Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 27.900
Filtrar
1.
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
2.
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
3.
Nat Commun ; 11(1): 4140, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32811829

RESUMO

Investigating the evolution of complex phenotypes and the underlying molecular bases of their variation is critical to understand how organisms adapt to their environment. Applying classical quantitative genetics on a segregating population derived from a Can-0xCol-0 cross, we identify the MADS-box transcription factor FLOWERING LOCUS M (FLM) as a player of the phenotypic variation in plant growth and color. We show that allelic variation at FLM modulates plant growth strategy along the leaf economics spectrum, a trade-off between resource acquisition and resource conservation, observable across thousands of plant species. Functional differences at FLM rely on a single intronic substitution, disturbing transcript splicing and leading to the accumulation of non-functional FLM transcripts. Associations between this substitution and phenotypic and climatic data across Arabidopsis natural populations, show how noncoding genetic variation at a single gene might be adaptive through pleiotropic effects.


Assuntos
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Regulação da Expressão Gênica de Plantas/genética , Proteínas de Domínio MADS/genética , Proteínas de Domínio MADS/metabolismo , Processamento de RNA/genética , Alelos , Arabidopsis/metabolismo , Evolução Molecular , Pleiotropia Genética , Variação Genética , Íntrons , Fenótipo , Folhas de Planta/genética , Folhas de Planta/fisiologia , Locos de Características Quantitativas/genética , Temperatura
4.
Nat Commun ; 11(1): 4178, 2020 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-32826895

RESUMO

Friedreich's ataxia (FRDA) is an autosomal-recessive neurodegenerative and cardiac disorder which occurs when transcription of the FXN gene is silenced due to an excessive expansion of GAA repeats into its first intron. Herein, we generate dorsal root ganglia organoids (DRG organoids) by in vitro differentiation of human iPSCs. Bulk and single-cell RNA sequencing show that DRG organoids present a transcriptional signature similar to native DRGs and display the main peripheral sensory neuronal and glial cell subtypes. Furthermore, when co-cultured with human intrafusal muscle fibers, DRG organoid sensory neurons contact their peripheral targets and reconstitute the muscle spindle proprioceptive receptors. FRDA DRG organoids model some molecular and cellular deficits of the disease that are rescued when the entire FXN intron 1 is removed, and not with the excision of the expanded GAA tract. These results strongly suggest that removal of the repressed chromatin flanking the GAA tract might contribute to rescue FXN total expression and fully revert the pathological hallmarks of FRDA DRG neurons.


Assuntos
Ataxia de Friedreich/genética , Ataxia de Friedreich/patologia , Gânglios Espinais/metabolismo , Edição de Genes/métodos , Proteínas de Ligação ao Ferro/genética , Organoides/metabolismo , Células Receptoras Sensoriais/metabolismo , Antioxidantes/farmacologia , Sistemas CRISPR-Cas , Diferenciação Celular , Cromatina/metabolismo , Ataxia de Friedreich/tratamento farmacológico , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/patologia , Predisposição Genética para Doença/genética , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Íntrons , Mitocôndrias/metabolismo , Organoides/efeitos dos fármacos , Organoides/patologia , Células Receptoras Sensoriais/patologia , Análise de Sequência de RNA , Transcriptoma
5.
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
6.
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
7.
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
8.
Gene ; 762: 145016, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-32777522

RESUMO

Mcl-1 is a member of the Bcl-2 anti-apoptotic protein family with important roles in the development, lifespan and metabolism of lymphocytes, as well as oncogenesis. Mcl-1 displays the shortest half-life of all Bcl-2 family members, with miRNA interference and proteasomal degradation being major pathways for Mcl-1 downregulation. In this study, we have identified a previously undescribed control mechanism active at the RNA level. A divergently transcribed lncRNA LOC107985203 (named here mcl1-AS1) negatively modulated Mcl-1 expression resulting in downregulation of Mcl-1 at both mRNA and protein level in a time-dependent manner. Using reporter assays, we confirmed that the mcl1-AS1 lncRNA promoter was located within Mcl-1 coding region. We next placed mcl1-AS1 under tetracycline-inducible control and demonstrated decreased viability in HEK293 cells upon doxycycline induction. Inhibition of mcl1-AS1 with shRNA reversed drug sensitivity. Bioinformatics surveys predicted direct mcl1-AS1 lncRNA binding to Mcl-1 transcripts, suggesting its mechanism in Mcl-1 expression is at the transcriptional level, consistent with a common role for anti-sense transcripts. The identification of a bi-directional promoter and lncRNA controlling Mcl-1 expression will have implications for controlling Mcl-1 activity in cancer cells, or for the purpose of enhancing the lifespan and quality of anti-cancer T lymphocytes.


Assuntos
Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , RNA Antissenso/genética , RNA Longo não Codificante/genética , Éxons , Células HEK293 , Humanos , Íntrons , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Regiões Promotoras Genéticas , RNA Antissenso/metabolismo , RNA Longo não Codificante/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
9.
PLoS One ; 15(7): e0235613, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32634176

RESUMO

Germline variants inactivating the mismatch repair (MMR) genes MLH1, MSH2, MSH6 and PMS2 cause Lynch syndrome that implies an increased cancer risk, where colon and endometrial cancer are the most frequent. Identification of these pathogenic variants is important to identify endometrial cancer patients with inherited increased risk of new cancers, in order to offer them lifesaving surveillance. However, several other genes are also part of the MMR pathway. It is therefore relevant to search for variants in additional genes that may be associated with cancer risk by including all known genes involved in the MMR pathway. Next-generation sequencing was used to screen 22 genes involved in the MMR pathway in constitutional DNA extracted from full blood from 199 unselected endometrial cancer patients. Bioinformatic pipelines were developed for identification and functional annotation of variants, using several different software tools and custom programs. This facilitated identification of 22 exonic, 4 UTR and 9 intronic variants that could be classified according to pathogenicity. This study has identified several germline variants in genes of the MMR pathway that potentially may be associated with an increased risk for cancer, in particular endometrial cancer, and therefore are relevant for further investigation. We have also developed bioinformatics strategies to analyse targeted sequencing data, including low quality data and genomic regions outside of the protein coding exons of the relevant genes.


