be highly susceptible to SARS-CoV-2. Urgent actions have been undertaken to limit the exposure of great apes to humans, and similar efforts may be necessary for many other primate species.


Asunto(s)
Betacoronavirus/patogenicidad , Infecciones por Coronavirus/veterinaria , Especificidad del Huésped/genética , Pandemias/veterinaria , Peptidil-Dipeptidasa A/genética , Neumonía Viral/veterinaria , Enfermedades de los Primates/enzimología , Primates/genética , Receptores Virales/genética , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Animales , Betacoronavirus/fisiología , Evolución Biológica , Quirópteros/genética , Secuencia Conservada , Infecciones por Coronavirus/epidemiología , Infecciones por Coronavirus/transmisión , Predisposición Genética a la Enfermedad , Mamíferos/genética , Modelos Moleculares , Mutación Missense , Peptidil-Dipeptidasa A/química , Filogenia , Neumonía Viral/epidemiología , Neumonía Viral/transmisión , Mutación Puntual , Enfermedades de los Primates/virología , Unión Proteica , Conformación Proteica , Riesgo , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Especificidad de la Especie , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/metabolismo
14.
Euro Surveill ; 25(39)2020 10.
Artículo en Inglés | MEDLINE | ID: mdl-33006300

RESUMEN

We found that a single nucleotide polymorphism (SNP) in the nucleoprotein gene of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from a patient interfered with detection in a widely used commercial assay. Some 0.2% of the isolates in the EpiCoV database contain this SNP. Although SARS-CoV-2 was still detected by the other probe in the assay, this underlines the necessity of targeting two independent essential regions of a pathogen for reliable detection.


Asunto(s)
Betacoronavirus/genética , Técnicas de Laboratorio Clínico , Infecciones por Coronavirus/diagnóstico , Nucleoproteínas/genética , Pandemias , Neumonía Viral/diagnóstico , Mutación Puntual , Polimorfismo de Nucleótido Simple , ARN Viral/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Virales/genética , Secuencia de Bases , Betacoronavirus/aislamiento & purificación , Técnicas de Laboratorio Clínico/métodos , Trazado de Contacto , Infecciones por Coronavirus/virología , Cartilla de ADN , Errores Diagnósticos , Reacciones Falso Negativas , Femenino , Genes Virales , Humanos , Persona de Mediana Edad , Nasofaringe/virología , Nucleoproteínas/análisis , Filogenia , Neumonía Viral/virología , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Rumanía , Enfermedad Relacionada con los Viajes , Proteínas Virales/análisis
15.
Nat Protoc ; 15(12): 3777-3787, 2020 12.
Artículo en Inglés | MEDLINE | ID: mdl-33106680

RESUMEN

The research community is in a race to understand the molecular mechanisms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, to repurpose currently available antiviral drugs and to develop new therapies and vaccines against coronavirus disease 2019 (COVID-19). One major challenge in achieving these goals is the paucity of suitable preclinical animal models. Mice constitute ~70% of all the laboratory animal species used in biomedical research. Unfortunately, SARS-CoV-2 infects mice only if they have been genetically modified to express human ACE2. The inherent resistance of wild-type mice to SARS-CoV-2, combined with a wealth of genetic tools that are available only for modifying mice, offers a unique opportunity to create a versatile set of genetically engineered mouse models useful for COVID-19 research. We propose three broad categories of these models and more than two dozen designs that may be useful for SARS-CoV-2 research and for fighting COVID-19.


Asunto(s)
/genética , Modelos Animales de Enfermedad , /genética , Animales , Secuencia de Bases , Técnicas de Sustitución del Gen , Ingeniería Genética , Sitios Genéticos/genética , Ratones , Ratones Transgénicos , Mutación Puntual
16.
Nat Commun ; 11(1): 5435, 2020 10 28.
Artículo en Inglés | MEDLINE | ID: mdl-33116135

RESUMEN

Memory B cells (MBCs) are long-lived and produce high-affinity, generally, class-switched antibodies. Here, we use a multiparameter approach involving CD27 to segregate naïve B cells (NBC), IgD+ unswitched (unsw)MBCs and IgG+ or IgA+ class-switched (sw)MBCs from humans of different age, sex and race. Conserved antibody variable gene expression indicates that MBCs emerge through unbiased selection from NBCs. Integrative analyses of mRNAs, miRNAs, lncRNAs, chromatin accessibility and cis-regulatory elements uncover a core mRNA-ncRNA transcriptional signature shared by IgG+ and IgA+ swMBCs and distinct from NBCs, while unswMBCs display a transitional transcriptome. Some swMBC transcriptional signature loci are accessible but not expressed in NBCs. Profiling miRNAs reveals downregulated MIR181, and concomitantly upregulated MIR181 target genes such as RASSF6, TOX, TRERF1, TRPV3 and RORα, in swMBCs. Finally, lncRNAs differentially expressed in swMBCs cluster proximal to the IgH chain locus on chromosome 14. Our findings thus provide new insights into MBC transcriptional programs and epigenetic regulation, opening new investigative avenues on these critical cell elements in human health and disease.


