RESUMO
Viral genomes frequently harbor overlapping genes, complicating the development of virus-vectored vaccines and gene therapies. This study introduces a novel conditional splicing system to precisely control the expression of such overlapping genes through recombinase-mediated conditional splicing. We refined site-specific recombinase (SSR) conditional splicing systems and explored their mechanisms. The systems demonstrated exceptional inducibility (116,700-fold increase) with negligible background expression, facilitating the conditional expression of overlapping genes in adenovirus-associated virus (AAV) and human immunodeficiency virus type 1. Notably, this approach enabled the establishment of stable AAV producer cell lines, encapsulating all necessary packaging genes. Our findings underscore the potential of the SSR-conditional splicing system to significantly advance vector engineering, enhancing the efficacy and scalability of viral-vector-based therapies and vaccines. IMPORTANCE: Regulating overlapping genes is vital for gene therapy and vaccine development using viral vectors. The regulation of overlapping genes presents challenges, including cytotoxicity and impacts on vector capacity and genome stability, which restrict stable packaging cell line development and broad application. To address these challenges, we present a "loxp-splice-loxp"-based conditional splicing system, offering a novel solution for conditional expression of overlapping genes and stable cell line establishment. This system may also regulate other cytotoxic genes, representing a significant advancement in cell engineering and gene therapy as well as biomass production.
Assuntos
Dependovirus , Homologia de Genes , Genes Virais , Engenharia Genética , HIV-1 , Splicing de RNA , Humanos , Linhagem Celular , Dependovirus/genética , DNA Nucleotidiltransferases/genética , DNA Nucleotidiltransferases/metabolismo , Regulação Viral da Expressão Gênica , Homologia de Genes/genética , Genes Virais/genética , Engenharia Genética/métodos , Terapia Genética/métodos , Vetores Genéticos/genética , HIV-1/genética , Splicing de RNA/genética , Vacinas/biossíntese , Vacinas/genética , Empacotamento do Genoma Viral/genéticaRESUMO
Osteosarcoma (OS) is the most common primary malignant tumor of bone. Remodelin, an inhibitor of the N (4)-Acetylcytidine (ac4C) acetylation modifying enzyme N-acetyltransferase 10 (NAT10), has been shown to have therapeutic effects on cancer in several studies, and our previous studies have confirmed the inhibitory effect of Remodelin on OS cells, however, the mechanism of action has not yet been elucidated. We used network pharmacological analysis to quantify the therapeutic targets of Remodelin against OS. acRIP-seq and RNA-seq were performed to investigate the inhibitory activity of Remodelin on acetylation and its effect on the transcriptome after intervening in OS cells U2OS with Remodelin in vitro. Key target genes were deduced based on their pharmacological properties, combined with network pharmacology results and sequencing results. Finally, the deduced target genes were validated with vitro experiments. Network pharmacological analysis showed that 2291 OS-related target genes and 369 Remodelin-related target genes were obtained, and 116 overlapping genes were identified as Remodelin targets for OS treatment. Sequencing results showed that a total of 13,736 statistically significant ac4C modification peaks were detected by acRIP-seq, including 6938 hypoacetylation modifications and 6798 hyperacetylation modifications. A total of 2350 statistically significant mRNAs were detected by RNA-seq, of which 830 were up-regulated and 1520 were down-regulated. Association analyses identified a total of 382 genes that were Hypoacetylated-down, consistent with inhibition of mRNA acetylation and expression by Remodelin. Five genes, CASP3, ESR2, FGFR2, IGF1 and MAPK1, were identified as key therapeutic targets of Remodelin against OS. Finally, in vitro experiments, CCK-8 and qRT-PCR demonstrated that Remodelin indeed inhibited the proliferation of OS cells and reduced the expression of three genes: ESR2, IGF1, and MAPK1. In conclusion, ESR2, IGF1 and MAPK1 were identified as key therapeutic targets of Remodelin against OS. This reveals the target of Remodelin's pharmacological action on OS and provides new ideas for the treatment of OS.
