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1.
Cell ; 173(2): 321-337.e10, 2018 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-29625050

RESUMO

Genetic alterations in signaling pathways that control cell-cycle progression, apoptosis, and cell growth are common hallmarks of cancer, but the extent, mechanisms, and co-occurrence of alterations in these pathways differ between individual tumors and tumor types. Using mutations, copy-number changes, mRNA expression, gene fusions and DNA methylation in 9,125 tumors profiled by The Cancer Genome Atlas (TCGA), we analyzed the mechanisms and patterns of somatic alterations in ten canonical pathways: cell cycle, Hippo, Myc, Notch, Nrf2, PI-3-Kinase/Akt, RTK-RAS, TGFß signaling, p53 and ß-catenin/Wnt. We charted the detailed landscape of pathway alterations in 33 cancer types, stratified into 64 subtypes, and identified patterns of co-occurrence and mutual exclusivity. Eighty-nine percent of tumors had at least one driver alteration in these pathways, and 57% percent of tumors had at least one alteration potentially targetable by currently available drugs. Thirty percent of tumors had multiple targetable alterations, indicating opportunities for combination therapy.


Assuntos
Bases de Dados Genéticas , Neoplasias/patologia , Transdução de Sinais/genética , Genes Neoplásicos , Humanos , Neoplasias/genética , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas Wnt/genética , Proteínas Wnt/metabolismo
2.
EMBO J ; 40(19): e107988, 2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34423452

RESUMO

The intricate process of human mtDNA replication requires the coordinated action of both transcription and replication machineries. Transcription and replication events at the lagging strand of mtDNA prompt the formation of a stem-loop structure (OriL) and the synthesis of a ∼25 nt RNA primer by mitochondrial RNA polymerase (mtRNAP). The mechanisms by which mtRNAP recognizes OriL, initiates transcription, and transfers the primer to the replisome are poorly understood. We found that transcription initiation at OriL involves slippage of the nascent transcript. The transcript slippage is essential for initiation complex stability and its ability to translocate the mitochondrial DNA polymerase gamma, PolG, which pre-binds to OriL, downstream of the replication origin thus allowing for the primer synthesis. Our data suggest the primosome assembly at OriL-a complex of mtRNAP and PolG-can efficiently generate the primer, transfer it to the replisome, and protect it from degradation by mitochondrial endonucleases.


Assuntos
Replicação do DNA , DNA Mitocondrial , Mitocôndrias/genética , Origem de Replicação , Iniciação da Transcrição Genética , Sequência de Bases , DNA Mitocondrial/química , DNA Mitocondrial/genética , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Mitocôndrias/metabolismo , Proteínas Mitocondriais/química , Proteínas Mitocondriais/metabolismo , Modelos Moleculares , Conformação Molecular , Conformação de Ácido Nucleico , RNA/química , RNA/genética , Relação Estrutura-Atividade
3.
Nucleic Acids Res ; 50(5): 2765-2781, 2022 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-35191499

RESUMO

Recognition of mammalian mitochondrial promoters requires the concerted action of mitochondrial RNA polymerase (mtRNAP) and transcription initiation factors TFAM and TFB2M. In this work, we found that transcript slippage results in heterogeneity of the human mitochondrial transcripts in vivo and in vitro. This allowed us to correctly interpret the RNAseq data, identify the bona fide transcription start sites (TSS), and assign mitochondrial promoters for > 50% of mammalian species and some other vertebrates. The divergent structure of the mammalian promoters reveals previously unappreciated aspects of mtDNA evolution. The correct assignment of TSS also enabled us to establish the precise register of the DNA in the initiation complex and permitted investigation of the sequence-specific protein-DNA interactions. We determined the molecular basis of promoter recognition by mtRNAP and TFB2M, which cooperatively recognize bases near TSS in a species-specific manner. Our findings reveal a role of mitochondrial transcription machinery in mitonuclear coevolution and speciation.