Asunto(s)
Linfocitos B/inmunología , Memoria Inmunológica/genética , Adulto , Linfocitos B/clasificación , Linfocitos B/metabolismo , Cromatina/genética , Cromatina/inmunología , Regiones Determinantes de Complementariedad , Epigénesis Genética , Femenino , Perfilación de la Expresión Génica , Humanos , Cambio de Clase de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/genética , Masculino , MicroARNs/genética , MicroARNs/metabolismo , Mutación Puntual , Transducción de Señal/genética , Factores de Transcripción/genética , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/metabolismo , Adulto Joven
17.
Beijing Da Xue Xue Bao Yi Xue Ban ; 52(5): 971-974, 2020 Oct 18.
Artículo en Chino | MEDLINE | ID: mdl-33047739

RESUMEN

The rearrangement of the gene encoding the transcription factor ETS-related gene (ERG) is thought to play a key role in the development of prostate cancer. However, the studies on the ERG mutations have been rarely reported in non-small cell lung carcinoma (NSCLC). Here, we reported genetic features regarding a case of a 68-year-old male patient who presented the primary synchronous multiple tumor lesions in the separated lungs. The patient was hospitalized due to the presence of tumor lesions at the right and left lungs revealed by a chest computerized tomography (CT) scan. After conducting lobectomies at the both lungs, the tumor nodules were all removed, and the histological analysis suggested adenocarcinoma at the both tumor lesions. The patient was diagnosed with synchronous multiple primary lung cancer (SMPLC) based on Martini-Melamed criteria and American College of Chest Physicians practice guidelines. An exome analysis of 315 genes in the two tumor lesions and a non-tumor lesion was conducted by using Illumina Nextseq500 platform from each tumor region to decipher a potential evolutional progress of SMPLC. Single or pair-end reads were first mapped to a human genome reference and filtered based on the mapping quality score. The read depth was ≥ 1 000× and the depth of coverage was 95%. The data revealed a discordant epidermal growth factor receptor (EGFR) from the separate lungs; additionally, a high frequency of point mutation on exon 9 H310P of the ERG gene was detected at the both sites of the tumor lesions. This case showed that a potential role of the molecular features analysis from each tumor lesion might contribute to the understanding of the evolutional development of SMPLC. This study suggests that the same environment may contribute certain gene(s) mutations in the same sites in the early stages of polyclonal tumor origins; meanwhile the extensive studies on these genes may help us understand the evolution and progress of tumor clones.


Asunto(s)
Adenocarcinoma , Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Neoplasias Primarias Múltiples , Anciano , Humanos , Neoplasias Pulmonares/genética , Masculino , Neoplasias Primarias Múltiples/genética , Mutación Puntual , Regulador Transcripcional ERG
18.
BMC Bioinformatics ; 21(1): 467, 2020 Oct 20.
Artículo en Inglés | MEDLINE | ID: mdl-33081688

RESUMEN

BACKGROUND: Genomic profiling of solid human tumors by projects such as The Cancer Genome Atlas (TCGA) has provided important information regarding the somatic alterations that drive cancer progression and patient survival. Although researchers have successfully leveraged TCGA data to build prognostic models, most efforts have focused on specific cancer types and a targeted set of gene-level predictors. Less is known about the prognostic ability of pathway-level variables in a pan-cancer setting. To address these limitations, we systematically evaluated and compared the prognostic ability of somatic point mutation (SPM) and copy number variation (CNV) data, gene-level and pathway-level models for a diverse set of TCGA cancer types and predictive modeling approaches. RESULTS: We evaluated gene-level and pathway-level penalized Cox proportional hazards models using SPM and CNV data for 29 different TCGA cohorts. We measured predictive accuracy as the concordance index for predicting survival outcomes. Our comprehensive analysis suggests that the use of pathway-level predictors did not offer superior predictive power relative to gene-level models for all cancer types but had the advantages of robustness and parsimony. We identified a set of cohorts for which somatic alterations could not predict prognosis, and a unique cohort LGG, for which SPM data was more predictive than CNV data and the predictive accuracy is good for all model types. We found that the pathway-level predictors provide superior interpretative value and that there is often a serious collinearity issue for the gene-level models while pathway-level models avoided this issue. CONCLUSION: Our comprehensive analysis suggests that when using somatic alterations data for cancer prognosis prediction, pathway-level models are more interpretable, stable and parsimonious compared to gene-level models. Pathway-level models also avoid the issue of collinearity, which can be serious for gene-level somatic alterations. The prognostic power of somatic alterations is highly variable across different cancer types and we have identified a set of cohorts for which somatic alterations could not predict prognosis. In general, CNV data predicts prognosis better than SPM data with the exception of the LGG cohort.


