RESUMO
Upstream open reading frames (uORFs) are initiated by AUG or near-cognate start codons and have been identified in the transcript leader sequences of the majority of eukaryotic transcripts. Functionally, uORFs are implicated in downstream translational regulation of the main protein coding sequence and may serve as a source of non-canonical peptides. Genetic defects in uORF sequences have been linked to the development of various diseases, including cancer. To simplify uORF-related research, the initial release of uORFdb in 2014 provided a comprehensive and manually curated collection of uORF-related literature. Here, we present an updated sequence-based version of uORFdb, accessible at https://www.bioinformatics.uni-muenster.de/tools/uorfdb. The new uORFdb enables users to directly access sequence information, graphical displays, and genetic variation data for over 2.4 million human uORFs. It also includes sequence data of >4.2 million uORFs in 12 additional species. Multiple uORFs can be displayed in transcript- and reading-frame-specific models to visualize the translational context. A variety of filters, sequence-related information, and links to external resources (UCSC Genome Browser, dbSNP, ClinVar) facilitate immediate in-depth analysis of individual uORFs. The database also contains uORF-related somatic variation data obtained from whole-genome sequencing (WGS) analyses of 677 cancer samples collected by the TCGA consortium.
Assuntos
Bases de Dados Genéticas , Fases de Leitura Aberta , Humanos , Regiões 5' não Traduzidas , Códon de Iniciação , Eucariotos/genética , Neoplasias/genética , Fases de Leitura Aberta/genética , Biossíntese de ProteínasRESUMO
As one of the major structural constituents, mobile elements comprise more than half of the human genome, among which Alu, L1, and SVA elements are still active and continue to generate new offspring. One of the major characteristics of L1 and SVA elements is their ability to co-mobilize adjacent downstream sequences to new loci in a process called 3' DNA transduction. Transductions influence the structure and content of the genome in different ways, such as increasing genome variation, exon shuffling, and gene duplication. Moreover, given their mutagenicity capability, 3' transductions are often involved in tumorigenesis or in the development of some diseases. In this study, we analyzed 3202 genomes sequenced at high coverage by the New York Genome Center to catalog and characterize putative 3' transduced segments mediated by L1s and SVAs. Here, we present a genome-wide map of inter/intrachromosomal 3' transduction variants, including their genomic and functional location, length, progenitor location, and allelic frequency across 26 populations. In total, we identified 7103 polymorphic L1s and 3040 polymorphic SVAs. Of these, 268 and 162 variants were annotated as high-confidence L1 and SVA 3' transductions, respectively, with lengths that ranged from 7 to 997 nucleotides. We found specific loci within chromosomes X, 6, 7, and 6_GL000253v2_alt as master L1s and SVAs that had yielded more transductions, among others. Together, our results demonstrate the dynamic nature of transduction events within the genome and among individuals and their contribution to the structural variations of the human genome.
RESUMO
Upstream open reading frame (uORF)-mediated translational control has emerged as an important regulatory mechanism in human health and disease. However, a systematic search for cancer-associated somatic uORF mutations has not been performed. Here, we analyzed the genetic variability at canonical (uAUG) and alternative translational initiation sites (aTISs), as well as the associated upstream termination codons (uStops) in 3394 whole-exome-sequencing datasets from patient samples of breast, colon, lung, prostate, and skin cancer and of acute myeloid leukemia, provided by The Cancer Genome Atlas research network. We found that 66.5% of patient samples were affected by at least one of 5277 recurrent uORF-associated somatic single nucleotide variants altering 446 uAUG, 347 uStop, and 4733 aTIS codons. While twelve uORF variants were detected in all entities, 17 variants occurred in all five types of solid cancer analyzed here. Highest frequencies of individual somatic variants in the TLSs of NBPF20 and CHCHD2 reached 10.1% among LAML and 8.1% among skin cancer patients, respectively. Functional evaluation by dual luciferase reporter assays identified 19 uORF variants causing significant translational deregulation of the associated main coding sequence, ranging from 1.73-fold induction for an AUG.1 > UUG variant in SETD4 to 0.006-fold repression for a CUG.6 > GUG variant in HLA-DRB1. These data suggest that somatic uORF mutations are highly prevalent in human malignancies and that defective translational regulation of protein expression may contribute to the onset or progression of cancer.