Assuntos
Reparo de Erro de Pareamento de DNA , Neoplasias do Endométrio/patologia , Endonuclease PMS2 de Reparo de Erro de Pareamento/genética , Proteína 1 Homóloga a MutL/genética , Proteína 2 Homóloga a MutS/genética , Neoplasias Colorretais Hereditárias sem Polipose/genética , Neoplasias Colorretais Hereditárias sem Polipose/patologia , Variações do Número de Cópias de DNA , DNA de Neoplasias/sangue , DNA de Neoplasias/química , DNA de Neoplasias/metabolismo , Neoplasias do Endométrio/genética , Éxons , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Íntrons , Fatores de Risco , Regiões não Traduzidas/genética
10.
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
11.
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
12.
Nat Commun ; 11(1): 3501, 2020 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-32647108

RESUMO

While most monogenic diseases are caused by loss or reduction of protein function, the need for technologies that can selectively increase levels of protein in native tissues remains. Here we demonstrate that antisense-mediated modulation of pre-mRNA splicing can increase endogenous expression of full-length protein by preventing naturally occurring non-productive alternative splicing and promoting generation of productive mRNA. Bioinformatics analysis of RNA sequencing data identifies non-productive splicing events in 7,757 protein-coding human genes, of which 1,246 are disease-associated. Antisense oligonucleotides targeting multiple types of non-productive splicing events lead to increases in productive mRNA and protein in a dose-dependent manner in vitro. Moreover, intracerebroventricular injection of two antisense oligonucleotides in wild-type mice leads to a dose-dependent increase in productive mRNA and protein in the brain. The targeting of natural non-productive alternative splicing to upregulate expression from wild-type or hypomorphic alleles provides a unique approach to treating genetic diseases.


Assuntos
Processamento Alternativo , Regulação da Expressão Gênica , Oligonucleotídeos Antissenso/farmacologia , Alelos , Animais , Animais Recém-Nascidos , Encéfalo/metabolismo , Biologia Computacional , Éxons , Feminino , Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Íntrons , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/metabolismo , Ativação Transcricional/efeitos dos fármacos , Regulação para Cima
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.
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
15.
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
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
17.
In Vivo ; 34(3 Suppl): 1629-1632, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32503821

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 , Exorribonucleases/genética , Genes Virais , Netrina-1/genética , Proteínas não Estruturais Virais/genética , Proteínas Virais/genética , Animais , Antivirais/farmacologia , Antivirais/uso terapêutico , Sequência de Bases , Infecções por Coronavirus/tratamento farmacológico , DNA Complementar/genética , Endorribonucleases/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
18.
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
19.
Nat Commun ; 11(1): 2837, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32503992

RESUMO

Group II introns are ubiquitous self-splicing ribozymes and retrotransposable elements evolutionarily and chemically related to the eukaryotic spliceosome, with potential applications as gene-editing tools. Recent biochemical and structural data have captured the intron in multiple conformations at different stages of catalysis. Here, we employ enzymatic assays, X-ray crystallography, and molecular simulations to resolve the spatiotemporal location and function of conformational changes occurring between the first and the second step of splicing. We show that the first residue of the highly-conserved catalytic triad is protonated upon 5'-splice-site scission, promoting a reversible structural rearrangement of the active site (toggling). Protonation and active site dynamics induced by the first step of splicing facilitate the progression to the second step. Our insights into the mechanism of group II intron splicing parallels functional data on the spliceosome, thus reinforcing the notion that these evolutionarily-related molecular machines share the same enzymatic strategy.


Assuntos
Íntrons/genética , Precursores de RNA/metabolismo , Processamento de RNA , RNA Bacteriano/metabolismo , Spliceossomos/metabolismo , Bacillaceae/genética , Domínio Catalítico/genética , Cristalografia por Raios X , Simulação de Dinâmica Molecular , Mutagênese , Conformação de Ácido Nucleico , Precursores de RNA/genética , RNA Bacteriano/genética , Análise Espaço-Temporal
20.
Nat Commun ; 11(1): 2845, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32504065

RESUMO

Exonic splicing enhancers (ESEs) are enriched in exons relative to introns and bind splicing activators. This study considers a fundamental question of co-evolution: How did ESE motifs become enriched in exons prior to the evolution of ESE recognition? We hypothesize that the high exon to intron motif ratios necessary for ESE function were created by mutational bias coupled with purifying selection on the protein code. These two forces retain certain coding motifs in exons while passively depleting them from introns. Through the use of simulations, genomic analyses, and high throughput splicing assays, we confirm the key predictions of this hypothesis, including an overlap between protein and splicing information in ESEs. We discuss the implications of mutational bias as an evolutionary driver in other cis-regulatory systems.


Assuntos
Elementos Facilitadores Genéticos , Evolução Molecular , Éxons/genética , Genoma Humano , Processamento de RNA , Simulação por Computador , Genômica , Ensaios de Triagem em Larga Escala , Humanos , Íntrons/genética , Modelos Genéticos , Mutação
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA