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1.
Gene ; 766: 145146, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-32941952

RESUMO

The removal of introns from mRNA precursors (pre-mRNAs) is an essential step in eukaryotic gene expression. The splicing machinery heavily contributes to biological complexity and especially to the ability of cells to adapt to altered cellular conditions. Hypoxia also plays a key role in the pathophysiology of many diseases, including Alzheimer's disease (AD). In the presented study, we have examined the influence of cellular hypoxia on mRNA splice variant formation from Alzheimer's disease-related Tau and APP genes in brain cells. We have shown that the hypoxic microenvironment influenced the formation of Tau mRNA splice variants, but had no effect on APP mRNA splice variant formation. Additionally, our presented results indicate that splicing factor SRSF1 but not SRSF5 alters the formation of Tau cellular mRNA splice variants in hypoxic cells. Obtained results have also shown that hypoxic brain cells possess enhanced CLK1-4 kinase mRNA levels. This study underlines that cellular hypoxia can influence disease development through changing pre-mRNA splicing.


Assuntos
Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Hipóxia Celular/genética , RNA Mensageiro/genética , Proteínas tau/genética , Processamento Alternativo/genética , Encéfalo/metabolismo , Linhagem Celular Tumoral , Humanos , Íntrons/genética , Precursores de RNA/genética , Transcrição Genética/genética
2.
BMC Bioinformatics ; 21(1): 478, 2020 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-33099301

RESUMO

BACKGROUND: Introns have been shown to be spliced in a defined order, and this order influences both alternative splicing regulation and splicing fidelity, but previous studies have only considered neighbouring introns. The detailed intron splicing order remains unknown. RESULTS: In this work, a method was developed that can calculate the intron splicing orders of all introns in each transcript. A simulation study showed that this method can accurately calculate intron splicing orders. I further applied this method to real S. pombe, fruit fly, Arabidopsis thaliana, and human sequencing datasets and found that intron splicing orders change from gene to gene and that humans contain more not in-order spliced transcripts than S. pombe, fruit fly and Arabidopsis thaliana. In addition, I reconfirmed that the first introns in humans are spliced slower than those in S. pombe, fruit fly, and Arabidopsis thaliana genome-widely. Both the calculated most likely orders and the method developed here are available on the web. CONCLUSIONS: A novel computational method was developed to calculate the intron splicing orders and applied the method to real sequencing datasets. I obtained intron splicing orders for hundreds or thousands of genes in four organisms. I found humans contain more number of not in-order spliced transcripts.


Assuntos
Arabidopsis/genética , Biologia Computacional/métodos , Drosophila melanogaster/genética , Íntrons/genética , Processamento de RNA/genética , Schizosaccharomyces/genética , Processamento Alternativo , Animais , Sequência de Bases , Humanos
3.
BMC Med Genet ; 21(1): 191, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-33004005

RESUMO

BACKGROUND: Central nervous system (CNS) hemangioblastomas are the most frequent cause of mortality in patients with Von Hippel-Lindau (VHL) disease, an autosomal dominant genetic disease resulting from germline mutations in the VHL tumor suppressor gene, with most mutations occurring in the exons. To date, there have been no reports of CNS hemangioblastoma cases related to pathogenic variants in intron 2 of VHL, which encodes a tumor suppressor protein (i.e., pVHL) that regulates hypoxia-inducible factor proteins. CASE PRESENTATION: We report the presence of a base substitution of c.464-1G > C and c.464-2A > G in the intron 2 of VHL causing CNS hemangioblastomas in six patients with VHL from two Chinese families. The clinical information about the two pathogentic variants has been submitted to ClinVar database. The ClinVar accession for NM_000551.3(VHL):c.464-1G > C was SCV001371687. This finding may provide a new approach for diagnosing and researching VHL-associated hemangioblastomas. CONCLUSIONS: This is the first report of a pathogenic variant at intron 2 in VHL-associated hemangioblastomas. Gene sequencing showed that not only exonic but also intronic mutations can lead to the development of CNS hemangioblastomas.


Assuntos
Neoplasias do Sistema Nervoso Central/genética , Hemangioblastoma/genética , Mutação , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Doença de von Hippel-Lindau/genética , Adulto , Grupo com Ancestrais do Continente Asiático/genética , Sequência de Bases , Neoplasias do Sistema Nervoso Central/diagnóstico por imagem , Neoplasias do Sistema Nervoso Central/etnologia , China , Saúde da Família , Feminino , Hemangioblastoma/diagnóstico por imagem , Hemangioblastoma/etnologia , Humanos , Íntrons/genética , Imagem por Ressonância Magnética/métodos , Masculino , Pessoa de Meia-Idade , Linhagem , Análise de Sequência de DNA , Doença de von Hippel-Lindau/diagnóstico por imagem , Doença de von Hippel-Lindau/etnologia
4.
Nat Commun ; 11(1): 5060, 2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-33033246

RESUMO

Fusion oncogenes (FOs) are common in many cancer types and are powerful drivers of tumor development. Because their expression is exclusive to cancer cells and their elimination induces cell apoptosis in FO-driven cancers, FOs are attractive therapeutic targets. However, specifically targeting the resulting chimeric products is challenging. Based on CRISPR/Cas9 technology, here we devise a simple, efficient and non-patient-specific gene-editing strategy through targeting of two introns of the genes involved in the rearrangement, allowing for robust disruption of the FO specifically in cancer cells. As a proof-of-concept of its potential, we demonstrate the efficacy of intron-based targeting of transcription factors or tyrosine kinase FOs in reducing tumor burden/mortality in in vivo models. The FO targeting approach presented here might open new horizons for the selective elimination of cancer cells.


Assuntos
Sistemas CRISPR-Cas/genética , Neoplasias/genética , Fusão Oncogênica/genética , Animais , Sequência de Bases , Linhagem Celular Tumoral , Proliferação de Células/genética , Doxorrubicina/uso terapêutico , Proteínas de Fusão bcr-abl/genética , Deleção de Genes , Loci Gênicos , Instabilidade Genômica , Células HEK293 , Humanos , Íntrons/genética , Camundongos Nus , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Proteínas de Fusão Oncogênica/genética , RNA Guia/metabolismo , Reprodutibilidade dos Testes , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Mol Cell ; 80(1): 127-139.e6, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-33007253

RESUMO

Human spliceosomes contain numerous proteins absent in yeast, whose functions remain largely unknown. Here we report a 3D cryo-EM structure of the human spliceosomal C complex at 3.4 Å core resolution and 4.5-5.7 Å at its periphery, and aided by protein crosslinking we determine its molecular architecture. Our structure provides additional insights into the spliceosome's architecture between the catalytic steps of splicing, and how proteins aid formation of the spliceosome's catalytically active RNP (ribonucleoprotein) conformation. It reveals the spatial organization of the metazoan-specific proteins PPWD1, WDR70, FRG1, and CIR1 in human C complexes, indicating they stabilize functionally important protein domains and RNA structures rearranged/repositioned during the Bact to C transition. Structural comparisons with human Bact, C∗, and P complexes reveal an intricate cascade of RNP rearrangements during splicing catalysis, with intermediate RNP conformations not found in yeast, and additionally elucidate the structural basis for the sequential recruitment of metazoan-specific spliceosomal proteins.


Assuntos
Fatores de Processamento de RNA/química , Fatores de Processamento de RNA/metabolismo , Spliceossomos/metabolismo , Animais , Catálise , Células HeLa , Humanos , Íntrons/genética , Modelos Moleculares , Complexos Multiproteicos/metabolismo , Complexos Multiproteicos/ultraestrutura , Ligação Proteica , Estabilidade Proteica , RNA/química , RNA/metabolismo , Ribonucleoproteínas/metabolismo , Saccharomyces cerevisiae/metabolismo , Especificidade da Espécie , Fatores de Tempo
6.
DNA Cell Biol ; 39(11): 2095-2101, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33016778

