RESUMEN
Colorectal cancers are one of the most prevalent tumour types worldwide and, despite the emergence of targeted and biologic therapies, have among the highest mortality rates. The Personalized OncoGenomics (POG) program at BC Cancer performs whole genome and transcriptome analysis (WGTA) to identify specific alterations in an individual's cancer that may be most effectively targeted. Informed using WGTA, a patient with advanced mismatch repair-deficient colorectal cancer was treated with the antihypertensive drug irbesartan and experienced a profound and durable response. We describe the subsequent relapse of this patient and potential mechanisms of response using WGTA and multiplex immunohistochemistry (m-IHC) profiling of biopsies before and after treatment from the same metastatic site of the L3 spine. We did not observe marked differences in the genomic landscape before and after treatment. Analyses revealed an increase in immune signalling and infiltrating immune cells, particularly CD8+ T cells, in the relapsed tumour. These results indicate that the observed anti-tumour response to irbesartan may have been due to an activated immune response. Determining whether there may be other cancer contexts in which irbesartan may be similarly valuable will require additional studies.
Asunto(s)
Antihipertensivos , Neoplasias Colorrectales , Humanos , Irbesartán/uso terapéutico , Antihipertensivos/uso terapéutico , Linfocitos T CD8-positivos/patología , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patologíaRESUMEN
BACKGROUND: Recent advances are enabling delivery of precision genomic medicine to cancer clinics. While the majority of approaches profile panels of selected genes or hotspot regions, comprehensive data provided by whole-genome and transcriptome sequencing and analysis (WGTA) present an opportunity to align a much larger proportion of patients to therapies. PATIENTS AND METHODS: Samples from 570 patients with advanced or metastatic cancer of diverse types enrolled in the Personalized OncoGenomics (POG) program underwent WGTA. DNA-based data, including mutations, copy number and mutation signatures, were combined with RNA-based data, including gene expression and fusions, to generate comprehensive WGTA profiles. A multidisciplinary molecular tumour board used WGTA profiles to identify and prioritize clinically actionable alterations and inform therapy. Patient responses to WGTA-informed therapies were collected. RESULTS: Clinically actionable targets were identified for 83% of patients, of which 37% of patients received WGTA-informed treatments. RNA expression data were particularly informative, contributing to 67% of WGTA-informed treatments; 25% of treatments were informed by RNA expression alone. Of a total 248 WGTA-informed treatments, 46% resulted in clinical benefit. RNA expression data were comparable to DNA-based mutation and copy number data in aligning to clinically beneficial treatments. Genome signatures also guided therapeutics including platinum, poly-ADP ribose polymerase inhibitors and immunotherapies. Patients accessed WGTA-informed treatments through clinical trials (19%), off-label use (35%) and as standard therapies (46%) including those which would not otherwise have been the next choice of therapy, demonstrating the utility of genomic information to direct use of chemotherapies as well as targeted therapies. CONCLUSIONS: Integrating RNA expression and genome data illuminated treatment options that resulted in 46% of treated patients experiencing positive clinical benefit, supporting the use of comprehensive WGTA profiling in clinical cancer care.
Asunto(s)
Neoplasias , Perfilación de la Expresión Génica , Genómica/métodos , Humanos , Mutación , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Medicina de Precisión/métodos , ARN , TranscriptomaRESUMEN
BACKGROUND: NRG1 fusion-positive lung cancers have emerged as potentially actionable events in lung cancer, but clinical support is currently limited and no evidence of efficacy of this approach in cancers beyond lung has been shown. PATIENTS AND METHODS: Here, we describe two patients with advanced cancers refractory to standard therapies. Patient 1 had lung adenocarcinoma and patient 2 cholangiocarcinoma. Whole-genome and transcriptome sequencing were carried out for these cases with select findings validated by fluorescence in situ hybridization. RESULTS: Both tumors were found to be positive for NRG1 gene fusions. In patient 1, an SDC4-NRG1 gene fusion was detected, similar gene fusions having been described in lung cancers previously. In patient 2, a novel ATP1B1-NRG1 gene fusion was detected. Cholangiocarcinoma is not a disease type in which NRG1 fusions had been described previously. Integrative genome analysis was used to assess the potential functional significance of the detected genomic events including the gene fusions, prioritizing therapeutic strategies targeting the HER-family of growth factor receptors. Both patients were treated with the pan HER-family kinase inhibitor afatinib and both displayed significant and durable response to treatment. Upon progression sites of disease were sequenced. The lack of obvious genomic events to describe the disease progression indicated that broad transcriptomic or epigenetic mechanisms could be attributed to the lack of prolonged response to afatinib. CONCLUSION: These observations lend further support to the use of pan HER-tyrosine kinase inhibitors for the treatment of NRG1 fusion-positive in both cancers of lung and hepatocellular origin and indicate more broadly that cancers found to be NRG1 fusion-positive may benefit from such a clinical approach regardless of their site of origin. CLINICAL TRIAL INFORMATION: Personalized Oncogenomics (POG) Program of British Columbia: Utilization of Genomic Analysis to Better Understand Tumour Heterogeneity and Evolution (NCT02155621).