Assuntos
Neoplasias Ósseas , Hidrazonas , Osteossarcoma , Tiazóis , Humanos , Farmacologia em Rede , Osteossarcoma/tratamento farmacológico , Osteossarcoma/genética , Homologia de Genes , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/genéticaRESUMO
OBJECTIVE: As one of the most common endocrine disorders in women of reproductive age, polycystic ovary syndrome (PCOS) is highly heterogeneous with varied clinical features and diverse gestational complications among individuals. The patients with PCOS have 2-fold higher risk of preterm labor which is associated with substantial infant morbidity and mortality and great socioeconomic cost. The study was designated to identify molecular subtypes and the related hub genes to facilitate the susceptibility assessment of preterm labor in women with PCOS. METHODS: Four mRNA datasets (GSE84958, GSE5090, GSE43264 and GSE98421) were obtained from Gene Expression Omnibus database. Twenty-eight candidate genes related to preterm labor or labor were yielded from the researches and our unpublished data. Then, we utilized unsupervised clustering to identify molecular subtypes in PCOS based on the expression of above candidate genes. Key modules were generated with weighted gene co-expression network analysis R package, and their hub genes were generated with CytoHubba. The probable biological function and mechanism were explored through Gene Ontology analysis and Kyoto Encyclopedia of Genes and Genomes pathway analysis. In addition, STRING and Cytoscape software were used to identify the protein-protein interaction (PPI) network, and the molecular complex detection (MCODE) was used to identify the hub genes. Then the overlapping hub genes were predicted. RESULTS: Two molecular subtypes were found in women with PCOS based on the expression similarity of preterm labor or labor-related genes, in which two modules were highlighted. The key modules and PPI network have five overlapping five hub genes, two of which, GTF2F2 and MYO6 gene, were further confirmed by the comparison between clustering subgroups according to the expression of hub genes. CONCLUSIONS: Distinct PCOS molecular subtypes were identified with preterm labor or labor-related genes, which might uncover the potential mechanism underlying heterogeneity of clinical pregnancy complications in women with PCOS.
Assuntos
Trabalho de Parto Prematuro , Síndrome do Ovário Policístico , Feminino , Humanos , Gravidez , Análise por Conglomerados , Homologia de Genes , Funções Verossimilhança , Síndrome do Ovário Policístico/genéticaRESUMO
PARP inhibitors (PARPi) have revolutionized the therapeutic landscape of epithelial ovarian cancer (EOC) treatment with outstanding benefits in regard to progression-free survival, especially in patients either carrying BRCA1/2 mutations or harboring defects in the homologous recombination repair system. Yet, it remains uncertain which PARPi to apply and how to predict responders when platinum sensitivity is unknown. To shed light on the predictive power of genes previously suggested to be associated with PARPi response, we systematically reviewed the literature and identified 79 publications investigating a total of 93 genes. The top candidate genes were further tested using a comprehensive CRISPR-Cas9 mutagenesis screening in combination with olaparib treatment. Therefore, we generated six constitutive Cas9+ EOC cell lines and profiled 33 genes in a CRISPR-Cas9 cell competition assay using non-essential (AAVS1) and essential (RPA3 and PCNA) genes for cell fitness as negative and positive controls, respectively. We identified only ATM, MUS81, NBN, BRCA2, and RAD51B as predictive markers for olaparib response. As the major survival benefit of PARPi treatment was reported in platinum-sensitive tumors, we next assessed nine top candidate genes in combination with three PARPi and carboplatin. Interestingly, we observed similar dropout rates in a gene and compound independent manner, supporting the strong correlation of cancer cell response to compounds that rely on DNA repair for their effectiveness. In addition, we report on CDK12 as a common vulnerability for EOC cell survival and proliferation without altering the olaparib response, highlighting its potential as a therapeutic target in EOC.
Assuntos
Neoplasias Ovarianas , Inibidores de Poli(ADP-Ribose) Polimerases , Humanos , Feminino , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Carcinoma Epitelial do Ovário/tratamento farmacológico , Carcinoma Epitelial do Ovário/genética , Carboplatina/farmacologia , Carboplatina/uso terapêutico , Sistemas CRISPR-Cas/genética , Detecção Precoce de Câncer , Ftalazinas/farmacologia , Ftalazinas/uso terapêutico , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Homologia de GenesRESUMO
Background: The tumor microenvironment (TME) primarily comprises cancer cells, cancer-infiltrating immune cells, and stromal cells. The tumor cells alter the TME by secreting signaling molecules to induce immune tolerance. The immune cell infiltrating the TME influences the prognosis of patients with cancers. However, immune cell infiltration (ICI) in the TME of patients with prostate cancer (PC) has not yet been studied. Methods: In this study, we used Cell-type Identification by Estimating Relative Subsets of RNA Transcripts (CIBERSORT) and Estimation of Stromal and Immune cells in Malignant Tumors using Expression data (ESTIMATE) algorithms to identify three ICI clusters based on 1,099 genes associated with ICI in the TME. The patients were classified into three distinct ICI gene clusters based on overlapping differentially expressed genes in ICI clusters. Furthermore, the ICI scores were calculated using principal component analysis. Results: The results revealed that patients with high ICI scores had poor prognoses and reduced expression of immune-checkpoint genes and immune-related genes. Furthermore, the transforming growth factor-beta (TGF-ß) and WNT-ß signaling pathways were enriched in the high ICI score subgroup, which suggests that suppression of T cells could contribute to poor prognosis of patients with PC. A positive correlation was observed between the high-ICI-score subgroup and the high tumor mutation burden (TMB) value. Patients with low ICI scores could benefit from immunotherapy, indicating that the ICI score could be used to predict the efficacy of immunotherapeutic response. Conclusions: In summary, we provide a comprehensive overview of the landscape of ICI in PC, which could aid in designing the strategies for immunotherapy for patients with PC.