RESUMO

Angiotensin-converting enzyme 2 (ACE2) is known as the counter-regulator of the renin-angiotensin system, it cleaves angiotensin II to render Ag 1-7, a potent vasodilator with multiple roles in cardiovascular protection. A few studies have pinpointed ACE2 polymorphisms and their relationship with heart function and hypertension in a sex-dependent manner. These observations still lack replication mostly for admixed populations. This study aimed to report minor allele frequencies of four ACE2 intron variants, rs2285666, rs2048683, rs2106809, and rs4240157, derived from previous research using the GSA, v1.0, microarray in 1231 hypertensive and nonhypertensive patients. Logistic and multiple linear regression models were developed to identify potential associations with hypertension status and systolic and diastolic blood pressure (SBP and DBP). Allele frequency differences were identified for ACE2 rs2048683 and rs4240157 in populations with European ancestry and people of the Americas. Regression analyses identified a significant association of ACE2 rs2048683 and rs4240157 with SBP/DBP in males or females. Our observations confirm sex differences in ACE2 genetic associations with SBP and DBP and contribute to the collection of genetic variation in ACE2 for admixed populations.


Assuntos
Pressão Sanguínea/genética , Hipertensão Essencial/genética , Predisposição Genética para Doença , Peptidil Dipeptidase A/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Grupo com Ancestrais do Continente Asiático/genética , Hipertensão Essencial/patologia , Feminino , Frequência do Gene , Estudos de Associação Genética , Genótipo , Humanos , Íntrons/genética , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genética , Adulto Jovem
7.
PLoS Genet ; 16(9): e1009027, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32966296

RESUMO

The availability of genomes for many species has advanced our understanding of the non-protein-coding fraction of the genome. Comparative genomics has proven itself to be an invaluable approach for the systematic, genome-wide identification of conserved non-protein-coding elements (CNEs). However, for many non-mammalian model species, including chicken, our capability to interpret the functional importance of variants overlapping CNEs has been limited by current genomic annotations, which rely on a single information type (e.g. conservation). We here studied CNEs in chicken using a combination of population genomics and comparative genomics. To investigate the functional importance of variants found in CNEs we develop a ch(icken) Combined Annotation-Dependent Depletion (chCADD) model, a variant effect prediction tool first introduced for humans and later on for mouse and pig. We show that 73 Mb of the chicken genome has been conserved across more than 280 million years of vertebrate evolution. The vast majority of the conserved elements are in non-protein-coding regions, which display SNP densities and allele frequency distributions characteristic of genomic regions constrained by purifying selection. By annotating SNPs with the chCADD score we are able to pinpoint specific subregions of the CNEs to be of higher functional importance, as supported by SNPs found in these subregions are associated with known disease genes in humans, mice, and rats. Taken together, our findings indicate that CNEs harbor variants of functional significance that should be object of further investigation along with protein-coding mutations. We therefore anticipate chCADD to be of great use to the scientific community and breeding companies in future functional studies in chicken.


Assuntos
Galinhas/genética , DNA Intergênico/genética , Genômica/métodos , Alelos , Animais , Sequência Conservada/genética , DNA Intergênico/metabolismo , Evolução Molecular , Frequência do Gene/genética , Variação Genética/genética , Genoma/genética , Íntrons/genética , Metagenômica/métodos , Polimorfismo de Nucleotídeo Único/genética , Análise de Sequência/métodos
8.
PLoS One ; 15(8): e0236759, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32745105

RESUMO

The fall armyworm (Spodoptera frugiperda) is a moth pest native to the Western Hemisphere that has recently become a global problem, invading Africa, Asia, and Australia. The species has a broad host range, long-distance migration capability, and a propensity for the generation of pesticide resistance traits that make it a formidable invasive threat and a difficult pest to control. While fall armyworm migration has been extensively studied in North America, where annual migrations of thousands of kilometers are the norm, migration patterns in South America are less understood. As a first step to address this issue we have been genetically characterizing fall armyworm populations in Ecuador, a country in the northern portion of South America that has not been extensively surveyed for this pest. These studies confirm and extend past findings indicating similarities in the fall armyworm populations from Ecuador, Trinidad-Tobago, Peru, and Bolivia that suggest substantial migratory interactions. Specifically, we found that populations throughout Ecuador are genetically homogeneous, indicating that the Andes mountain range is not a long-term barrier to fall armyworm migration. Quantification of genetic variation in an intron sequence describe patterns of similarity between fall armyworm from different locations in South America with implications for how migration might be occurring. In addition, we unexpectedly found these observations only apply to one subset of fall armyworm (the C-strain), as the other group (R-strain) was not present in Ecuador. The results suggest differences in migration behavior between fall armyworm groups in South America that appear to be related to differences in host plant preferences.