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The cardinal virulence factor of human-pathogenic enterohaemorrhagic Escherichia coli (EHEC) is Shiga toxin (Stx), which causes severe extraintestinal complications including kidney failure by damaging renal endothelial cells. In EHEC pathogenesis, the disturbance of the kidney epithelium by Stx becomes increasingly recognised, but how this exactly occurs is unknown. To explore this molecularly, we investigated the Stx receptor content and transcriptomic profile of two human renal epithelial cell lines: highly Stx-sensitive ACHN cells and largely Stx-insensitive Caki-2 cells. Though both lines exhibited the Stx receptor globotriaosylceramide, RNAseq revealed strikingly different transcriptomic responses to an Stx challenge. Using RNAi to silence factors involved in ACHN cells' Stx response, the greatest protection occurred when silencing RAB5A and TRAPPC6B, two host factors that we newly link to Stx trafficking. Silencing these factors alongside YKT6 fully prevented the cytotoxic Stx effect. Overall, our approach reveals novel subcellular targets for potential therapies against Stx-mediated kidney failure.
Assuntos
Células Epiteliais/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Rim/efeitos dos fármacos , Toxina Shiga II/farmacologia , Proteínas de Transporte Vesicular/antagonistas & inibidores , Proteínas rab5 de Ligação ao GTP/antagonistas & inibidores , Células Cultivadas , Células Epiteliais/metabolismo , Perfilação da Expressão Gênica , Humanos , Rim/metabolismoRESUMO
Analysis of ENCODE long RNA-Seq and ChIP-seq (Chromatin Immunoprecipitation Sequencing) datasets for HepG2 and HeLa cell lines uncovered 1647 and 1958 transcripts that interfere with transcription factor binding to human enhancer domains. TFBSs (Transcription Factor Binding Sites) intersected by these 'Enhancer Occlusion Transcripts' (EOTrs) displayed significantly lower relative transcription factor (TF) binding affinities compared to TFBSs for the same TF devoid of EOTrs. Expression of most EOTrs was regulated in a cell line specific manner; analysis for the same TFBSs across cell lines, i.e. in the absence or presence of EOTrs, yielded consistently higher relative TF/DNA-binding affinities for TFBSs devoid of EOTrs. Lower activities of EOTr-associated enhancer domains coincided with reduced occupancy levels for histone tail modifications H3K27ac and H3K9ac. Similarly, the analysis of EOTrs with allele-specific expression identified lower activities for alleles associated with EOTrs. ChIA-PET (Chromatin Interaction Analysis by Paired-End Tag Sequencing) and 5C (Carbon Copy Chromosome Conformation Capture) uncovered that enhancer domains associated with EOTrs preferentially interacted with poised gene promoters. Analysis of EOTr regions with GRO-seq (Global run-on) data established the correlation of RNA polymerase pausing and occlusion of TF-binding. Our results implied that EOTr expression regulates human enhancer domains via transcriptional interference.
Assuntos
Elementos Facilitadores Genéticos , Fatores de Transcrição/metabolismo , Transcrição Gênica , Alelos , Sítios de Ligação , Cromatina/química , Sequenciamento de Cromatina por Imunoprecipitação , RNA Polimerases Dirigidas por DNA/metabolismo , Células HeLa , Células Hep G2 , Código das Histonas , Humanos , Matrizes de Pontuação de Posição Específica , Regiões Promotoras Genéticas , RNA-Seq , Fatores de Transcrição de p300-CBP/metabolismoRESUMO
Valvular heart disease is observed in approximately 2% of the general population1. Although the initial observation is often localized (for example, to the aortic or mitral valve), disease manifestations are regularly observed in the other valves and patients frequently require surgery. Despite the high frequency of heart valve disease, only a handful of genes have so far been identified as the monogenic causes of disease2-7. Here we identify two consanguineous families, each with two affected family members presenting with progressive heart valve disease early in life. Whole-exome sequencing revealed homozygous, truncating nonsense alleles in ADAMTS19 in all four affected individuals. Homozygous knockout mice for Adamts19 show aortic valve dysfunction, recapitulating aspects of the human phenotype. Expression analysis using a lacZ reporter and single-cell RNA sequencing highlight Adamts19 as a novel marker for valvular interstitial cells; inference of gene regulatory networks in valvular interstitial cells positions Adamts19 in a highly discriminatory network driven by the transcription factor lymphoid enhancer-binding factor 1 downstream of the Wnt signaling pathway. Upregulation of endocardial Krüppel-like factor 2 in Adamts19 knockout mice precedes hemodynamic perturbation, showing that a tight balance in the Wnt-Adamts19-Klf2 axis is required for proper valve maturation and maintenance.