Assuntos
Neoplasias da Próstata , Humanos , Masculino , Algoritmos , Transporte Biológico , Homologia de Genes , Tolerância Imunológica , Neoplasias da Próstata/genética , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/terapia , Microambiente Tumoral/genéticaRESUMO
So-called oncocytic papillary renal cell carcinoma (OPRCC) is a poorly defined variant of papillary renal cell carcinoma. Since its first description, several studies were published with conflicting results, and thus precise definition is lacking. A cohort of 39 PRCCs composed of oncocytic cells were analyzed. Cases were divided into 3 groups based on copy number variation (CNV) pattern. The first group consisted of 23 cases with CNV equal to renal oncocytoma. The second group consisted of 7 cases with polysomy of chromosomes 7 and 17 and the last group of 9 cases included those with variable CNV. Epidemiologic, morphologic and immunohistochemical features varied among the groups. There were not any particular histomorphologic features correlating with any of the genetic subgroups. Further, a combination of morphologic, immunohistochemical, and molecular-genetic features did not allow to precisely predict biologic behavior. Owing to variable CNV pattern in OPRCC, strict adherence to morphology and immunohistochemical profile is recommended, particularly in limited samples (i.e., core biopsy). Applying CNV pattern as a part of a diagnostic algorithm can be potentially misleading. OPRCC is a highly variable group of tumors, which might be misdiagnosed as renal oncocytoma. Using the term OPRCC as a distinct diagnostic entity is, thanks to its high heterogeneity, questionable.
Assuntos
Adenoma Oxífilo/genética , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Neoplasias Renais/genética , Células Oxífilas/metabolismo , Adenoma Oxífilo/diagnóstico , Adenoma Oxífilo/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/metabolismo , Biópsia com Agulha de Grande Calibre/normas , Carcinoma de Células Renais/epidemiologia , Aberrações Cromossômicas , Variações do Número de Cópias de DNA/genética , Diagnóstico Diferencial , Erros de Diagnóstico , Feminino , Homologia de Genes/genética , Humanos , Imuno-Histoquímica/métodos , Hibridização in Situ Fluorescente/métodos , Neoplasias Renais/diagnóstico , Neoplasias Renais/patologia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias/métodos , Células Oxífilas/patologiaRESUMO
Heterobifunctional proteolysis-targeting chimeric compounds leverage the activity of E3 ligases to induce degradation of target oncoproteins and exhibit potent preclinical antitumor activity. To dissect the mechanisms regulating tumor cell sensitivity to different classes of pharmacological "degraders" of oncoproteins, we performed genome-scale CRISPR-Cas9-based gene editing studies. We observed that myeloma cell resistance to degraders of different targets (BET bromodomain proteins, CDK9) and operating through CRBN (degronimids) or VHL is primarily mediated by prevention of, rather than adaptation to, breakdown of the target oncoprotein; and this involves loss of function of the cognate E3 ligase or interactors/regulators of the respective cullin-RING ligase (CRL) complex. The substantial gene-level differences for resistance mechanisms to CRBN- versus VHL-based degraders explains mechanistically the lack of cross-resistance with sequential administration of these two degrader classes. Development of degraders leveraging more diverse E3 ligases/CRLs may facilitate sequential/alternating versus combined uses of these agents toward potentially delaying or preventing resistance.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Antineoplásicos/farmacologia , Mieloma Múltiplo/genética , Mieloma Múltiplo/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Animais , Sistemas CRISPR-Cas , Linhagem Celular Tumoral , Quinase 9 Dependente de Ciclina/metabolismo , Resistencia a Medicamentos Antineoplásicos , Edição de Genes , Regulação Neoplásica da Expressão Gênica , Homologia de Genes , Estudo de Associação Genômica Ampla , Genômica/métodos , Humanos , Camundongos , Mieloma Múltiplo/tratamento farmacológico , Proteínas Oncogênicas/metabolismo , Proteínas/antagonistas & inibidores , Proteínas/metabolismo , Proteólise , Células Tumorais CultivadasRESUMO
Overlapping genes originate by a mechanism of overprinting, in which nucleotide substitutions in a pre-existing frame induce the expression of a de novo protein from an alternative frame. In this study, I assembled a dataset of 319 viral overlapping genes, which included 82 overlaps whose expression is experimentally known and the respective 237 homologs. Principal component analysis revealed that overlapping genes have a common pattern of nucleotide and amino acid composition. Discriminant analysis separated overlapping from non-overlapping genes with an accuracy of 97%. When applied to overlapping genes with known genealogy, it separated ancestral from de novo frames with an accuracy close to 100%. This high discriminant power was crucial to computationally design variants of de novo viral proteins known to possess selective anticancer toxicity (apoptin) or protection against neurodegeneration (X protein), as well as to detect two new potential overlapping genes in the genome of the new coronavirus SARS-CoV-2.