Assuntos
Haplótipos/genética , Spodoptera/genética , Migração Animal , Animais , Equador , Complexo IV da Cadeia de Transporte de Elétrons/genética , Marcadores Genéticos , Íntrons/genética , Controle de Pragas , Filogenia , Filogeografia , América do Sul
9.
Nucleic Acids Res ; 48(15): 8724-8739, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32735645

RESUMO

T cell activation is a well-established model for studying cellular responses to exogenous stimulation. Motivated by our previous finding that intron retention (IR) could lead to transcript instability, in this study, we performed BruChase-Seq to experimentally monitor the expression dynamics of nascent transcripts in resting and activated CD4+ T cells. Computational modeling was then applied to quantify the stability of spliced and intron-retained transcripts on a genome-wide scale. Beyond substantiating that intron-retained transcripts were considerably less stable than spliced transcripts, we found a global stabilization of spliced mRNAs upon T cell activation, although the stability of intron-retained transcripts remained relatively constant. In addition, we identified that La-related protein 4 (LARP4), an RNA-binding protein (RBP) known to enhance mRNA stability, was involved in T cell activation-dependent mRNA stabilization. Knocking out Larp4 in mice destabilized Nfκb1 mRNAs and reduced secretion of interleukin-2 (IL2) and interferon-gamma (IFNγ), two factors critical for T cell proliferation and function. We propose that coordination between splicing regulation and mRNA stability may provide a novel paradigm to control spatiotemporal gene expression during T cell activation.


Assuntos
Interferon gama/genética , Interleucina-2/genética , Proteínas/genética , Estabilidade de RNA/genética , Transcriptoma/genética , Processamento Alternativo/genética , Animais , Humanos , Íntrons/genética , Ativação Linfocitária/genética , Camundongos , NF-kappa B/genética , Ligação Proteica/genética , RNA Mensageiro/genética , Linfócitos T/metabolismo
10.
PLoS One ; 15(8): e0237367, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32810148

RESUMO

Bacterial group II introns mostly behave as versatile retromobile genetic elements going through distinct cycles of gain and loss. These large RNA molecules are also ribozymes splicing autocatalytically from their interrupted pre-mRNA transcripts by two different concurrent pathways, branching and circularization. These two splicing pathways were shown to release in bacterial cells significant amounts of branched intron lariats and perfect end-to-end intron circles respectively. On one hand, released intron lariats can invade new sites in RNA and/or DNA by reverse branching while released intron circles are dead end spliced products since they cannot reverse splice through circularization. The presence of two parallel and competing group II intron splicing pathways in bacteria led us to investigate the conditions that influence the overall circle to lariat ratio in vivo. Here we unveil that removing a prominent processing site within the Ll.LtrB group II intron, raising growth temperature of Lactococcus lactis host cells and increasing the expression level of the intron-interrupted gene all increased the relative amount of released intron circles compared to lariats. Strengthening and weakening the base pairing interaction between the intron and its upstream exon respectively increased and decreased the overall levels of released intron circles in comparison to lariats. Host environment was also found to impact the circle to lariat ratio of the Ll.LtrB and Ll.RlxA group II introns from L. lactis and the Ef.PcfG intron from Enterococcus faecalis. Overall, our data show that multiple factors significantly influence the balance between released intron circles and lariats in bacterial cells.