Assuntos
Proteínas ADAMTS/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Doenças das Valvas Cardíacas/etiologia , Proteínas ADAMTS/genética , Animais , Família , Feminino , Doenças das Valvas Cardíacas/patologia , Humanos , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Masculino , Camundongos , Camundongos Knockout , Linhagem , Análise de Célula Única , Via de Sinalização WntRESUMO
Ezrin, radixin, moesin, and merlin are cytoskeletal proteins, whose functions are specific to metazoans. They participate in cell cortex rearrangement, including cell-cell contact formation, and play an important role in cancer progression. Here, we have performed a comprehensive phylogenetic analysis of the proteins spanning 87 species. The results describe a possible mechanism for the protein family origin in the root of Metazoa, paralogs diversification in vertebrates, and acquisition of novel functions, including tumor suppression. In addition, a merlin paralog, present in most vertebrates but lost in mammals, has been described here for the first time. We have also highlighted a set of amino acid variations within the conserved motifs as the candidates for determining physiological differences between ERM paralogs.
Assuntos
Proteínas do Citoesqueleto/genética , Evolução Molecular , Proteínas de Membrana/genética , Proteínas dos Microfilamentos/genética , Neurofibromina 2/genética , Motivos de Aminoácidos , Animais , Proteínas do Citoesqueleto/química , Peixes/genética , Humanos , Proteínas de Membrana/química , Proteínas dos Microfilamentos/química , Família Multigênica , Neurofibromina 2/química , Filogenia , SinteniaRESUMO
The clonal architecture of tumors plays a vital role in their pathogenesis and invasiveness; however, it is not yet clear how this clonality contributes to different malignancies. In this study we sought to address mutational intratumor heterogeneity (ITH) in adult T-cell leukemia/lymphoma (ATL). ATL is a malignancy with an incompletely understood molecular pathogenesis caused by infection with human T-cell leukemia virus type-1 (HTLV-1). To determine the clonal structure through tumor genetic diversity profiles, we investigated 142 whole-exome sequencing data of tumor and matched normal samples from 71 ATL patients. Based on SciClone analysis, the ATL samples showed a wide spectrum of modes over clonal/subclonal frequencies ranging from one to nine clusters. The average number of clusters was six across samples, but the number of clusters differed among different samples. Of these ATL samples, 94% had more than two clusters. Aggressive ATL cases had slightly more clonal clusters than indolent types, indicating the presence of ITH during earlier stages of disease. The known significantly mutated genes in ATL were frequently clustered together and possibly coexisted in the same clone. IRF4, CCR4, TP53, and PLCG1 mutations were almost clustered in subclones with a moderate variant allele frequency (VAF), whereas HLA-B, CARD11, and NOTCH1 mutations were clustered in subclones with lower VAFs. Taken together, these results show that ATL displays a high degree of ITH and a complex subclonal structure. Our findings suggest that clonal/subclonal architecture might be a useful measure for prognostic purposes and personalized assessment of the therapeutic response.
Assuntos
Heterogeneidade Genética , Predisposição Genética para Doença , Leucemia-Linfoma de Células T do Adulto/etiologia , Leucemia-Linfoma de Células T do Adulto/patologia , Mutação , Biomarcadores , Biologia Computacional/métodos , Variações do Número de Cópias de DNA , Suscetibilidade a Doenças , Variação Genética , Estudo de Associação Genômica Ampla , Infecções por HTLV-I/complicações , Infecções por HTLV-I/virologia , Vírus Linfotrópico T Tipo 1 Humano , Humanos , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Ribosome profiling revealed widespread translational activity at upstream open reading frames (uORFs) and validated uORF-mediated translational control as a commonly repressive mechanism of gene expression. Translational activation of proto-oncogenes through loss-of-uORF mutations has been demonstrated, yet a systematic search for cancer-associated genetic alterations in uORFs is lacking. Here, we applied a PCR-based, multiplex identifier-tagged deep sequencing approach to screen 404 uORF translation initiation sites of 83 human tyrosine kinases and 49 other proto-oncogenes in 308 human malignancies. We identified loss-of-function uORF mutations in EPHB1 in two samples derived from breast and colon cancer, and in MAP2K6 in a sample of colon adenocarcinoma. Both mutations were associated with enhanced translation, suggesting that loss-of-uORF-mediated translational induction of the downstream main protein coding sequence may have contributed to carcinogenesis. Computational analysis of whole exome sequencing datasets of 464 colon adenocarcinomas subsequently revealed another 53 non-recurrent somatic mutations functionally deleting 22 uORF initiation and 31 uORF termination codons, respectively. These data provide evidence for somatic mutations affecting uORF initiation and termination codons in human cancer. The insufficient coverage of uORF regions in current whole exome sequencing datasets demands for future genome-wide analyses to ultimately define the contribution of uORF-mediated translational deregulation in oncogenesis.