Assuntos
Betacoronavirus/genética , Evolução Molecular , Homologia de Genes , Genes Virais , Algoritmos , Sequência de Aminoácidos , Sequência de Bases , Biologia Computacional , Simulação por Computador , Análise Discriminante , Análise dos Mínimos Quadrados , Análise de Componente Principal , SARS-CoV-2RESUMO
AIMS: Evaluation of nucleic acids in plasma exosomes is a noninvasive method that can be used to detect different types of cancer. The aim of this study was to determine the value of exosomal long noncoding RNAs (lncRNAs) in detecting lung squamous cell carcinoma (LSCC). MATERIALS AND METHODS: A total of 75 LSCC patients and 79 negative control subjects were enrolled in the study. Twenty differentially expressed lncRNAs were evaluated as potential candidates. Exosomes were isolated by ultracentrifugation, and lncRNA levels in exosomes were determined using real-time polymerase chain reaction. Receiver Operating Characteristic (ROC) curves were used to determine specificity and sensitivity. RESULTS: Exosomal SOX2-OT was significantly upregulated in LSCC patients and showed the strongest power in detecting LSCC. The area under the ROC curve was 0.815, and the sensitivity and specificity were 76% and 73.17%, respectively. Moreover, exosomal SOX2-OT levels were significantly correlated with tumor size, TNM stage, and lymph node metastasis. Exosomal SOX2-OT levels were significantly decreased in the postoperative plasma of LSCC patients. CONCLUSION: SOX2-OT may serve as a promising noninvasive plasma-based tumor biomarker for LSCC.
Assuntos
Carcinoma de Células Escamosas/genética , Complexo Multienzimático de Ribonucleases do Exossomo/genética , Fatores de Transcrição SOXB1/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Complexo Multienzimático de Ribonucleases do Exossomo/sangue , Exossomos/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Homologia de Genes/genética , Humanos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Metástase Linfática , Masculino , Pessoa de Meia-Idade , RNA Longo não Codificante/análise , RNA Longo não Codificante/genética , Curva ROC , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Airflow obstruction is a hallmark of chronic obstructive pulmonary disease (COPD), and is defined as either the ratio between forced expiratory volume in one second and forced vital capacity (FEV1/FVC) < 70% or < lower limit of normal (LLN). This study aimed to assess the overlap between genome-wide association studies (GWAS) on airflow obstruction using these two definitions in the same population stratified by smoking. METHODS: GWASes were performed in the LifeLines Cohort Study for both airflow obstruction definitions in never-smokers (NS = 5071) and ever-smokers (ES = 4855). The FEV1/FVC < 70% models were adjusted for sex, age, and height; FEV1/FVC < LLN models were not adjusted. Ever-smokers models were additionally adjusted for pack-years and current-smoking. The overlap in significantly associated SNPs between the two definitions and never/ever-smokers was assessed using several p-value thresholds. To quantify the agreement, the Pearson correlation coefficient was calculated between the p-values and ORs. Replication was performed in the Vlagtwedde-Vlaardingen study (NS = 432, ES = 823). The overlapping SNPs with p < 10- 4 were validated in the Vlagtwedde-Vlaardingen and Rotterdam Study cohorts (NS = 1966, ES = 3134) and analysed for expression quantitative trait loci (eQTL) in lung tissue (n = 1087). RESULTS: In the LifeLines cohort, 96% and 93% of the never- and ever-smokers were classified concordantly based on the two definitions. 26 and 29% of the investigated SNPs were overlapping at p < 0.05 in never- and ever-smokers, respectively. At p < 10- 4 the overlap was 4% and 6% respectively, which could be change findings as shown by simulation studies. The effect estimates of the SNPs of the two definitions correlated strongly, but the p-values showed more variation and correlated only moderately. Similar observations were made in the Vlagtwedde-Vlaardingen study. Two overlapping SNPs in never-smokers (NFYC and FABP7) had the same direction of effect in the validation cohorts and the NFYC SNP was an eQTL for NFYC-AS1. NFYC is a transcription factor that binds to several known COPD genes, and FABP7 may be involved in abnormal pulmonary development. CONCLUSIONS: The definition of airflow obstruction and the population under study may be important determinants of which SNPs are associated with airflow obstruction. The genes FABP7 and NFYC(-AS1) could play a role in airflow obstruction in never-smokers specifically.