Assuntos
Íntrons/genética , Lactococcus lactis/genética , Regulação Bacteriana da Expressão Gênica , Temperatura
11.
Parasitol Res ; 119(8): 2485-2494, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32617724

RESUMO

Echinostomes are a diverse group of digenetic trematodes that are difficult to classify by predominantly traditional techniques and contain many cryptic species. Application of contemporary genetic/molecular markers can provide an alternative choice for comprehensive classification or systematic analysis. In this study, we successfully characterized the intron 5 of domain 1 of the taurocyamine kinase gene (TkD1Int5) of Artyfechinostomum malayanum and the other two species of the 37 collar-spined group, Echinostoma revolutum and Echinostoma miyagawai, whereas TkD1Int5 of Hypoderaeum conoideum cannot be amplified. High levels of nucleotide polymorphism were detected in TkD1Int5 within E. revolutum and E. miyagawai, but not in A. malayanum. Thus, TkD1Int5 can be potentially used as genetic marker for genetic investigation of E. miyagawai and E. revolutum. We therefore used TkD1Int5 to explore genetic variation within and genetic differentiation between 58 samples of E. miyagawai and five samples of E. revolutum. Heterozygosity was observed in 17 and two samples with 16 and three insertion/deletion (indel) patterns in E. miyagawai and E. revolutum, respectively. Heterozygous samples were then cloned and nucleotide sequence was performed revealing the combined haplotypes in a particular sample. Based on nucleotide variable sites (excluding indels), the 72 E. miyagawai and seven E. revolutum haplotypes were subsequently classified. The haplotype network revealed clear genetic differentiation between E. miyagawai and E. revolutum haplogroups, but no genetic structure correlated with geographical localities was detected. High polymorphism and heterogeneity of the TkD1Int5 sequence found in our study suggest that it can be used in subsequent studies as an alternate independent potential genetic marker to investigate the population genetics, genetic structure, and possible hybridization of the other echinostomes, especially the 37 collar-spined group distributed worldwide.


Assuntos
Echinostoma/genética , Variação Genética , Íntrons/genética , Animais , Echinostoma/classificação , Haplótipos
12.
Nat Commun ; 11(1): 3304, 2020 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-32620809

RESUMO

A main assumption of molecular population genetics is that genomic mutation rate does not depend on sequence function. Challenging this assumption, a recent study has found a reduction in the mutation rate in exons compared to introns in somatic cells, ascribed to an enhanced exonic mismatch repair system activity. If this reduction happens also in the germline, it can compromise studies of population genomics, including the detection of selection when using introns as proxies for neutrality. Here we compile and analyze published germline de novo mutation data to test if the exonic mutation rate is also reduced in germ cells. After controlling for sampling bias in datasets with diseased probands and extended nucleotide context dependency, we find no reduction in the mutation rate in exons compared to introns in the germline. Therefore, there is no evidence that enhanced exonic mismatch repair activity determines the mutation rate in germline cells.


Assuntos
Éxons/genética , Mutação em Linhagem Germinativa , Íntrons/genética , Taxa de Mutação , Algoritmos , Reparo de Erro de Pareamento de DNA/genética , Evolução Molecular , Células Germinativas/metabolismo , Humanos , Modelos Genéticos , Mutação , Sequenciamento Completo do Exoma/métodos
13.
PLoS Genet ; 16(7): e1008944, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32730252

RESUMO

Efficient nuclear transgene expression in the green microalga Chlamydomonas reinhardtii is generally hindered by low transcription rates. Introns can increase transcript abundance by a process called Intron-Mediated Enhancement (IME) in this alga and has been broadly observed in other eukaryotes. However, the mechanisms of IME in microalgae are poorly understood. Here, we identified 33 native introns from highly expressed genes in C. reinhardtii selected from transcriptome studies as well as 13 non-native introns. We investigated their IME capacities and probed the mechanism of action by modification of splice sites, internal sequence motifs, and position within transgenes. Several introns were found to elicit strong IME and found to be broadly applicable in different expression constructs. We determined that IME in C. reinhardtii exclusively occurs from introns within transcribed ORFs regardless of the promoter and is not induced by traditional enhancers of transcription. Our results elucidate some mechanistic details of IME in C. reinhardtii, which are similar to those observed in higher plants yet underly distinctly different induction processes. Our findings narrow the focus of targets responsible for algal IME and provides evidence that introns are underestimated regulators of C. reinhardtii nuclear gene expression.