Asunto(s)
Variaciones en el Número de Copia de ADN/genética , Mutación Puntual/genética , Humanos , Pronóstico
19.
Proc Natl Acad Sci U S A ; 117(38): 23652-23662, 2020 09 22.
Artículo en Inglés | MEDLINE | ID: mdl-32868447

RESUMEN

The magnitude of the COVID-19 pandemic underscores the urgency for a safe and effective vaccine. Many vaccine candidates focus on the Spike protein, as it is targeted by neutralizing antibodies and plays a key role in viral entry. Here we investigate the diversity seen in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequences and compare it to the sequence on which most vaccine candidates are based. Using 18,514 sequences, we perform phylogenetic, population genetics, and structural bioinformatics analyses. We find limited diversity across SARS-CoV-2 genomes: Only 11 sites show polymorphisms in >5% of sequences; yet two mutations, including the D614G mutation in Spike, have already become consensus. Because SARS-CoV-2 is being transmitted more rapidly than it evolves, the viral population is becoming more homogeneous, with a median of seven nucleotide substitutions between genomes. There is evidence of purifying selection but little evidence of diversifying selection, with substitution rates comparable across structural versus nonstructural genes. Finally, the Wuhan-Hu-1 reference sequence for the Spike protein, which is the basis for different vaccine candidates, matches optimized vaccine inserts, being identical to an ancestral sequence and one mutation away from the consensus. While the rapid spread of the D614G mutation warrants further study, our results indicate that drift and bottleneck events can explain the minimal diversity found among SARS-CoV-2 sequences. These findings suggest that a single vaccine candidate should be efficacious against currently circulating lineages.


Asunto(s)
Betacoronavirus/genética , Genoma Viral , Vacunas Virales/genética , Betacoronavirus/inmunología , Infecciones por Coronavirus/genética , Infecciones por Coronavirus/prevención & control , Variación Genética , Humanos , Pandemias/prevención & control , Neumonía Viral/prevención & control , Mutación Puntual , Selección Genética
20.
PLoS One ; 15(9): e0238529, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32966289

RESUMEN

Amelanotic/hypomelanotic melanoma is a clinicopathologic subtype with absent or minimal melanin. This study assessed previously reported coding variants in albinism genes (TYR, OCA2, TYRP1, SLC45A2, SLC24A5, LRMDA) and common intronic, regulatory variants of OCA2 in individuals with amelanotic/hypomelanotic melanoma, pigmented melanoma cases and controls. Exome sequencing was available for 28 individuals with amelanotic/hypomelanotic melanoma and 303 individuals with pigmented melanoma, which were compared to whole exome data from 1144 Australian controls. Microarray genotyping was available for a further 17 amelanotic/hypomelanotic melanoma, 86 pigmented melanoma, 147 melanoma cases (pigmentation unknown) and 652 unaffected controls. Rare deleterious variants in TYR/OCA1 were more common in amelanotic/hypomelanotic melanoma cases than pigmented melanoma cases (set mixed model association tests P = 0.0088). The OCA2 hypomorphic allele p.V443I was more common in melanoma cases (1.8%) than controls (1.0%, X2 P = 0.02), and more so in amelanotic/hypomelanotic melanoma (4.4%, X2 P = 0.007). No amelanotic/hypomelanotic melanoma cases carried an eye and skin darkening haplotype of OCA2 (including rs7174027), present in 7.1% of pigmented melanoma cases (P = 0.0005) and 9.4% controls. Variants in TYR and OCA2 may play a role in amelanotic/hypomelanotic melanoma susceptibility. We suggest that somatic loss of function at these loci could contribute to the loss of tumor pigmentation, consistent with this we found a higher rate of somatic mutation in TYR/OCA2 in amelanotic/hypomelanotic melanoma vs pigmented melanoma samples (28.6% vs 3.0%; P = 0.021) from The Cancer Genome Atlas Skin Cutaneous Melanoma collection.


Asunto(s)
Albinismo/genética , Melanoma/genética , Proteínas de Transporte de Membrana/genética , Monofenol Monooxigenasa/genética , Neoplasias Cutáneas/genética , Variación Genética , Mutación de Línea Germinal , Humanos , Mutación Puntual , Polimorfismo de Nucleótido Simple , Secuenciación del Exoma Completo
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