Assuntos
Fator de Ligação a CCAAT/genética , Proteína 7 de Ligação a Ácidos Graxos/genética , Estudo de Associação Genômica Ampla , Doença Pulmonar Obstrutiva Crônica/genética , Fumar/genética , Proteínas Supressoras de Tumor/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Feminino , Volume Expiratório Forçado , Homologia de Genes/genética , Predisposição Genética para Doença , Humanos , Modelos Lineares , Modelos Logísticos , Pulmão/fisiopatologia , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas/genética , Fumar/efeitos adversos , Espirometria , Capacidade Vital , Adulto JovemRESUMO
Mechanisms by which members of the AP-1 family of transcription factors play non-redundant biological roles despite recognizing the same DNA sequence remain poorly understood. To address this question, here we investigate the molecular functions and genome-wide DNA binding patterns of AP-1 family members in primary and immortalized mouse macrophages. ChIP-sequencing shows overlapping and distinct binding profiles for each factor that were remodeled following TLR4 ligation. Development of a machine learning approach that jointly weighs hundreds of DNA recognition elements yields dozens of motifs predicted to drive factor-specific binding profiles. Machine learning-based predictions are confirmed by analysis of the effects of mutations in genetically diverse mice and by loss of function experiments. These findings provide evidence that non-redundant genomic locations of different AP-1 family members in macrophages largely result from collaborative interactions with diverse, locus-specific ensembles of transcription factors and suggest a general mechanism for encoding functional specificities of their common recognition motif.
Assuntos
DNA/metabolismo , Genoma , Macrófagos/metabolismo , Fator de Transcrição AP-1/genética , Fator de Transcrição AP-1/metabolismo , Fator 3 Ativador da Transcrição , Animais , Sequência de Bases , Sítios de Ligação/genética , Linhagem Celular , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/genética , Técnicas de Inativação de Genes , Homologia de Genes , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Mutação , Motivos de Nucleotídeos , Domínios Proteicos , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Receptor 4 Toll-Like/metabolismoRESUMO
Genetic coding in bacteria largely operates via the "one gene-one protein" paradigm. However, the peculiarities of the mRNA structure, the versatility of the genetic code, and the dynamic nature of translation sometimes allow organisms to deviate from the standard rules of protein encoding. Bacteria can use several unorthodox modes of translation to express more than one protein from a single mRNA cistron. One such alternative path is the use of additional translation initiation sites within the gene. Proteins whose translation is initiated at different start sites within the same reading frame will differ in their N termini but will have identical C-terminal segments. On the other hand, alternative initiation of translation in a register different from the frame dictated by the primary start codon will yield a protein whose sequence is entirely different from the one encoded in the main frame. The use of internal mRNA codons as translation start sites is controlled by the nucleotide sequence and the mRNA folding. The proteins of the alternative proteome generated via the "genes-within-genes" strategy may carry important functions. In this review, we summarize the currently known examples of bacterial genes encoding more than one protein due to the utilization of additional translation start sites and discuss the known or proposed functions of the alternative polypeptides in relation to the main protein product of the gene. We also discuss recent proteome- and genome-wide approaches that will allow the discovery of novel translation initiation sites in a systematic fashion.
Assuntos
Bactérias/genética , Códon de Iniciação , Genoma Bacteriano/genética , Proteínas de Bactérias/genética , Regulação Bacteriana da Expressão Gênica , Homologia de Genes , Código Genético , Fases de Leitura Aberta , Iniciação Traducional da Cadeia PeptídicaRESUMO
Gene overlap plays various regulatory functions on transcriptional and post-transcriptional levels. Most current studies focus on protein-coding genes overlapping with non-protein-coding counterparts, the so called natural antisense transcripts. Considerably less is known about the role of gene overlap in the case of two protein-coding genes. Here, we provide OverGeneDB, a database of human and mouse 5' end protein-coding overlapping genes. The database contains 582 human and 113 mouse gene pairs that are transcribed using overlapping promoters in at least one analyzed library. Gene pairs were identified based on the analysis of the transcription start site (TSS) coordinates in 73 human and 10 mouse organs, tissues and cell lines. Beside TSS data, resources for 26 human lung adenocarcinoma cell lines also contain RNA-Seq and ChIP-Seq data for seven histone modifications and RNA Polymerase II activity. The collected data revealed that the overlap region is rarely conserved between the studied species and tissues. In â¼50% of the overlapping genes, transcription started explicitly in the overlap regions. In the remaining half of overlapping genes, transcription was initiated both from overlapping and non-overlapping TSSs. OverGeneDB is accessible at http://overgenedb.amu.edu.pl.