Assuntos
Carcinogênese/genética , Mutação , Proteínas de Neoplasias/genética , Neoplasias/genética , Fases de Leitura Aberta , Proto-Oncogenes , Regiões 5' não Traduzidas , Carcinogênese/metabolismo , Carcinogênese/patologia , Linhagem Celular Tumoral , Códon de Terminação , Genes Reporter , Estudo de Associação Genômica Ampla , Células HEK293 , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Luciferases/genética , Luciferases/metabolismo , MAP Quinase Quinase 6 , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Iniciação Traducional da Cadeia Peptídica , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Receptor EphB1/genética , Receptor EphB1/metabolismoRESUMO
Proximal promoter regions (PPR) are heavily transcribed yielding different types of small RNAs. The act of transcription within PPRs might regulate downstream gene expression via transcriptional interference (TI). For analysis, we investigated capped and polyadenylated small RNA transcripts within PPRs of human RefSeq genes in eight different cell lines. Transcripts of our datasets overlapped with experimentally determined transcription factor binding sites (TFBS). For TFBSs intersected by these small RNA transcripts, we established negative correlation of sRNA expression levels and transcription factor (TF) DNA binding affinities; suggesting that the transcripts acted via TI. Accordingly, datasets were designated as TFbiTrs (TF-binding interfering transcripts). Expression of most TFbiTrs was restricted to certain cell lines. This facilitated the analysis of effects related to TFbiTr expression for the same RefSeq genes across cell lines. We consistently uncovered higher relative TF/DNA binding affinities and concomitantly higher expression levels for RefSeq genes in the absence of TFbiTrs. Analysis of corresponding chromatin landscapes supported these results. ChIA-PET revealed the participation of distal enhancers in TFbiTr transcription. Enhancers regulating TFbiTrs, in effect, act as repressors for corresponding downstream RefSeq genes. We demonstrate the significant impact of TI on gene expression using selected small RNA datasets.
Assuntos
DNA/genética , Regiões Promotoras Genéticas , RNA Mensageiro/genética , Fatores de Transcrição/genética , Transcrição Gênica , Células A549 , Sítios de Ligação , Linhagem Celular , Cromatina/química , Cromatina/metabolismo , DNA/metabolismo , Conjuntos de Dados como Assunto , Elementos Facilitadores Genéticos , Células HeLa , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Células K562 , Células MCF-7 , Neurônios/citologia , Neurônios/metabolismo , Ligação Proteica , RNA Mensageiro/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Here, we employed cDNA amplicon sequencing using a long-read portable sequencer, MinION, to characterize various types of mutations in cancer-related genes, namely, EGFR, KRAS, NRAS and NF1. For homozygous SNVs, the precision and recall rates were 87.5% and 91.3%, respectively. For previously reported hotspot mutations, the precision and recall rates reached 100%. The precise junctions of EML4-ALK, CCDC6-RET and five other gene fusions were also detected. Taking advantages of long-read sequencing, we conducted phasing of EGFR mutations and elucidated the mutational allelic backgrounds of anti-tumor drug-sensitive and resistant mutations, which could provide useful information for selecting therapeutic approaches. In the H1975 cells, 72% of the reads harbored both L858R and T790M mutations, and 22% of the reads harbored neither mutation. To ensure that the clinical requirements can be met in potentially low cancer cell populations, we further conducted a serial dilution analysis of the template for EGFR mutations. Several percent of the mutant alleles could be detected depending on the yield and quality of the sequencing data. Finally, we characterized the mutation genotypes in eight clinical samples. This method could be a convenient long-read sequencing-based analytical approach and thus may change the current approaches used for cancer genome sequencing.