Assuntos
Mitocôndrias/genética , Transcrição Gênica , Animais , DNA Mitocondrial/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Humanos , Mamíferos/genética , Mamíferos/metabolismo , Mitocôndrias/enzimologia , Proteínas Mitocondriais/metabolismo , Fatores de Transcrição/química , Fatores de Transcrição/genética , Sítio de Iniciação de Transcrição
4.
J Biol Chem ; 293(15): 5585-5599, 2018 04 13.
Artigo em Inglês | MEDLINE | ID: mdl-29475949

RESUMO

Cytochrome b (Cytb) is the only mitochondrial encoded subunit from the bc1 complex. Cbp3 and Cbp6 are chaperones necessary for translation of the COB mRNA and Cytb hemylation. Here we demonstrate that their role in translation is dispensable in some laboratory strains, whereas their role in Cytb hemylation seems to be universally conserved. BY4742 yeast requires Cbp3 and Cbp6 for efficient COB mRNA translation, whereas the D273-10b strain synthesizes Cytb at wildtype levels in the absence of Cbp3 and Cbp6. Steady-state levels of Cytb are close to wildtype in mutant D273-10b cells, and Cytb forms non-functional, supercomplex-like species with cytochrome c oxidase, in which at least core 1, cytochrome c1, and Rieske iron-sulfur subunits are present. We demonstrated that Cbp3 interacts with the mitochondrial ribosome and with the COB mRNA in both BY4742 and D273-10b strains. The polymorphism(s) causing the differential function of Cbp3, Cbp6, and the assembly feedback regulation of Cytb synthesis is of nuclear origin rather than mitochondrial, and Smt1, a COB mRNA-binding protein, does not seem to be involved in the observed differential phenotype. Our results indicate that the essential role of Cbp3 and Cbp6 is to assist Cytb hemylation and demonstrate that in the absence of heme b, Cytb can form non-functional supercomplexes with cytochrome c oxidase. Our observations support that an additional protein or proteins are involved in Cytb synthesis in some yeast strains.


Assuntos
Citocromos b/biossíntese , Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Proteínas Mitocondriais/biossíntese , Chaperonas Moleculares/metabolismo , Biossíntese de Proteínas , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Citocromos b/genética , Citocromos c1/genética , Citocromos c1/metabolismo , Proteínas de Membrana/genética , Metiltransferases/genética , Metiltransferases/metabolismo , Mitocôndrias/genética , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Chaperonas Moleculares/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética
5.
Nucleic Acids Res ; 45(11): 6628-6643, 2017 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-28520979

RESUMO

Members of the DEAD-box family are often multifunctional proteins involved in several RNA transactions. Among them, yeast Saccharomyces cerevisiae Mss116 participates in mitochondrial intron splicing and, under cold stress, also in mitochondrial transcription elongation. Here, we show that Mss116 interacts with the mitoribosome assembly factor Mrh4, is required for efficient mitoribosome biogenesis, and consequently, maintenance of the overall mitochondrial protein synthesis rate. Additionally, Mss116 is required for efficient COX1 mRNA translation initiation and elongation. Mss116 interacts with a COX1 mRNA-specific translational activator, the pentatricopeptide repeat protein Pet309. In the absence of Mss116, Pet309 is virtually absent, and although mitoribosome loading onto COX1 mRNA can occur, activation of COX1 mRNA translation is impaired. Mutations abolishing the helicase activity of Mss116 do not prevent the interaction of Mss116 with Pet309 but also do not allow COX1 mRNA translation. We propose that Pet309 acts as an adaptor protein for Mss116 action on the COX1 mRNA 5΄-UTR to promote efficient Cox1 synthesis. Overall, we conclude that the different functions of Mss116 in the biogenesis and functioning of the mitochondrial translation machinery depend on Mss116 interplay with its protein cofactors.


Assuntos
RNA Helicases DEAD-box/fisiologia , Ribossomos Mitocondriais/metabolismo , Proteínas de Saccharomyces cerevisiae/fisiologia , Saccharomyces cerevisiae/genética , Regiões 5' não Traduzidas , Sequência de Bases , Sítios de Ligação , RNA Helicases DEAD-box/metabolismo , DNA Fúngico/genética , DNA Mitocondrial/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Regulação Fúngica da Expressão Gênica , Mitocôndrias/metabolismo , Proteínas Mitocondriais/biossíntese , Iniciação Traducional da Cadeia Peptídica , Estabilidade Proteica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo
6.
J Biol Chem ; 291(17): 9343-55, 2016 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-26929411