Assuntos
Bases de Dados Genéticas , Homologia de Genes , Animais , Expressão Gênica , Código das Histonas , Humanos , Camundongos , Família Multigênica , Fases de Leitura Aberta , Regiões Promotoras Genéticas , RNA Polimerase II/metabolismo , Análise de Sequência de RNA , Fatores de Transcrição/metabolismo , Sítio de Iniciação de TranscriçãoRESUMO
Cancer database is a systematic collection and analysis of information on various human cancers at genomic and molecular level that can be utilized to understand various steps in carcinogenesis and for therapeutic advancement in cancer field. Oral cancer is one of the leading causes of morbidity and mortality all over the world. The current research efforts in this field are aimed at cancer etiology and therapy. Advanced genomic technologies including microarrays, proteomics, transcrpitomics, and gene sequencing development have culminated in generation of extensive data and subjection of several genes and microRNAs that are distinctively expressed and this information is stored in the form of various databases. Extensive data from various resources have brought the need for collaboration and data sharing to make effective use of this new knowledge. The current review provides comprehensive information of various publicly accessible databases that contain information pertinent to oral squamous cell carcinoma (OSCC) and databases designed exclusively for OSCC. The databases discussed in this paper are Protein-Coding Gene Databases and microRNA Databases. This paper also describes gene overlap in various databases, which will help researchers to reduce redundancy and focus on only those genes, which are common to more than one databases. We hope such introduction will promote awareness and facilitate the usage of these resources in the cancer research community, and researchers can explore the molecular mechanisms involved in the development of cancer, which can help in subsequent crafting of therapeutic strategies.
Assuntos
Pesquisa Biomédica , Bases de Dados Factuais , Internet , Neoplasias Bucais/genética , Carcinogênese/genética , Carcinoma de Células Escamosas/genética , Regulação Neoplásica da Expressão Gênica , Genes Neoplásicos/genética , Homologia de Genes , Genes Supressores de Tumor , Humanos , MicroRNAs/genética , Fases de Leitura Aberta/genética , PesquisadoresRESUMO
Overlapping genes exist in all domains of life and are much more abundant than expected upon their first discovery in the late 1970s. Assuming that the reference gene is read in frame +0, an overlapping gene can be encoded in two reading frames in the sense strand, denoted by +1 and +2, and in three reading frames in the opposite strand, denoted by -0, -1, and -2. This motivated numerous researchers to study the constraints induced by the genetic code on the various overlapping frames, mostly based on information theory. Our focus in this paper is on the constraints induced on two overlapping genes in terms of amino acids, as well as polypeptides. We show that simple linear constraints bind the amino-acid composition of two proteins encoded by overlapping genes. Novel constraints are revealed when polypeptides are considered, and not just single amino acids. For example, in double-coding sequences with an overlapping reading frame -2, each Tyrosine (denoted as Tyr or Y) in the overlapping frame overlaps a Tyrosine in the reference frame +0 (and reciprocally), whereas specific words (e.g. YY) never occur. We thus distinguish between null constraints (YY = 0 in frame -2) and non-null constraints (Y in frame +0 â Y in frame -2). Our equivalence-based constraints are symmetrical and thus enable the characterization of the joint composition of overlapping proteins. We describe several formal frameworks and a graph algorithm to characterize and compute these constraints. As expected, the degrees of freedom left by these constraints vary drastically among the different overlapping frames. Interestingly, the biological meaning of constraints induced on two overlapping proteins (hydropathy, forbidden di-peptides, expected overlap length ) is also specific to the reading frame. We study the combinatorics of these constraints for overlapping polypeptides of length n, pointing out that, (i) except for frame -2, non-null constraints are deduced from the amino-acid (length = 1) constraints and (ii) null constraints are deduced from the di-peptide (length = 2) constraints. These results yield support for understanding the mechanisms and evolution of overlapping genes, and for developing novel overlapping gene detection methods.