Assuntos
Chlamydomonas reinhardtii/genética , Íntrons/genética , Processamento de Proteína Pós-Traducional/genética , Processamento de RNA/genética , Regulação da Expressão Gênica de Plantas/genética , Microalgas/genética , Regiões Promotoras Genéticas , Transcriptoma/genética
14.
BMC Evol Biol ; 20(1): 85, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32664916

RESUMO

BACKGROUND: ATP-binding cassette (ABC) transporters are involved in the active transportation of various endogenous or exogenous substances. Two ABCG2 gene subfamily members have been identified in birds. A detailed comparative study of the ABCG2 and ABCG2-like genes aid our understanding of their evolutionary history at the molecular level and provide a theoretical reference for studying the specific functions of ABCG2 and ABCG2-like genes in birds. RESULTS: We first identified 77 ABCG2/ABCG2-like gene sequences in the genomes of 41 birds. Further analysis showed that both the nucleic acid and amino acid sequences of ABCG2 and ABCG2-like genes were highly conserved and exhibited high homology in birds. However, significant differences in the N-terminal structure were found between the ABCG2 and ABCG2-like amino acid sequences. A selective pressure analysis showed that the ABCG2 and ABCG2-like genes were affected by purifying selection during the process of bird evolution. CONCLUSIONS: We believe that multiple members of the ABCG2 gene subfamily exist on chromosome 4 in the ancestors of birds. Over the long course of evolution, only the ABCG2 gene was retained on chromosome 4 in birds. The ABCG2-like gene on chromosome 6 might have originated from chromosome replication or fusion. The structural differences between the N terminus of ABCG2 protein and those of ABCG2-like proteins might lead to functional differences between the corresponding genes.


Assuntos
Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Aves/genética , Evolução Molecular , Homologia de Sequência de Aminoácidos , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/química , Sequência de Aminoácidos , Animais , Cromossomos/genética , Sequência Conservada/genética , Éxons/genética , Regulação da Expressão Gênica , Genoma , Íntrons/genética , Família Multigênica , Fases de Leitura Aberta/genética , Fosforilação , Filogenia , Domínios Proteicos , Sítios de Splice de RNA/genética , Seleção Genética , Sintenia/genética
15.
Nat Commun ; 11(1): 3354, 2020 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-32620797

RESUMO

Expansion of an intronic (GGGGCC)n repeat region within the C9orf72 gene is a main cause of familial amyotrophic lateral sclerosis and frontotemporal dementia (c9ALS/FTD). A hallmark of c9ALS/FTD is the accumulation of misprocessed RNAs, which are often targets of cellular RNA surveillance. Here, we show that RNA decay mechanisms involving upstream frameshift 1 (UPF1), including nonsense-mediated decay (NMD), are inhibited in c9ALS/FTD brains and in cultured cells expressing either of two arginine-rich dipeptide repeats (R-DPRs), poly(GR) and poly(PR). Mechanistically, although R-DPRs cause the recruitment of UPF1 to stress granules, stress granule formation is independent of NMD inhibition. Instead, NMD inhibition is primarily a result from global translational repression caused by R-DPRs. Overexpression of UPF1, but none of its NMD-deficient mutants, enhanced the survival of neurons treated by R-DPRs, suggesting that R-DPRs cause neurotoxicity in part by inhibiting cellular RNA surveillance.


Assuntos
Esclerose Amiotrófica Lateral/genética , Proteína C9orf72/genética , Demência Frontotemporal/genética , Degradação do RNAm Mediada por Códon sem Sentido , RNA Helicases/metabolismo , Transativadores/metabolismo , Esclerose Amiotrófica Lateral/patologia , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Expansão das Repetições de DNA , Conjuntos de Dados como Assunto , Embrião de Mamíferos , Feminino , Lobo Frontal/patologia , Demência Frontotemporal/patologia , Humanos , Íntrons/genética , Camundongos , Neurônios/metabolismo , Neurônios/patologia , Cultura Primária de Células , Biossíntese de Proteínas , RNA Mensageiro/metabolismo , RNA-Seq , Transativadores/genética
16.
In Vivo ; 34(3 Suppl): 1629-1632, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: covidwho-534630

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a positive-sense single-stranded RNA virus. It is contagious in humans and is the cause of the coronavirus disease 2019 (COVID-19) pandemic. In the current analysis, we searched for SARS-CoV-2 sequences within the human genome. To compare the SARS-CoV-2 genome to the human genome, we used the blast-like alignment tool (BLAT) of the University of California, Santa Cruz Genome Browser. BLAT can align a user sequence of 25 bases or more to the genome. BLAT search results revealed a 117-base pair SARS-CoV-2 sequence in the human genome with 94.6% identity. The sequence was in chromosome 1p within an intronic region of the netrin G1 (NTNG1) gene. The sequence matched a sequence in the SARS-CoV-2 orf1b (open reading frames) gene. The SARS-CoV-2 human sequence lies within non-structural proteins 14 and 15 (NSP14 and NSP15), and is quite close to the viral spike sequence, separated only by NSP16, a 904-base pair sequence. The mechanism for SARS-CoV-2 infection is the binding of the virus spike protein to the membrane-bound form of angiotensin-converting enzyme 2 and internalization of the complex by the host cell. It is probably no accident that a sequence from the SARS-CoV-2 orf1b gene is found in the human NTNG1 gene, implicated in schizophrenia, and that haloperidol, used to treat schizophrenia, may also be a treatment for COVID-19. We suggest, therefore, that it is important to investigate other haloperidol analogs. Among them are benperidol, bromperidol, bromperidol decanoate, droperidol, seperidol hydrochloride, and trifluperidol. These analogs might be valuable in the treatment of COVID-19 and other coronavirus infections.


Assuntos
Betacoronavirus/genética , Cromossomos Humanos Par 1/genética , Genes Virais , Netrina-1/genética , Proteínas Virais/genética , Animais , Antivirais/farmacologia , Antivirais/uso terapêutico , Sequência de Bases , Infecções por Coronavirus/tratamento farmacológico , DNA Complementar/genética , Endorribonucleases/genética , Exorribonucleases/genética , Haloperidol/análogos & derivados , Haloperidol/farmacologia , Haloperidol/uso terapêutico , Humanos , Íntrons/genética , Pan troglodytes/genética , Pandemias , Pneumonia Viral/tratamento farmacológico , RNA Viral/genética , Esquizofrenia/tratamento farmacológico , Esquizofrenia/genética , Alinhamento de Sequência , Homologia de Sequência do Ácido Nucleico , Especificidade da Espécie , Proteínas não Estruturais Virais/genética
18.
Nature ; 584(7822): 619-623, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32581359

RESUMO

Autoimmune thyroid disease is the most common autoimmune disease and is highly heritable1. Here, by using a genome-wide association study of 30,234 cases and 725,172 controls from Iceland and the UK Biobank, we find 99 sequence variants at 93 loci, of which 84 variants are previously unreported2-7. A low-frequency (1.36%) intronic variant in FLT3 (rs76428106-C) has the largest effect on risk of autoimmune thyroid disease (odds ratio (OR) = 1.46, P = 2.37 × 10-24). rs76428106-C is also associated with systemic lupus erythematosus (OR = 1.90, P = 6.46 × 10-4), rheumatoid factor and/or anti-CCP-positive rheumatoid arthritis (OR = 1.41, P = 4.31 × 10-4) and coeliac disease (OR = 1.62, P = 1.20 × 10-4). FLT3 encodes fms-related tyrosine kinase 3, a receptor that regulates haematopoietic progenitor and dendritic cells. RNA sequencing revealed that rs76428106-C generates a cryptic splice site, which introduces a stop codon in 30% of transcripts that are predicted to encode a truncated protein, which lacks its tyrosine kinase domains. Each copy of rs76428106-C doubles the plasma levels of the FTL3 ligand. Activating somatic mutations in FLT3 are associated with acute myeloid leukaemia8 with a poor prognosis and rs76428106-C also predisposes individuals to acute myeloid leukaemia (OR = 1.90, P = 5.40 × 10-3). Thus, a predicted loss-of-function germline mutation in FLT3 causes a reduction in full-length FLT3, with a compensatory increase in the levels of its ligand and an increased disease risk, similar to that of a gain-of-function mutation.