Assuntos
Adenocarcinoma/genética , Receptores ErbB/genética , Sequenciamento de Nucleotídeos em Larga Escala/instrumentação , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Neoplasias Pulmonares/genética , Mutação , Análise de Sequência de DNA/métodos , Biomarcadores Tumorais/genética , HumanosRESUMO
BACKGROUND: Human T cell leukemia virus type 1 (HTLV-1) causes adult T cell leukemia (ATL) in a proportion of infected individuals after a long latency period. Development of ATL is a multistep clonal process that can be investigated by monitoring the clonal expansion of HTLV-1-infected cells by isolation of provirus integration sites. The clonal composition (size, number, and combinations of clones) during the latency period in a given infected individual has not been clearly elucidated. METHODS: We used high-throughput sequencing technology coupled with a tag system for isolating integration sites and measuring clone sizes from 60 clinical samples. We assessed the role of clonality and clone size dynamics in ATL onset by modeling data from high-throughput monitoring of HTLV-1 integration sites using single- and multiple-time-point samples. RESULTS: From four size categories analyzed, we found that big clones (B; 513-2048 infected cells) and very big clones (VB; >2048 infected cells) had prognostic value. No sample harbored two or more VB clones or three or more B clones. We examined the role of clone size, clone combination, and the number of integration sites in the prognosis of infected individuals. We found a moderate reverse correlation between the total number of clones and the size of the largest clone. We devised a data-driven model that allows intuitive representation of clonal composition. CONCLUSIONS: This integration site-based clonality tree model represents the complexity of clonality and provides a global view of clonality data that facilitates the analysis, interpretation, understanding, and visualization of the behavior of clones on inter- and intra-individual scales. It is fully data-driven, intuitively depicts the clonality patterns of HTLV-1-infected individuals and can assist in early risk assessment of ATL onset by reflecting the prognosis of infected individuals. This model should assist in assimilating information on clonal composition and understanding clonal expansion in HTLV-1-infected individuals.
Assuntos
Gráficos por Computador , DNA Viral/genética , Vírus Linfotrópico T Tipo 1 Humano/genética , Leucemia-Linfoma de Células T do Adulto/genética , Integração Viral/genética , Adulto , Células Clonais , Estudos Transversais , Humanos , Leucemia-Linfoma de Células T do Adulto/virologia , Estudos LongitudinaisRESUMO
Current models of early mouse development assign roles to stochastic processes and epigenetic regulation, which are considered to be as influential as the genetic differences that exist between strains of the species Mus musculus. The aim of this study was to test whether mouse oocytes vary from each other in the abundance of gene products that could influence, prime, or even predetermine developmental trajectories and features of derivative embryos. Using the paradigm of inbred mouse strains, we quantified 2010 protein groups (SILAC LC-MS/MS) and 15205 transcripts (RNA deep sequencing) present simultaneously in oocytes of four strains tested (129/Sv, C57Bl/6J, C3H/HeN, DBA/2J). Oocytes differed according to donor strain in the abundance of catalytic and regulatory proteins, as confirmed for a subset (bromodomain adjacent to zinc finger domain, 1B [BAZ1B], heme oxygenase 1 [HMOX1], estrogen related receptor, beta [ESRRB]) via immunofluorescence in situ. Given a Pearson's r correlation coefficient of 0.18-0.20, the abundance of oocytic proteins could not be predicted from that of cognate mRNAs. Our results document that a prerequisite to generate embryo diversity, namely the different abundances of maternal proteins in oocytes, can be studied in the model of inbred mouse strains. Thus, we highlight the importance of proteomic quantifications in modern embryology. All MS data have been deposited in the ProteomeXchange with identifier PXD001059 (http://proteomecentral.proteomexchange.org/dataset/PXD001059).