Assuntos
Sequência de Aminoácidos/genética , Homologia de Genes , Fases de Leitura Aberta , Proteínas/genética , Algoritmos , Animais , Evolução Biológica , HumanosRESUMO
BACKGROUND: Iron (Fe) and phosphorus (P) are essential mineral nutrients in plants. Knowledge regarding global changes in the abundance of Fe-responsive genes under Pi deficiency as well as the processes these genes are involved in remains largely unavailable at the genome level. In the current study, we comparatively analyzed RNA sequencing data sets relative to Fe deficiency (NCBI: SRP044814) and Pi starvation (NCBI: SRA050356.1). RESULTS: Analysis showed a total of 579 overlapping genes that are responsible for both Fe deficiency and Pi starvation in Arabidopsis roots. A subset of 137 genes had greater than twofold changes in transcript abundant as a result of the treatments. Gene ontology (GO) analysis showed that the stress-related processes 'response to salt stress', 'response to oxidative stress', and 'response to zinc ion' were enriched in the 579 genes, while Fe response-related processes, including 'cellular response to nitric oxide', 'cellular response to iron ion', and 'cellular iron ion homeostasis', were also enriched in the subset of 137 genes. Co-expression analysis of the 579 genes using the MACCU toolbox yielded a network consisting of 292 nodes (genes). Further analysis revealed that a subset of 90 genes were up-regulated under Fe shortage, but down-regulated under Pi starvation. GO analysis in this group of genes revealed an increased cellular response to iron ion/nitric oxide/ethylene stimuli. Promoter analysis was performed in 35 of the 90 genes with a 1.5-fold or greater change in abundance, showing that 12 genes contained the PHOSPHATE STARVATION RESPONSE1-binding GNATATNC cis-element within their promoter regions. Quantitative real-time PCR showed that the decreased abundance of Fe acquisition genes under Pi deficiency exclusively relied on Fe concentration in Pi-deficient media. CONCLUSIONS: Comprehensive analysis of the overlapping genes derived from Fe deficiency and Pi starvation provides more information to understand the link between Pi and Fe homeostasis. Gene clustering and root-specific co-expression analysis revealed several potentially important genes which likely function as putative novel players in response to Fe and Pi deficiency or in cross-talk between Fe-deficient responses and Pi-deficient signaling.
Assuntos
Arabidopsis/genética , Epistasia Genética , Regulação da Expressão Gênica de Plantas , Homologia de Genes , Estudo de Associação Genômica Ampla , Deficiências de Ferro , Fosfatos/deficiência , Raízes de Plantas/genética , Sequência de Bases , Análise por Conglomerados , Regulação para Baixo/genética , Perfilação da Expressão Gênica , Ontologia Genética , Genes de Plantas , Dados de Sequência Molecular , Motivos de Nucleotídeos/genética , Raízes de Plantas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo RealRESUMO
DNA polymerase ν (POLN) is one of 16 DNA polymerases encoded in vertebrate genomes. It is important to determine its gene expression patterns, biological roles, and biochemical activities. By quantitative analysis of mRNA expression, we found that POLN from the zebrafish Danio rerio is expressed predominantly in testis. POLN is not detectably expressed in zebrafish embryos or in mouse embryonic stem cells. Consistent with this, injection of POLN-specific morpholino antisense oligonucleotides did not interfere with zebrafish embryonic development. Analysis of transcripts revealed that vertebrate POLN has an unusual gene expression arrangement, sharing a first exon with HAUS3, the gene encoding augmin-like complex subunit 3. HAUS3 is broadly expressed in embryonic and adult tissues, in contrast to POLN. Differential expression of POLN and HAUS3 appears to arise by alternate splicing of transcripts in mammalian cells and zebrafish. When POLN was ectopically overexpressed in human cells, it specifically coimmunoprecipitated with the homologous recombination factors BRCA1 and FANCJ, but not with previously suggested interaction partners (HELQ and members of the Fanconi anemia core complex). Purified zebrafish POLN protein is capable of thymine glycol bypass and strand displacement, with activity dependent on a basic amino acid residue known to stabilize the primer-template. These properties are conserved with the human enzyme. Although the physiological function of pol ν remains to be clarified, this study uncovers distinctive aspects of its expression control and evolutionarily conserved properties of this DNA polymerase.
Assuntos
DNA Polimerase Dirigida por DNA/metabolismo , Regulação da Expressão Gênica , Testículo/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Processamento Alternativo , Animais , Proteína BRCA1/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , DNA/química , Dano ao DNA , Éxons , Proteínas de Grupos de Complementação da Anemia de Fanconi/metabolismo , Ordem dos Genes , Homologia de Genes , Células HEK293 , Células HeLa , Humanos , Masculino , Camundongos , Recombinação Genética , Transgenes , Peixe-ZebraRESUMO
The growing number of available microbial genomes offers the possibility to identify features that could be used for identification. In this work, the possibility to exploit overlapping genes to develop a simple PCR based method of identification, was explored. Using the Burkholderia cepacia complex as a model, genomic analyses were performed to check the phylogenetic distribution of an overlap between marC and hisH genes and then, a PCR specific for Burkholderia was designed, set up and tested on a panel of strains and on DNA extracted from the sputum of cystic fibrosis patients. Results obtained revealed the usefulness of this approach, which could then be used to develop PCR for the identification of specific bacteria species or genera.