Asunto(s)
Adenocarcinoma/tratamiento farmacológico , Neoplasias de los Conductos Biliares/tratamiento farmacológico , Colangiocarcinoma/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Neurregulina-1/genética , Neurregulina-1/metabolismo , Quinazolinas/uso terapéutico , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma del Pulmón , Adulto , Afatinib , Neoplasias de los Conductos Biliares/genética , Neoplasias de los Conductos Biliares/metabolismo , Colangiocarcinoma/genética , Colangiocarcinoma/metabolismo , Femenino , Perfilación de la Expresión Génica , Humanos , Hibridación Fluorescente in Situ , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Proteínas de Fusión Oncogénica/genética , Proteínas de Fusión Oncogénica/metabolismo , Inhibidores de Proteínas Quinasas/uso terapéutico , Sindecano-4/genéticaRESUMEN
BACKGROUND: A patient suffering from metastatic colorectal cancer, treatment-related toxicity and resistance to standard chemotherapy and radiation was assessed as part of a personalized oncogenomics initiative to derive potential alternative therapeutic strategies. PATIENTS AND METHODS: Whole-genome and transcriptome sequencing was used to interrogate a metastatic tumor refractory to standard treatments of a patient with mismatch repair-deficient metastatic colorectal cancer. RESULTS: Integrative genomic analysis indicated overexpression of the AP-1 transcriptional complex suggesting experimental therapeutic rationales, including blockade of the renin-angiotensin system. This led to the repurposing of the angiotensin II receptor antagonist, irbesartan, as an anticancer therapy, resulting in the patient experiencing a dramatic and durable response. CONCLUSIONS: This case highlights the utility of comprehensive integrative genomic profiling and bioinformatics analysis to provide hypothetical rationales for personalized treatment options.
Asunto(s)
Compuestos de Bifenilo/administración & dosificación , Neoplasias Colorrectales/tratamiento farmacológico , Medicina de Precisión , Tetrazoles/administración & dosificación , Factor de Transcripción AP-1/genética , Anciano , Antagonistas de Receptores de Angiotensina/administración & dosificación , Angiotensinas/antagonistas & inhibidores , Angiotensinas/genética , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Biología Computacional , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Irbesartán , Metástasis de la Neoplasia , Sistema Renina-Angiotensina/efectos de los fármacos , Transcriptoma/genéticaRESUMEN
We had previously defined by allele loss studies a minimal region at 6q27 (between D6S264 and D6S297) to contain a putative tumour suppressor gene. The p90 ribosomal S6 kinase-3 gene (p90 Rsk-3, RPS6KA2) maps in this interval. It is a serine-threonine kinase that signals downstream of the mitogen-activated protein kinase pathway. It is expressed in normal ovarian epithelium, whereas reduced or absent in tumours or cell lines. We show that RPS6KA2 is monoallelically expressed in the ovary suggesting that loss of a single expressed allele is sufficient to cause complete loss of expression in cancer cells. Further, we have identified two new isoforms of RPS6KA2 with an alternative start codon. Homozygous deletions were identified within the RPS6KA2 gene in two cell lines. Re-expression of RPS6KA2 in ovarian cancer cell lines suppressed colony formation. In UCI101 cells, the expression of RPS6KA2 reduced proliferation, caused G1 arrest, increased apoptosis, reduced levels of phosphorylated extracellular signal-regulated kinase and altered other cell cycle proteins. In contrast, small interfering RNA against RPS6KA2 showed the opposite effect in 41M cells. The above results suggest that RPS6KA2 is a putative tumour suppressor gene to explain allele loss at 6q27.
Asunto(s)
Cromosomas Humanos Par 6/genética , Genes Supresores de Tumor/fisiología , Neoplasias Glandulares y Epiteliales/genética , Neoplasias Ováricas/genética , Proteínas Quinasas S6 Ribosómicas 90-kDa/metabolismo , Adenocarcinoma de Células Claras/genética , Adenocarcinoma de Células Claras/metabolismo , Adenocarcinoma de Células Claras/patología , Adenocarcinoma Mucinoso/genética , Adenocarcinoma Mucinoso/metabolismo , Adenocarcinoma Mucinoso/patología , Secuencia de Aminoácidos , Apoptosis , Carcinoma Endometrioide/genética , Carcinoma Endometrioide/metabolismo , Carcinoma Endometrioide/patología , Ciclo Celular , Proliferación Celular , Deleción Cromosómica , Mapeo Cromosómico , Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/metabolismo , Cistadenocarcinoma Seroso/patología , Metilación de ADN , ADN de Neoplasias/genética , ADN de Neoplasias/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Inmunoprecipitación , Pérdida de Heterocigocidad , Sistema de Señalización de MAP Quinasas/genética , Proteínas Quinasas Activadas por Mitógenos , Datos de Secuencia Molecular , Neoplasias Glandulares y Epiteliales/metabolismo , Neoplasias Glandulares y Epiteliales/patología , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Reacción en Cadena de la Polimerasa , Polimorfismo Conformacional Retorcido-Simple , Interferencia de ARN , Proteínas Quinasas S6 Ribosómicas 90-kDa/genética , Homología de Secuencia de Aminoácido , Transfección , Células Tumorales CultivadasRESUMEN
Deletions of chromosome 6 are a common abnormality in diverse human malignancies including astrocytic tumours, suggesting the presence of tumour suppressor genes (TSG). In order to help identify candidate TSGs, we have constructed a chromosome 6 tile path microarray. The array contains 1,780 clones (778 P1-derived artificial chromosome and 1,002 bacterial artificial chromosome) that cover 98.3% of the published chromosome 6 sequences. A total of 104 adult astrocytic tumours (10 diffuse astrocytomas, 30 anaplastic astrocytomas (AA), 64 glioblastomas (GB)) were analysed using this array. Single copy number change was successfully detected and the result was in general concordant with a microsatellite analysis. The pattern of copy number change was complex with multiple interstitial deletions/gains. However, a predominance of telomeric 6q deletions was seen. Two small common and overlapping regions of deletion at 6q26 were identified. One was 1,002 kb in size and contained PACRG and QKI, while the second was 199 kb and harbours a single gene, ARID1B. The data show that the chromosome 6 tile path array is useful in mapping copy number changes with high resolution and accuracy. We confirmed the high frequency of chromosome 6 deletions in AA and GB, and identified two novel commonly deleted regions that may harbour TSGs.
Asunto(s)
Astrocitoma/genética , Deleción Cromosómica , Cromosomas Humanos Par 6/genética , Glioblastoma/genética , Hibridación de Ácido Nucleico , Análisis de Secuencia por Matrices de Oligonucleótidos , Astrocitoma/patología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Cromosomas Artificiales Bacterianos , ADN de Neoplasias/análisis , Dosificación de Gen , Glioblastoma/patología , Humanos , Repeticiones de Microsatélite , Telómero/genéticaRESUMEN
Chromosome 6 is a metacentric chromosome that constitutes about 6% of the human genome. The finished sequence comprises 166,880,988 base pairs, representing the largest chromosome sequenced so far. The entire sequence has been subjected to high-quality manual annotation, resulting in the evidence-supported identification of 1,557 genes and 633 pseudogenes. Here we report that at least 96% of the protein-coding genes have been identified, as assessed by multi-species comparative sequence analysis, and provide evidence for the presence of further, otherwise unsupported exons/genes. Among these are genes directly implicated in cancer, schizophrenia, autoimmunity and many other diseases. Chromosome 6 harbours the largest transfer RNA gene cluster in the genome; we show that this cluster co-localizes with a region of high transcriptional activity. Within the essential immune loci of the major histocompatibility complex, we find HLA-B to be the most polymorphic gene on chromosome 6 and in the human genome.
Asunto(s)
Cromosomas Humanos Par 6/genética , Genes/genética , Mapeo Físico de Cromosoma , Animales , Exones/genética , Enfermedades Genéticas Congénitas/genética , Antígenos HLA-B/genética , Humanos , Seudogenes/genética , ARN de Transferencia/genética , Análisis de Secuencia de ADNRESUMEN
We have mapped and sequenced both chromosome breakpoints of a balanced t(6;11)(q14.2;q25) chromosome translocation that segregates with a schizophrenia-like psychosis. Bioinformatics analysis of the regions revealed a number of confirmed and predicted transcripts. No confirmed transcripts are disrupted by either breakpoint. The chromosome 6 breakpoint region is gene poor, the closest transcript being the serotonin receptor 1E (HTR1E) at 625 kb telomeric to the breakpoint. The chromosome 11 breakpoint is situated close to the telomere. The closest gene, beta-1,3-glucuronyltransferase (B3GAT1 or GlcAT-P), is 299 kb centromeric to the breakpoint. B3GAT1 is the key enzyme during the biosynthesis of the carbohydrate epitope HNK-1, which is present on a number of cell adhesion molecules important in neurodevelopment. Mice deleted for the B3GAT1 gene show defects in hippocampal long-term potentiation and in spatial memory formation. We propose that the translocation causes a positional effect on B3GAT1, affecting expression levels and making it a plausible candidate for the psychosis found in this family. More generally, regions close to telomeres are highly polymorphic in both sequence and length in the general population and several studies have implicated subtelomeric deletions as a common cause of idiopathic mental retardation. This leads us to the hypothesis that polymorphic or other variation of the 11q telomere may affect the activity of B3GAT1 and be a risk factor for schizophrenia and related psychoses in the general population.
Asunto(s)
Cromosomas Humanos Par 11/genética , Cromosomas Humanos Par 6/genética , Glucuronosiltransferasa/genética , Trastornos Psicóticos/genética , Telómero/ultraestructura , Translocación Genética , Secuencia de Bases , Rotura Cromosómica , Mapeo Cromosómico/métodos , Cromosomas Humanos Par 11/ultraestructura , Cromosomas Humanos Par 6/ultraestructura , Depresión/genética , Etiquetas de Secuencia Expresada , Femenino , Glucuronosiltransferasa/fisiología , Humanos , Masculino , Datos de Secuencia Molecular , Linaje , Trastornos Psicóticos/epidemiología , Factores de Riesgo , Eliminación de Secuencia , Suicidio , Intento de SuicidioRESUMEN
Chromosomal translocation t(6;14)(p21.1;q32.3) has been reported as a rare but recurrent event not only in myeloma and plasma cell leukemia but also in diffuse large B-cell non-Hodgkin lymphoma (B-NHL) (diffuse large B-cell lymphoma [DLBCL]) and splenic lymphoma with villous lymphocytes (SLVL); however, the nature of the target gene(s) has not been determined. This study identified t(6;14)(p21.1;q32.3) in 3 cases of transformed extranodal marginal zone B-NHL, in 1 case of SLVL, and in 1 case of a low-grade B-cell lymphoproliferative disorder. In a sixth case, a CD5(+) DLBCL, the translocation was identified by molecular cloning in the absence of cytogenetically detectable change. Two chromosomal translocation breakpoints were cloned by using long-distance inverse polymerase chain reaction methods. Comparison with the genomic sequence for chromosome 6p21.1 showed breakpoints approximately 59 and 73.5 kilobases 5' of the cyclin D3 (CCND3) gene with no other identifiable transcribed sequences in the intervening region. Although Southern blotting with derived genomic 6p21.1 probes failed to detect other rearrangements, fluorescent in situ hybridization assays, using BAC (bacterial artificial chromosome) clones spanning and flanking the CCND3 locus, along with probes for IGH confirmed localization of 6p21.1 breakpoints within the same region, as well as fusion of the CCND3 and IGH loci. Furthermore, in all cases, high-level expression of CCND3 was demonstrated at RNA and/or protein levels by Northern and Western blotting and by immunohistochemistry. These data implicate CCND3 as a dominant oncogene in the pathogenesis and transformation in several histologic subtypes of mature B-cell malignancies with t(6;14)(p21.1;q32.3) and suggest that CCND3 overexpression seen in about 10% of DLBCL cases may have a genetic basis.
Asunto(s)
Cromosomas Humanos Par 14 , Cromosomas Humanos Par 6 , Ciclinas/genética , Linfoma de Células B/genética , Trastornos Linfoproliferativos/genética , Translocación Genética/genética , Adulto , Anciano , Linfocitos B/química , Linfocitos B/patología , Linfocitos B/ultraestructura , Secuencia de Bases , Rotura Cromosómica/genética , Clonación Molecular , Ciclina D3 , Femenino , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Inmunohistoquímica , Hibridación Fluorescente in Situ , Linfoma de Células B/química , Linfoma de Células B/patología , Trastornos Linfoproliferativos/patología , Masculino , Persona de Mediana Edad , Datos de Secuencia MolecularRESUMEN
Parkin has been identified as a causative gene of the autosomal recessive juvenile parkinsonism (AR-JP). In this study, we determined the genomic structure of the Parkin gene and identified a core promoter region based on the DNA sequence of 1.4 Mb. The 5'-flanking region contained no apparent TATA or CAAT box elements but several putative cis-elements for various transcription factors. The GC- and CpG-rich regions were observed not only in the 5'-flanking sequence but also in the 5'-part of the first intron of Parkin. We identified an exact starting point of Parkin transcription. A core promoter region was determined by transfecting a series of deletion constructs with a dual luciferase reporter system into human neuroblastoma cells. Furthermore, we located a neighboring novel gene in a head-to-head direction with Parkin with only a 198-bp interval.
Asunto(s)
Regiones Promotoras Genéticas , Secuencia de Bases , Encéfalo/metabolismo , Islas de CpG , Citosina/química , ADN Complementario/metabolismo , Exones , Eliminación de Gen , Biblioteca de Genes , Genes Reporteros , Guanina/química , Humanos , Luciferasas/metabolismo , Modelos Genéticos , Datos de Secuencia Molecular , Músculo Esquelético/metabolismo , Mutagénesis , Mutación , Programas Informáticos , TransfecciónRESUMEN
The mouse Vanin-1 molecule plays a role in thymic reconstitution following damage by irradiation. We recently demonstrated that it is a membrane pantetheinase (EC 3.56.1.-). This molecule is the prototypic member of a larger Vanin family encoded by at least two mouse (Vanin-1 and Vanin-3) and three human (VNN1, VNN2, VNN3) orthologous genes. We now report (1) the structural characterization of the human and mouse Vanin genes and their organization in clusters on the 6q22-24 and 10A2B1 chromosomes, respectively; (2) identification of the human VNN3 gene and the demonstration that the mouse Vanin-3 molecule is secreted by cells, and (3) that the Vanin genes encode different isoforms of the mammalian pantetheinase activity. Thus, the Vanin family represents a novel class of secreted or membrane-associated ectoenzymes. We discuss here their possible role in processes pertaining to tissue repair in the context of oxidative stress.
Asunto(s)
Amidohidrolasas/genética , Moléculas de Adhesión Celular/genética , Cromosomas Humanos Par 6/genética , Proteínas de la Membrana/genética , Familia de Multigenes , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Secuencia Conservada , Proteínas Ligadas a GPI , Humanos , Ratones , Datos de Secuencia Molecular , Isoformas de Proteínas/genética , Homología de Secuencia de Aminoácido , Distribución TisularRESUMEN
Olfactory receptor (OR) loci frequently cluster and are present on most human chromosomes. They are members of the seven transmembrane receptor (7-TM) superfamily and, as such, are part of one of the largest mammalian multigene families, with an estimated copy number of up to 1000 ORs per haploid genome. As their name implies, ORs are known to be involved in the perception of odors and possibly also in other, nonolfaction-related, functions. Here, we report the characterization of ORs that are part of the MHC-linked OR clusters in human and mouse (partial sequence only). These clusters are of particular interest because of their possible involvement in olfaction-driven mate selection. In total, we describe 50 novel OR loci (36 human, 14 murine), making the human MHC-linked cluster the largest sequenced OR cluster in any organism so far. Comparative and phylogenetic analyses confirm the cluster to be MHC-linked but divergent in both species and allow the identification of at least one ortholog that will be useful for future regulatory and functional studies. Quantitative feature analysis shows clear evidence of duplications of blocks of OR genes and reveals the entire cluster to have a genomic environment that is very different from its neighboring regions. Based on in silico transcript analysis, we also present evidence of extensive long-distance splicing in the 5'-untranslated regions and, for the first time, of alternative splicing within the single coding exon of ORs. Taken together with our previous finding that ORs are also polymorphic, the presented data indicate that the expression, function, and evolution of these interesting genes might be more complex than previously thought.
Asunto(s)
Ligamiento Genético/genética , Ligamiento Genético/inmunología , Complejo Mayor de Histocompatibilidad/genética , Familia de Multigenes/genética , Receptores Odorantes/genética , Secuencia de Aminoácidos , Animales , Composición de Base , Secuencia de Bases , Mapeo Cromosómico , Orden Génico , Humanos , Ratones , Datos de Secuencia Molecular , Filogenia , Análisis de Secuencia de ADNRESUMEN
We constructed maps for eight chromosomes (1, 6, 9, 10, 13, 20, X and (previously) 22), representing one-third of the genome, by building landmark maps, isolating bacterial clones and assembling contigs. By this approach, we could establish the long-range organization of the maps early in the project, and all contig extension, gap closure and problem-solving was simplified by containment within local regions. The maps currently represent more than 94% of the euchromatic (gene-containing) regions of these chromosomes in 176 contigs, and contain 96% of the chromosome-specific markers in the human gene map. By measuring the remaining gaps, we can assess chromosome length and coverage in sequenced clones.
Asunto(s)
Cromosomas Humanos Par 10 , Cromosomas Humanos Par 13 , Cromosomas Humanos Par 1 , Cromosomas Humanos Par 20 , Cromosomas Humanos Par 6 , Mapeo Contig , Genoma Humano , Cromosoma X , HumanosRESUMEN
Recombination is the exchange of information between two homologous chromosomes during meiosis. The rate of recombination per nucleotide, which profoundly affects the evolution of chromosomal segments, is calculated by comparing genetic and physical maps. Human physical maps have been constructed using cytogenetics, overlapping DNA clones and radiation hybrids; but the ultimate and by far the most accurate physical map is the actual nucleotide sequence. The completion of the draft human genomic sequence provides us with the best opportunity yet to compare the genetic and physical maps. Here we describe our estimates of female, male and sex-average recombination rates for about 60% of the genome. Recombination rates varied greatly along each chromosome, from 0 to at least 9 centiMorgans per megabase (cM Mb(-1)). Among several sequence and marker parameters tested, only relative marker position along the metacentric chromosomes in males correlated strongly with recombination rate. We identified several chromosomal regions up to 6 Mb in length with particularly low (deserts) or high (jungles) recombination rates. Linkage disequilibrium was much more common and extended for greater distances in the deserts than in the jungles.
Asunto(s)
Mapeo Físico de Cromosoma , Recombinación Genética , Femenino , Humanos , Desequilibrio de Ligamiento , Masculino , Caracteres Sexuales , Secuencias Repetidas en TándemRESUMEN
Two new genes were cloned from region 6q21 and characterized. One gene, C6orf4-6, expresses three mRNA isoforms diverging at the 5' and 3' ends, and encodes two protein isoforms that differ by nine amino acids at their amino terminus. The second gene, C6UAS, is transcribed in the antisense orientation from the complementary strand of C6orf4-6. C6UAS overlaps the second exon of C6orf4, where the start codon of protein isoform 1 is located. C6UAS has no apparent ORF and most likely represents a structural RNA gene that is transcribed but not translated. This feature and the antisense polarity of transcription suggest that C6UAS could play a regulatory role on the expression of C6orf4, as indicated by a significant decrease of endogenous C6orf4 expression after transfection of C6UAS cDNA in human fibroblasts. Neither C6UAS nor C6orf4-6 genes show any homology with known human genes. The two genes were cloned from a subregion at 6q21 containing a replicative senescence gene, a tumor suppressor gene and a gene involved in hereditary schizophrenia. In addition, the common fragile site FRA6F was mapped in the same region. Cloning and characterization of C6orf4-6 and C6UAS may help to clarify the structure and the functional role of this important region.
Asunto(s)
Senescencia Celular/genética , Cromosomas Humanos Par 6/genética , Genes Sobrepuestos/genética , Proteínas/genética , Esquizofrenia/genética , Secuencia de Aminoácidos , Secuencia de Bases , Northern Blotting , Línea Celular , Línea Celular Transformada , Mapeo Cromosómico , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , Exones , Femenino , Expresión Génica , Genes/genética , Humanos , Intrones , Masculino , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Estructura Secundaria de Proteína , ARN/química , ARN/genética , ARN/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Análisis de Secuencia de ADN , Distribución Tisular , Transcripción GenéticaAsunto(s)
Cromosomas Humanos Par 6/genética , Clonación Molecular , Biología Computacional , Bases de Datos Factuales , Genes , Enfermedades Genéticas Congénitas/genética , Humanos , Complejo Mayor de Histocompatibilidad/genética , Mapeo Físico de Cromosoma , Análisis de Secuencia de ADN , Transcripción Genética/genéticaRESUMEN
Genetic studies have previously assigned a quantitative trait locus (QTL) for hemoglobin F and F cells to a region of approximately 4 Mb between the markers D6S408 and D6S292 on chromosome 6q23. An initial yeast artificial chromosome contig of 13 clones spanning this region was generated. Further linkage analysis of an extended kindred refined the candidate interval to 1-2 cM, and key recombination events now place the QTL within a region of <800 kb. We describe a high-resolution bacterial clone contig spanning 3 Mb covering this critical region. The map consists of 223 bacterial artificial chromosome (BAC) and 100 P1 artificial chromosome (PAC) clones ordered by sequence-tagged site (STS) content and restriction fragment fingerprinting with a minimum tiling path of 22 BACs and 1 PAC. A total of 194 STSs map to this interval of 3 Mb, giving an average marker resolution of approximately one per 15 kb. About half of the markers were novel and were isolated in the present study, including three CA repeats and 13 single nucleotide polymorphisms. Altogether 24 expressed sequence tags, 6 of which are unique genes, have been mapped to the contig.
Asunto(s)
Cromosomas Humanos Par 6/genética , Mapeo Contig , Hemoglobina Fetal/genética , Carácter Cuantitativo Heredable , Secuencia de Bases , Cromosomas Artificiales de Levadura , Etiquetas de Secuencia Expresada , Femenino , Ligamiento Genético , Haplotipos , Humanos , Masculino , Datos de Secuencia Molecular , Linaje , Reacción en Cadena de la Polimerasa , Polimorfismo de Longitud del Fragmento de RestricciónRESUMEN
Recently, we reported the localization of a gene for transient neonatal diabetes mellitus (TNDM), a rare form of childhood diabetes, to an approximately 5.4 Mb region of chromosome 6q24. We have also shown that TNDM is associated with both paternal uniparental disomy (UPD) of chromosome 6 and paternal duplications of the critical region. The sequencing of P1-derived artificial chromosome clones from within the region of interest has allowed us to further localize the gene and to investigate the methylation status of the region. The gene is now known to reside in a 300-400 kb region of 6q24 which contains several CpG islands. At one island we have demonstrated differential DNA methylation between patients with paternal UPD of chromosome 6 and normal controls. In addition, two patients with TNDM, in whom neither paternal UPD of chromosome 6 nor duplication of 6q24 have been found, show a DNA methylation pattern identical to that of patients with paternal UPD of chromosome 6. Control individuals show a hemizygous methylation pattern. These results show that TNDM can be associated with a methylation change and identify a novel methylation imprint on chromosome 6 associated with TNDM.