RESUMO

Cytochrome c oxidase assembly requires the synthesis of the mitochondria-encoded core subunits, Cox1, Cox2, and Cox3. In yeast, Pet54 protein is required to activate translation of the COX3 mRNA and to process the aI5ß intron on the COX1 transcript. Here we report a third, novel function of Pet54 on Cox1 synthesis. We observed that Pet54 is necessary to achieve an efficient Cox1 synthesis. Translation of the COX1 mRNA is coupled to the assembly of cytochrome c oxidase by a mechanism that involves Mss51. This protein activates translation of the COX1 mRNA by acting on the COX1 5'-UTR, and, in addition, it interacts with the newly synthesized Cox1 protein in high molecular weight complexes that include the factors Coa3 and Cox14. Deletion of Pet54 decreased Cox1 synthesis, and, in contrast to what is commonly observed for other assembly mutants, double deletion of cox14 or coa3 did not recover Cox1 synthesis. Our results show that Pet54 is a positive regulator of Cox1 synthesis that renders Mss51 competent as a translational activator of the COX1 mRNA and that this role is independent of the assembly feedback regulatory loop of Cox1 synthesis. Pet54 may play a role in Mss51 hemylation/conformational change necessary for translational activity. Moreover, Pet54 physically interacts with the COX1 mRNA, and this binding was independent of the presence of Mss51.


Assuntos
Complexo IV da Cadeia de Transporte de Elétrons/biossíntese , Proteínas Mitocondriais/biossíntese , Biossíntese de Proteínas/fisiologia , Proteínas de Ligação a RNA/metabolismo , Proteínas de Saccharomyces cerevisiae/biossíntese , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Regiões 5' não Traduzidas/fisiologia , Complexo IV da Cadeia de Transporte de Elétrons/genética , Proteínas Mitocondriais/genética , RNA Fúngico/genética , RNA Fúngico/metabolismo , Proteínas de Ligação a RNA/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
7.
RNA Biol ; 11(7): 953-67, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25181249

RESUMO

Mitochondrial synthesis of Cox1, the largest subunit of the cytochrome c oxidase complex, is controlled by Mss51 and Pet309, two mRNA-specific translational activators that act via the COX1 mRNA 5'-UTR through an unknown mechanism. Pet309 belongs to the pentatricopeptide repeat (PPR) protein family, which is involved in RNA metabolism in mitochondria and chloroplasts, and its sequence predicts at least 12 PPR motifs in the central portion of the protein. Deletion of these motifs selectively disrupted translation but not accumulation of the COX1 mRNA. We used RNA coimmunoprecipitation assays to show that Pet309 interacts with the COX1 mRNA in vivo and that this association is present before processing of the COX1 mRNA from the ATP8/6 polycistronic mRNA. This association was not affected by deletion of 8 of the PPR motifs but was undetectable after deletion of the entire 12-PPR region. However, interaction of the Pet309 protein lacking 12 PPR motifs with the COX1 mRNA was detected after overexpression of the mutated form of the protein, suggesting that deletion of this region decreased the binding affinity for the COX1 mRNA without abolishing it entirely. Moreover, binding of Pet309 to the COX1 mRNA was affected by deletion of Mss51. This work demonstrates an in vivo physical interaction between a yeast mitochondrial translational activator and its target mRNA and shows the cooperativity of the PPR domains of Pet309 in interaction with the COX1 mRNA.


Assuntos
Complexo IV da Cadeia de Transporte de Elétrons/genética , Proteínas de Membrana/metabolismo , Proteínas Mitocondriais/metabolismo , Fatores de Iniciação de Peptídeos/metabolismo , RNA Mensageiro/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Sítios de Ligação , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Regulação Fúngica da Expressão Gênica , Proteínas de Membrana/genética , Proteínas Mitocondriais/genética , Mutação , Fatores de Iniciação de Peptídeos/genética , RNA Fúngico/metabolismo , Ribossomos/metabolismo , Saccharomyces cerevisiae/genética , Fatores de Transcrição/genética
8.
Nat Commun ; 14(1): 8501, 2023 Dec 27.
Artigo em Inglês | MEDLINE | ID: mdl-38151585

RESUMO

DNA polymerase (DNAP) can correct errors in DNA during replication by proofreading, a process critical for cell viability. However, the mechanism by which an erroneously incorporated base translocates from the polymerase to the exonuclease site and the corrected DNA terminus returns has remained elusive. Here, we present an ensemble of nine high-resolution structures representing human mitochondrial DNA polymerase Gamma, Polγ, captured during consecutive proofreading steps. The structures reveal key events, including mismatched base recognition, its dissociation from the polymerase site, forward translocation of DNAP, alterations in DNA trajectory, repositioning and refolding of elements for primer separation, DNAP backtracking, and displacement of the mismatched base into the exonuclease site. Altogether, our findings suggest a conserved 'bolt-action' mechanism of proofreading based on iterative cycles of DNAP translocation without dissociation from the DNA, facilitating primer transfer between catalytic sites. Functional assays and mutagenesis corroborate this mechanism, connecting pathogenic mutations to crucial structural elements in proofreading steps.