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Embrião de Mamíferos/metabolismo , Camundongos Endogâmicos/embriologia , Oócitos/metabolismo , Proteoma/análise , Proteoma/metabolismo , Animais , Embrião de Mamíferos/química , Desenvolvimento Embrionário/fisiologia , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Sequenciamento de Nucleotídeos em Larga Escala , Marcação por Isótopo , Masculino , Espectrometria de Massas , Camundongos , Oócitos/química , Oócitos/crescimento & desenvolvimento , Proteoma/química , Proteoma/genética , Proteômica , Reprodutibilidade dos Testes , Análise de Sequência de RNA , TranscriptomaRESUMO
To understand the molecular mechanisms of parasitism in vivo, it is essential to elucidate how the transcriptomes of the human hosts and the infecting parasites affect one another. Here we report the RNA-seq analysis of 116 Indonesian patients infected with the malaria parasite Plasmodium falciparum (Pf). We extracted RNAs from their peripheral blood as a mixture of host and parasite transcripts and mapped the RNA-seq tags to the human and Pf reference genomes to separate the respective tags. We were thus able to simultaneously analyze expression patterns in both humans and parasites. We identified human and parasite genes and pathways that correlated with various clinical data, which may serve as primary targets for drug developments. Of particular importance, we revealed characteristic expression changes in the human innate immune response pathway genes including TLR2 and TICAM2 that correlated with the severity of the malaria infection. We also found a group of transcription regulatory factors, JUND, for example, and signaling molecules, TNFAIP3, for example, that were strongly correlated in the expression patterns of humans and parasites. We also identified several genetic variations in important anti-malaria drug resistance-related genes. Furthermore, we identified the genetic variations which are potentially associated with severe malaria symptoms both in humans and parasites. The newly generated data should collectively lay a unique foundation for understanding variable behaviors of the field malaria parasites, which are far more complex than those observed under laboratory conditions.
Assuntos
Genoma Humano , Genoma de Protozoário , Malária/genética , Plasmodium falciparum/genética , Transcriptoma , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adolescente , Adulto , Antimaláricos/uso terapêutico , Estudos de Casos e Controles , Criança , Pré-Escolar , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Resistência a Medicamentos/genética , Etiquetas de Sequências Expressas , Feminino , Interações Hospedeiro-Parasita/genética , Humanos , Imunidade Inata/genética , Lactente , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Malária/diagnóstico , Malária/tratamento farmacológico , Masculino , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Plasmodium falciparum/patogenicidade , Polimorfismo de Nucleotídeo Único , Proteínas Proto-Oncogênicas c-jun/genética , Proteínas Proto-Oncogênicas c-jun/metabolismo , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo , Proteína 3 Induzida por Fator de Necrose Tumoral alfa , Virulência/genéticaRESUMO
In mammals, the cell surface receptors encoded by the leukocyte receptor complex (LRC) regulate the activity of T lymphocytes and B lymphocytes, as well as that of natural killer cells, and thus provide protection against pathogens and parasites. The chicken genome encodes many Ig-like receptors that are homologous to the LRC receptors. The chicken Ig-like receptor (CHIR) genes are members of a large monophyletic gene family and are organized into genomic clusters, which are in conserved synteny with the mammalian LRC. One-third of CHIR genes encode polypeptide molecules that contain both activating and inhibitory motifs. These genes are present in different phylogenetic groups, suggesting that the primordial CHIR gene could have encoded both types of motifs in a single molecule. In contrast to the mammalian LRC genes, the CHIR genes with similar function (inhibition or activation) are evolutionarily closely related. We propose that, in addition to recombination, single nucleotide substitutions played an important role in the generation of receptors with different functions. Structural models and amino acid analyses of the CHIR proteins reveal the presence of different types of Ig-like domains in the same phylogenetic groups, as well as sharing of conserved residues and conserved changes of residues between different CHIR groups and between CHIRs and LRCs. Our data support the notion that the CHIR gene clusters are regions homologous to the mammalian LRC gene cluster and favor a model of evolution by repeated processes of birth and death (expansion-contraction) of the Ig-like receptor genes.
Assuntos
Galinhas/genética , Evolução Molecular , Leucócitos/metabolismo , Receptores de Superfície Celular/genética , Animais , Estrutura Terciária de Proteína , Receptores de Superfície Celular/metabolismo , Análise de Sequência de DNARESUMO
Factors affecting splicing of plant U12-dependent introns have been examined by extensive mutational analyses in an in vivo tobacco (Nicotiana tabacum) protoplast system using introns from three different Arabidopsis thaliana genes: CBP20, GSH2, and LD. The results provide evidence that splicing efficiency of plant U12 introns depends on a combination of factors, including UA content, exon bridging interactions between the U12 intron and flanking U2-dependent introns, and exon splicing enhancer sequences (ESEs). Unexpectedly, all three plant U12 introns required an adenosine at the upstream purine position in the branchpoint consensus UCCUURAUY. The exon upstream of the LD U12 intron is a major determinant of its higher level of splicing efficiency and potentially contains two ESE regions. These results suggest that in plants, U12 introns represent a level at which expression of their host genes can be regulated.