Assuntos
DNA Bacteriano/genética , Homologia de Genes/genética , Marcadores Genéticos/genética , Genoma Bacteriano/genética , Tipagem Molecular/métodos , Infecções por Burkholderia/diagnóstico , Infecções por Burkholderia/microbiologia , Complexo Burkholderia cepacia/genética , Complexo Burkholderia cepacia/isolamento & purificação , Fibrose Cística/microbiologia , Humanos , Reação em Cadeia da PolimeraseRESUMO
A inferência de homologia entre organismos é uma atividade da genômicacomparativa que possibilita compreender melhor a relação entre os mesmos e, porconseguinte, sua distância evolutiva. Especificamente, a identificação de genesortólogos, ou seja, aqueles que têm sua origem em um ancestral comum, permiteoferecer melhorias na anotação funcional de genes, uma vez que genes ortólogostendem a ter sua função conservada.Com a crescente disponibilidade de genomas através de técnicas de NGS, aconstrução e atualização de bases de dados de ortólogos representam um desafioconstante, pois demandam o estudo e identificação das relações entre os genes detais organismos, em um volume de dados cada vez mais extenso e a um custocomputacional cada vez mais elevado.Nesta tese propomos a solução para nuvem computacional elastic-OrthoSearch, umworkflow científico de genômica comparativa inspirado no OrthoSearch, responsávelpela inferência de homologia entre organismos com o uso de abordagem baseadaem melhores hits recíprocos e perfis de Markov.Também propomos uma metodologia para criação de bases de ortólogos construídaatravés do reuso do OrthoSearch. Esta metodologia mostrou-se capaz de alavancara oferta de grupos ortólogos e assim auxiliar, por exemplo, na identificação de alvosde protozoários...
Homology inference among organisms is a comparative genomics tasks which allowsfor a better understanding on how such organisms are related to each other and ontheir evolutionary distance. Specifically, the identification of orthologous genes those who share a common ancestor allows for functional gene annotationimprovements, as orthologous genes tend to preserve their functions.The increasing amount of genomic data provided by the NGS techniques makes theorthologous databases building and update processes a challenging task. It requiresthe identification and study of the organisms genes relationships, in an extensivedata volume and at an increasing computational cost.In this thesis we propose elastic-OrthoSearch, a cloud-enabled comparativegenomics scientific workflow, derived from OrthoSearch. It aims at providinghomology inference among organisms, in a reciprocal best hits and Markov profilesapproach.We also propose an improved orthologous database creation methodology built ontop of OrthoSearch. Such methodology has shown means to offer a broaderorthologous groups dataset, which could in turn aid on Protozoa target identification...
Assuntos
Animais , Homologia de Genes , Genoma de Protozoário , Genômica , Doenças Negligenciadas , Fluxo de Trabalho , Biologia ComputacionalRESUMO
A diversificação da produção industrial de alimentos de origem suína e o intercâmbio comercial de animais e seus derivados destinados ao consumo humano podem ser importantes disseminadores de sorovares de Salmonella spp. na cadeia alimentar. Objetivou-se avaliar em 86 cepas de Salmonella spp., isoladas em granja de terminação e no abate de suínos, a ocorrência de três genes de virulência (invA, agfA e lpfA), bem como a similaridade genética entre elas. A ocorrência do gene invA foi verificada em 100% das amostras. O gene lpfA foi detectado em 80,23% (69/86) das cepas, não foi detectado em S. Panama e estava presente em todas as cepas de S. Infantis. O gene agfA foi detectado em 63,95% (55/86) das amostras. S. Agona apresentou positividade para todos os genes de virulência estudados. A análise de homologia entre as cepas agrupou os diferentes sorovares em clusters. A similaridade foi independente do local de isolamento, o que demonstra a presença de clones ao longo da cadeia de produção e a existência de multiplicidade de fontes para a infecção dos animais, como a ração, e a contaminação cruzada das carcaças. A pesquisa de genes de virulência e a avaliação da proximidade gênica permitem a caracterização e um maior entendimento sobre cepas de Salmonella circulantes na cadeia produtiva de suínos e, assim, podem subsidiar medidas de controle durante o processo produtivo com o objetivo de garantir a saúde do consumidor...
The diversification of industrial food production of swine origin and trade of animals and their derivatives for human consumption may be important disseminators of serovars of Salmonella spp. in the food chain. This study aimed to evaluate 86 strains of Salmonella spp. isolated form in the finishing and slaughter of pigs, the occurrence of three virulence genes (invA, agfa and lpfA), as well as the genetic similarity between them. The occurrence of gene invA was observed in 100% of the samples. The gene lpfA was detected in 80.23% (69/86) strains and is not detected in S. Panama, but present in all strains of S. Infantis. The gene agfA was detected in 63.95% (55/86). S. Agona was positive for all virulence genes studied. The analysis of homology between the different serovars grouped the isolates in clusters. The similarity was regardless of the location of isolation, demonstrating the presence of clones along the production chain and that there are multiple sources for the infection of animals, such as feed, and cross-contamination of carcasses. A survey of virulence genes and evaluation of gene proximity allow characterization and better understanding of Salmonella strains circulating in the pig production chain, thus being able to support control measures during the production process in order to ensure consumer health...