Assuntos
Replicação do DNA , DNA Polimerase Dirigida por DNA , Humanos , Replicação do DNA/genética , DNA Polimerase Dirigida por DNA/metabolismo , DNA/genética , DNA/química , Exonucleases/metabolismo
9.
Nat Genet ; 55(10): 1632-1639, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37723262

RESUMO

Uniparental inheritance of mitochondrial DNA (mtDNA) is an evolutionary trait found in nearly all eukaryotes. In many species, including humans, the sperm mitochondria are introduced to the oocyte during fertilization1,2. The mechanisms hypothesized to prevent paternal mtDNA transmission include ubiquitination of the sperm mitochondria and mitophagy3,4. However, the causative mechanisms of paternal mtDNA elimination have not been defined5,6. We found that mitochondria in human spermatozoa are devoid of intact mtDNA and lack mitochondrial transcription factor A (TFAM)-the major nucleoid protein required to protect, maintain and transcribe mtDNA. During spermatogenesis, sperm cells express an isoform of TFAM, which retains the mitochondrial presequence, ordinarily removed upon mitochondrial import. Phosphorylation of this presequence prevents mitochondrial import and directs TFAM to the spermatozoon nucleus. TFAM relocalization from the mitochondria of spermatogonia to the spermatozoa nucleus directly correlates with the elimination of mtDNA, thereby explaining maternal inheritance in this species.


Assuntos
DNA Mitocondrial , Herança Materna , Humanos , Masculino , DNA Mitocondrial/genética , Herança Materna/genética , Sêmen/metabolismo , Mitocôndrias/genética , Espermatozoides/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo
10.
Cancer Res ; 83(23): 3861-3867, 2023 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-37668528

RESUMO

International cancer registries make real-world genomic and clinical data available, but their joint analysis remains a challenge. AACR Project GENIE, an international cancer registry collecting data from 19 cancer centers, makes data from >130,000 patients publicly available through the cBioPortal for Cancer Genomics (https://genie.cbioportal.org). For 25,000 patients, additional real-world longitudinal clinical data, including treatment and outcome data, are being collected by the AACR Project GENIE Biopharma Collaborative using the PRISSMM data curation model. Several thousand of these cases are now also available in cBioPortal. We have significantly enhanced the functionalities of cBioPortal to support the visualization and analysis of this rich clinico-genomic linked dataset, as well as datasets generated by other centers and consortia. Examples of these enhancements include (i) visualization of the longitudinal clinical and genomic data at the patient level, including timelines for diagnoses, treatments, and outcomes; (ii) the ability to select samples based on treatment status, facilitating a comparison of molecular and clinical attributes between samples before and after a specific treatment; and (iii) survival analysis estimates based on individual treatment regimens received. Together, these features provide cBioPortal users with a toolkit to interactively investigate complex clinico-genomic data to generate hypotheses and make discoveries about the impact of specific genomic variants on prognosis and therapeutic sensitivities in cancer. SIGNIFICANCE: Enhanced cBioPortal features allow clinicians and researchers to effectively investigate longitudinal clinico-genomic data from patients with cancer, which will improve exploration of data from the AACR Project GENIE Biopharma Collaborative and similar datasets.


Assuntos
Genômica , Neoplasias , Humanos , Neoplasias/genética , Neoplasias/terapia , Medicina de Precisão
11.
BMJ Open ; 12(6): e056295, 2022 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-35710244

RESUMO

INTRODUCTION: Asthma is a growing health problem in children in marginalised urban settings in low-income and middle-income countries. Asthma attacks are an important cause of emergency care attendance and long-term morbidity. We designed a prospective study, the Asthma Attacks study, to identify factors associated with recurrence of asthma attacks (or exacerbations) among children and adolescents attending emergency care in three Ecuadorian cities. METHODS AND ANALYSIS: Prospective cohort study designed to identify risk factors associated with recurrence of asthma attacks in 450 children and adolescents aged 5-17 years attending emergency care in public hospitals in three Ecuadorian cities (Quito, Cuenca and Portoviejo). The primary outcome will be rate of asthma attack recurrence during up to 12 months of follow-up. Data are being collected at baseline and during follow-up by questionnaire: sociodemographic data, asthma history and management (baseline only); recurrence of asthma symptoms and attacks (monthly); economic costs of asthma to family; Asthma Control Test; Pediatric Asthma Quality of life Questionnaire; and Newcastle Asthma Knowledge Questionnaire (baseline only). In addition, the following are being measured at baseline and during follow-up: lung function and reversibility by spirometry before and after salbutamol; fractional exhaled nitric oxide (FeNO); and presence of IgG antibodies to SARS-CoV-2 in blood. Recruitment started in 2019 but because of severe disruption to emergency services caused by the COVID-19 pandemic, eligibility criteria were modified to include asthmatic children with uncontrolled symptoms and registered with collaborating hospitals. Data will be analysed using logistic regression and survival analyses. ETHICS AND DISSEMINATION: Ethical approval was obtained from the Hospital General Docente de Calderon (CEISH-HGDC 2019-001) and Ecuadorian Ministry of Public Health (MSP-CGDES-2021-0041-O N° 096-2021). The study results will be disseminated through presentations at conferences and to key stakeholder groups including policy-makers, postgraduate theses, peer-review publications and a study website. Participants gave informed consent to participate in the study before taking part.


Assuntos
Asma , COVID-19 , Adolescente , Asma/diagnóstico , Asma/epidemiologia , Asma/terapia , COVID-19/epidemiologia , Criança , Cidades/epidemiologia , Equador/epidemiologia , Humanos , Pandemias , Estudos Prospectivos , Qualidade de Vida , SARS-CoV-2
12.
JCO Clin Cancer Inform ; 6: e2100144, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35148171

RESUMO

PURPOSE: Interpretation of genomic variants in tumor samples still presents a challenge in research and the clinical setting. A major issue is that information for variant interpretation is fragmented across disparate databases, and aggregation of information from these requires building extensive infrastructure. To this end, we have developed Genome Nexus, a one-stop shop for variant annotation with a user-friendly interface for cancer researchers and clinicians. METHODS: Genome Nexus (1) aggregates variant information from sources that are relevant to cancer research and clinical applications, (2) allows high-performance programmatic access to the aggregated data via a unified application programming interface, (3) provides a reference page for individual cancer variants, (4) provides user-friendly tools for annotating variants in patients, and (5) is freely available under an open source license and can be installed in a private cloud or local environment and integrated with local institutional resources. RESULTS: Genome Nexus is available at https://www.genomenexus.org. It displays annotations from more than a dozen resources including those that provide variant effect information (variant effect predictor), protein sequence annotation (Uniprot, Pfam, and dbPTM), functional consequence prediction (Polyphen-2, Mutation Assessor, and SIFT), population prevalences (gnomAD, dbSNP, and ExAC), cancer population prevalences (Cancer hotspots and SignalDB), and clinical actionability (OncoKB, CIViC, and ClinVar). We describe several use cases that demonstrate the utility of Genome Nexus to clinicians, researchers, and bioinformaticians. We cover single-variant annotation, cohort analysis, and programmatic use of the application programming interface. Genome Nexus is unique in providing a user-friendly interface specific to cancer that allows high-performance annotation of any variant including unknown ones. CONCLUSION: Interpretation of cancer genomic variants is improved tremendously by having an integrated resource for annotations. Genome Nexus is freely available under an open source license.


Assuntos
Neoplasias , Software , Genômica , Humanos , Anotação de Sequência Molecular , Mutação , Neoplasias/genética
13.
JCO Clin Cancer Inform ; 5: 221-230, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33625877

RESUMO

PURPOSE: Cancer classification is foundational for patient care and oncology research. Systems such as International Classification of Diseases for Oncology (ICD-O), Systematized Nomenclature of Medicine Clinical Terms (SNOMED-CT), and National Cancer Institute Thesaurus (NCIt) provide large sets of cancer classification terminologies but they lack a dynamic modernized cancer classification platform that addresses the fast-evolving needs in clinical reporting of genomic sequencing results and associated oncology research. METHODS: To meet these needs, we have developed OncoTree, an open-source cancer classification system. It is maintained by a cross-institutional committee of oncologists, pathologists, scientists, and engineers, accessible via an open-source Web user interface and an application programming interface. RESULTS: OncoTree currently includes 868 tumor types across 32 organ sites. OncoTree has been adopted as the tumor classification system for American Association for Cancer Research (AACR) Project Genomics Evidence Neoplasia Information Exchange (GENIE), a large genomic and clinical data-sharing consortium, and for clinical molecular testing efforts at Memorial Sloan Kettering Cancer Center and Dana-Farber Cancer Institute. It is also used by precision oncology tools such as OncoKB and cBioPortal for Cancer Genomics. CONCLUSION: OncoTree is a dynamic and flexible community-driven cancer classification platform encompassing rare and common cancers that provides clinically relevant and appropriately granular cancer classification for clinical decision support systems and oncology research.


Assuntos
Neoplasias , Genômica , Humanos , Oncologia , National Cancer Institute (U.S.) , Neoplasias/diagnóstico , Neoplasias/genética , Neoplasias/terapia , Medicina de Precisão , Estados Unidos
14.
Glob Public Health ; 14(6-7): 954-962, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30929572

RESUMO

The recognition of transgender women (TGW) as the most vulnerable population to HIV/AIDS in Peru and their inclusion as a specific key affected population in health research was the outcome of an extended process that culminated when TGW community organisations succeeded in articulating themselves as a population separate from men who have sex with men (MSM) and, in alliance with some academic research groups, documented their HIV prevalence and vulnerability factors. Prior to that process, TGW remained subsumed under the epidemiological category of men who have sex with men (MSM), invisible in the context of public health policies. Based on a growing body of academic research evidence, coupled with the increasing number and capacities of TGW representatives in technical and policy-related gatherings, a consensus emerged for the establishment of TGW health statistics separate from MSM by 2010. During the past decade, social and health research has contributed conclusive evidence on the living conditions of TGW and the structural barriers they face, beyond the focus of HIV/AIDS research. Despite such progress, pervasive barriers in public policies continue to hinder the use of existing research evidence and community experience in the development of sensitive HIV prevention and care strategies as part of a comprehensive health model for TGW in Peru.


Assuntos
Direito à Saúde , Pessoas Transgênero , Feminino , Infecções por HIV/epidemiologia , Infecções por HIV/prevenção & controle , Humanos , Peru/epidemiologia , Política Pública , Populações Vulneráveis
15.
J Clin Invest ; 129(10): 4276-4289, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31483290

RESUMO

BACKGROUNDAdenoid cystic carcinoma (ACC) is a rare malignancy arising in salivary glands and other sites, characterized by high rates of relapse and distant spread. Recurrent/metastatic (R/M) ACCs are generally incurable, due to a lack of active systemic therapies. To improve outcomes, deeper understanding of genetic alterations and vulnerabilities in R/M tumors is needed.METHODSAn integrated genomic analysis of 1,045 ACCs (177 primary, 868 R/M) was performed to identify alterations associated with advanced and metastatic tumors. Intratumoral genetic heterogeneity, germline mutations, and therapeutic actionability were assessed.RESULTSCompared with primary tumors, R/M tumors were enriched for alterations in key Notch (NOTCH1, 26.3% vs. 8.5%; NOTCH2, 4.6% vs. 2.3%; NOTCH3, 5.7% vs. 2.3%; NOTCH4, 3.6% vs. 0.6%) and chromatin-remodeling (KDM6A, 15.2% vs. 3.4%; KMT2C/MLL3, 14.3% vs. 4.0%; ARID1B, 14.1% vs. 4.0%) genes. TERT promoter mutations (13.1% of R/M cases) were mutually exclusive with both NOTCH1 mutations (q = 3.3 × 10-4) and MYB/MYBL1 fusions (q = 5.6 × 10-3), suggesting discrete, alternative mechanisms of tumorigenesis. This network of alterations defined 4 distinct ACC subgroups: MYB+NOTCH1+, MYB+/other, MYBWTNOTCH1+, and MYBWTTERT+. Despite low mutational load, we identified numerous samples with marked intratumoral genetic heterogeneity, including branching evolution across multiregion sequencing.CONCLUSIONThese observations collectively redefine the molecular underpinnings of ACC progression and identify further targets for precision therapies.FUNDINGAdenoid Cystic Carcinoma Research Foundation, Pershing Square Sohn Cancer Research grant, the PaineWebber Chair, Stand Up 2 Cancer, NIH R01 CA205426, the STARR Cancer Consortium, NCI R35 CA232097, the Frederick Adler Chair, Cycle for Survival, the Jayme Flowers Fund, The Sebastian Nativo Fund, NIH K08 DE024774 and R01 DE027738, and MSKCC through NIH/NCI Cancer Center Support Grant (P30 CA008748).