Assuntos
Códon sem Sentido/genética , Predisposição Genética para Doença/genética , Ligantes , Mutação , Tireoidite Autoimune/genética , Tirosina Quinase 3 Semelhante a fms/genética , Tirosina Quinase 3 Semelhante a fms/metabolismo , Alelos , Doenças Autoimunes/genética , Bases de Dados Factuais , Estudo de Associação Genômica Ampla , Mutação em Linhagem Germinativa , Humanos , Islândia , Íntrons/genética , Leucemia Mieloide Aguda , Mutação com Perda de Função , Sítios de Splice de RNA/genética , Reino Unido
19.
Nucleic Acids Res ; 48(12): 6889-6905, 2020 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-32479602

RESUMO

Mutations in the RNA-binding protein FUS cause amyotrophic lateral sclerosis (ALS), a devastating neurodegenerative disease. FUS plays a role in numerous aspects of RNA metabolism, including mRNA splicing. However, the impact of ALS-causative mutations on splicing has not been fully characterized, as most disease models have been based on overexpressing mutant FUS, which will alter RNA processing due to FUS autoregulation. We and others have recently created knockin models that overcome the overexpression problem, and have generated high depth RNA-sequencing on FUS mutants in parallel to FUS knockout, allowing us to compare mutation-induced changes to genuine loss of function. We find that FUS-ALS mutations induce a widespread loss of function on expression and splicing. Specifically, we find that mutant FUS directly alters intron retention levels in RNA-binding proteins. Moreover, we identify an intron retention event in FUS itself that is associated with its autoregulation. Altered FUS levels have been linked to disease, and we show here that this novel autoregulation mechanism is altered by FUS mutations. Crucially, we also observe this phenomenon in other genetic forms of ALS, including those caused by TDP-43, VCP and SOD1 mutations, supporting the concept that multiple ALS genes interact in a regulatory network.


Assuntos
Esclerose Amiotrófica Lateral/genética , Homeostase/genética , Proteína FUS de Ligação a RNA/genética , Animais , Citoplasma/genética , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Regulação da Expressão Gênica/genética , Humanos , Íntrons/genética , Mutação com Perda de Função , Camundongos , Camundongos Knockout , Mutação/genética , Processamento de RNA/genética , Superóxido Dismutase-1/genética , Proteína com Valosina/genética
20.
PLoS Genet ; 16(6): e1008830, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32502192

RESUMO

Many post-transcriptional mechanisms operate via mRNA 3'UTRs to regulate protein expression, and such controls are crucial for development. We show that homozygous mutations in two zebrafish exon junction complex (EJC) core genes rbm8a and magoh leads to muscle disorganization, neural cell death, and motor neuron outgrowth defects, as well as dysregulation of mRNAs subjected to nonsense-mediated mRNA decay (NMD) due to translation termination ≥ 50 nts upstream of the last exon-exon junction. Intriguingly, we find that EJC-dependent NMD also regulates a subset of transcripts that contain 3'UTR introns (3'UI) < 50 nts downstream of a stop codon. Some transcripts containing such stop codon-proximal 3'UI are also NMD-sensitive in cultured human cells and mouse embryonic stem cells. We identify 167 genes that contain a conserved proximal 3'UI in zebrafish, mouse and humans. foxo3b is one such proximal 3'UI-containing gene that is upregulated in zebrafish EJC mutant embryos, at both mRNA and protein levels, and loss of foxo3b function in EJC mutant embryos significantly rescues motor axon growth defects. These data are consistent with EJC-dependent NMD regulating foxo3b mRNA to control protein expression during zebrafish development. Our work shows that the EJC is critical for normal zebrafish development and suggests that proximal 3'UIs may serve gene regulatory function in vertebrates.


Assuntos
Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Neurogênese/genética , Degradação do RNAm Mediada por Códon sem Sentido/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Regiões 3' não Traduzidas/genética , Animais , Animais Geneticamente Modificados , Axônios/fisiologia , Códon de Terminação , Conjuntos de Dados como Assunto , Embrião não Mamífero , Éxons/genética , Redes Reguladoras de Genes/genética , Homozigoto , Humanos , Íntrons/genética , Camundongos , Músculo Esquelético/inervação , Mutagênese , Mutação , Crescimento Neuronal/genética , Proteínas Nucleares/genética , Terminação Traducional da Cadeia Peptídica , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genética , RNA-Seq , Alinhamento de Sequência , Regulação para Cima , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento
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