Assuntos
Carcinoma Adenoide Cístico/genética , Mutação , Adulto , Carcinoma Adenoide Cístico/patologia , Carcinoma Adenoide Cístico/secundário , Montagem e Desmontagem da Cromatina/genética , Feminino , Genes myb , Genômica , Humanos , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Receptores Notch/genética , Neoplasias das Glândulas Salivares/genética , Neoplasias das Glândulas Salivares/patologia , Telomerase/genética
16.
Sci Data ; 5: 180061, 2018 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-29664468

RESUMO

Driven by the recent advances of next generation sequencing (NGS) technologies and an urgent need to decode complex human diseases, a multitude of large-scale studies were conducted recently that have resulted in an unprecedented volume of whole transcriptome sequencing (RNA-seq) data, such as the Genotype Tissue Expression project (GTEx) and The Cancer Genome Atlas (TCGA). While these data offer new opportunities to identify the mechanisms underlying disease, the comparison of data from different sources remains challenging, due to differences in sample and data processing. Here, we developed a pipeline that processes and unifies RNA-seq data from different studies, which includes uniform realignment, gene expression quantification, and batch effect removal. We find that uniform alignment and quantification is not sufficient when combining RNA-seq data from different sources and that the removal of other batch effects is essential to facilitate data comparison. We have processed data from GTEx and TCGA and successfully corrected for study-specific biases, enabling comparative analysis between TCGA and GTEx. The normalized datasets are available for download on figshare.


Assuntos
Neoplasias/genética , RNA Neoplásico , RNA , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Análise de Sequência de RNA
17.
PLoS One ; 10(9): e0136929, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26327321

RESUMO

Protein degradation via the multistep ubiquitin/26S proteasome pathway is a rapid way to alter the protein profile and drive cell processes and developmental changes. Many key regulators of embryonic development are targeted for degradation by E3 ubiquitin ligases. The most studied family of E3 ubiquitin ligases is the SCF ubiquitin ligases, which use F-box adaptor proteins to recognize and recruit target proteins. Here, we used a bioinformatics screen and phylogenetic analysis to identify and annotate the family of F-box proteins in the Xenopus tropicalis genome. To shed light on the function of the F-box proteins, we analyzed expression of F-box genes during early stages of Xenopus development. Many F-box genes are broadly expressed with expression domains localized to diverse tissues including brain, spinal cord, eye, neural crest derivatives, somites, kidneys, and heart. All together, our genome-wide identification and expression profiling of the Xenopus F-box family of proteins provide a foundation for future research aimed to identify the precise role of F-box dependent E3 ubiquitin ligases and their targets in the regulatory circuits of development.


Assuntos
Proteínas F-Box/genética , Xenopus/genética , Animais , Regulação da Expressão Gênica no Desenvolvimento/genética , Estudo de Associação Genômica Ampla/métodos , Filogenia , Proteólise , Proteínas Ligases SKP Culina F-Box/genética , Ubiquitina-Proteína Ligases/genética
18.
Brain Res ; 1565: 8-17, 2014 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-24675025

RESUMO

Interactions between neural progenitor cells (NPC) and endothelial cells (EC) from adult vascular beds have been well explored previously. However, the factors and signaling mechanisms that regulate neurogenesis and angiogenesis are most prevalent during embryonic development. This study aimed to determine whether embryonic brain endothelial cells from the periventricular region (PVEC) present an advantage over adult brain EC in supporting NPC growth and differentiation. PVEC were isolated from E15 mouse brains, processed, and sorted with immunomagnetic beads using antibodies against CD31/PECAM. On immunofluorescence (IF) staining, nearly all cells were positive for EC markers CD31 and CD144/VE-Cadherin. In proliferation studies, NPC proliferation was highest in transwell co-culture with PVEC, approximately 2.3 fold increase compared to baseline versus 1.4 fold increase when co-cultured with adult brain endothelial cells (ABEC). These results correlated with the PVEC mediated delay in NPC differentiation, evidenced by high expression of progenitor marker Nestin evaluated by IF staining. Upon further characterization of PVEC in an angiogenesis assay measuring cord length, PVEC exhibited a high capacity to form cords in basal conditions compared to ABEC. This was enhanced in the presence of NPC, with both cell types displaying a preferential structural alignment resembling neurovascular networks. PVEC also expressed high Vegfa levels at baseline in comparison to NPC and ABEC. Vegfa levels increased when co-cultured with NPC. We demonstrate that PVEC and NPC co-cultures act synergistically to promote the formation of a neurovascular unit through dynamic and reciprocal communication. Our results suggest that PVEC/NPC could provide promising neuro-regenerative therapies for patients suffering brain injuries.


Assuntos
Ventrículos Cerebrais/embriologia , Células Endoteliais/citologia , Neovascularização Fisiológica/fisiologia , Células-Tronco Neurais/citologia , Animais , Células Cultivadas , Células Endoteliais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Neurais/metabolismo
19.
Biotechnol Prog ; 29(1): 230-6, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23074091

RESUMO

Phospholipids are a biologically and industrially important class of compounds whose physical properties can be improved for diverse applications by substitution of medium-chain fatty acids for their native fatty acid chains. In this study, phosphatidylcholine (PC) was enriched with medium-chain fatty acids (MCFAs) by acidolysis with phospholipase A(1) (PLA(1) ) immobilized on Duolite A568. Response surface methodology was employed to evaluate the effects of the molar ratio of substrates (PC to free MCFAs), enzyme loading, and reaction temperature on the incorporation of free MCFAs into PC and on PC recovery. Enzyme loading and molar ratio of substrates contributed positively, but temperature negatively, to the incorporation of free MCFAs into PC. Increases in enzyme loading and the molar ratio of PC to free MCFAs led to increased incorporation of the latter into the former, but increased temperature had the opposite effect. By contrast, an increase in enzyme loading led to decreased PC recovery. Increased temperature had also a negative effect on PC recovery. Optimal conditions for maximum incorporation and PC recovery were molar ratio of PC to free MCFAs of 1:16, enzyme loading of 16%, and 50°C. Under these conditions, the incorporation of free MCFAs was 41% and the PC recovery was 53%.


Assuntos
Enzimas Imobilizadas/metabolismo , Ácidos Graxos/metabolismo , Fosfatidilcolinas/metabolismo , Fosfolipases A1/metabolismo , Biocatálise , Enzimas Imobilizadas/química , Ácidos Graxos/química , Fosfatidilcolinas/química , Fosfolipases A1/química , Propriedades de Superfície , Temperatura
20.
J Biol Chem ; 283(3): 1472-1479, 2008 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-18039654

RESUMO

Pet309 is a protein essential for respiratory growth. It is involved in translation of the yeast mitochondrial COX1 gene, which encodes subunit I of the cytochrome c oxidase. Pet309 is also involved in stabilization of the COX1 mRNA. Mutations in a similar human protein, Lrp130, are associated with Leigh syndrome, where cytochrome c oxidase activity is affected. The sequence of Pet309 reveals the presence of at least seven pentatricopeptide repeats (PPRs) located in tandem in the central portion of the protein. Proteins containing PPR motifs are present in mitochondria and chloroplasts and are in general involved in RNA metabolism. Despite the increasing number of proteins from this family found to play essential roles in mitochondria and chloroplasts, little is understood about the mechanism of action of the PPR domains present in these proteins. In a series of in vivo analyses we constructed a pet309 mutant lacking the PPR motifs. Although the stability of the COX1 mRNA was not affected, synthesis of Cox1 was abolished. The deletion of one PPR motif at a time showed that all the PPR motifs are required for COX1 mRNA translation and respiratory growth. Mutations of basic residues in PPR3 caused reduced respiratory growth. According to a molecular model, these residues are facing a central cavity that could be involved in mRNA-binding activity, forming a possible path for this molecule on Pet309. Our results show that the RNA metabolism function of Pet309 is found in at least two separate domains of the protein.


Assuntos
Complexo IV da Cadeia de Transporte de Elétrons/genética , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Biossíntese de Proteínas , Estabilidade de RNA , RNA Mensageiro/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/enzimologia , Motivos de Aminoácidos , Aminoácidos , Complexo IV da Cadeia de Transporte de Elétrons/biossíntese , Regulação Fúngica da Expressão Gênica , Mitocôndrias/metabolismo , Proteínas Mitocondriais , Modelos Moleculares , Mutagênese , Fatores de Iniciação de Peptídeos , Estrutura Terciária de Proteína , Transporte Proteico , RNA Fúngico/metabolismo , RNA Mitocondrial , Sequências Repetitivas de Aminoácidos , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Proteínas de Saccharomyces cerevisiae/biossíntese , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/metabolismo , Relação Estrutura-Atividade
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