120 research outputs found

    The transcription factor Snf1p is involved in a Tup1p-independent manner in the glucose regulation of the major methanol metabolism genes of Hansenula polymorpha

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    Hansenula polymorpha is a methylotrophic yeast widely employed in biotechnology as a ''protein factory''. Most promoters used for heterologous protein expression, like MOX (methanol oxidase) and DAS (di-hydroxy acetone synthase), are involved in the peroxisomal methanol metabolism (C1 metabolism) and are under strong glucose repression. Interestingly, the MOX promoter is subjected to glucose regulation also in Saccharomyces cerevisiae, a non-methylotrophic yeast in which this phenomenon is well studied. In this species, the transcription factor Tup1p plays an essential role in glucose repression of several genes. This effect is counteracted by the activator Snf1p when glucose is exhausted from medium. Therefore, to test whether this regulatory circuit has been conserved in H. polymorpha, HpTUP1 and HpSNF1 were partially cloned and disrupted. Deletion of HpTUP1 did not affect glucose repression of the major C1 metabolism genes (MOX, DAS). Thus, though conserved, HpTUP1 does not seem to take part in a general glucose repression in H. polymorpha. In contrast, the deletion of HpSNF1 led to significant decreases in the activation of these genes in the absence of glucose. Therefore, the effect of HpSNF1 in transcriptional activation may be through an HpTUP1- independent circuit.521528Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

    No-match ORESTES explored as tumor markers

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    Sequencing technologies and new bioinformatics tools have led to the complete sequencing of various genomes. However, information regarding the human transcriptome and its annotation is yet to be completed. The Human Cancer Genome Project, using ORESTES (open reading frame EST sequences) methodology, contributed to this objective by generating data from about 1.2 million expressed sequence tags. Approximately 30% of these sequences did not align to ESTs in the public databases and were considered no-match ORESTES. On the basis that a set of these ESTs could represent new transcripts, we constructed a cDNA microarray. This platform was used to hybridize against 12 different normal or tumor tissues. We identified 3421 transcribed regions not associated with annotated transcripts, representing 83.3% of the platform. The total number of differentially expressed sequences was 1007. Also, 28% of analyzed sequences could represent noncoding RNAs. Our data reinforces the knowledge of the human genome being pervasively transcribed, and point out molecular marker candidates for different cancers. To reinforce our data, we confirmed, by real-time PCR, the differential expression of three out of eight potentially tumor markers in prostate tissues. Lists of 1007 differentially expressed sequences, and the 291 potentially noncoding tumor markers were provided

    No-match ORESTES explored as tumor markers

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    Sequencing technologies and new bioinformatics tools have led to the complete sequencing of various genomes. However, information regarding the human transcriptome and its annotation is yet to be completed. The Human Cancer Genome Project, using ORESTES (open reading frame EST sequences) methodology, contributed to this objective by generating data from about 1.2 million expressed sequence tags. Approximately 30% of these sequences did not align to ESTs in the public databases and were considered no-match ORESTES. On the basis that a set of these ESTs could represent new transcripts, we constructed a cDNA microarray. This platform was used to hybridize against 12 different normal or tumor tissues. We identified 3421 transcribed regions not associated with annotated transcripts, representing 83.3% of the platform. The total number of differentially expressed sequences was 1007. Also, 28% of analyzed sequences could represent noncoding RNAs. Our data reinforces the knowledge of the human genome being pervasively transcribed, and point out molecular marker candidates for different cancers. To reinforce our data, we confirmed, by real-time PCR, the differential expression of three out of eight potentially tumor markers in prostate tissues. Lists of 1007 differentially expressed sequences, and the 291 potentially noncoding tumor markers were provided

    Hereditary breast and ovarian cancer: assessment of \ud point mutations and copy number variations in \ud Brazilian patients

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    Background \ud Germ line mutations in BRCA1 and BRCA2 (BRCA1/2) and other susceptibility genes have \ud been identified as genetic causes of hereditary breast and ovarian cancer (HBOC). To identify \ud the disease-causing mutations in a cohort of 120 Brazilian women fulfilling criteria for \ud HBOC, we carried out a comprehensive screening of BRCA1/2, TP53 R337H, CHEK2 \ud 1100delC, followed by an analysis of copy number variations in 14 additional breast cancer \ud susceptibility genes (PTEN, ATM, NBN, RAD50, RAD51, BRIP1, PALB2, MLH1, MSH2, \ud MSH6, TP53, CDKN2A, CDH1 and CTNNB1). \ud Methods \ud Capillary sequencing and multiplex ligation-dependent probe amplification (MLPA) were \ud used for detecting point mutations and copy number variations (CNVs), respectively, for the \ud BRCA1 and BRCA2 genes; capillary sequencing was used for point mutation for both variants \ud TP53 R337H and CHEK2 1100delC, and finally array comparative genomic hybridization \ud (array-CGH) was used for identifying CNVs in the 14 additional genes. \ud Results \ud The positive detection rate in our series was 26%. BRCA1 pathogenic mutations were found \ud in 20 cases, including two cases with CNVs, whereas BRCA2 mutations were found in 7 \ud cases. We also found three patients with the TP53 R337H mutation and one patient with the \ud CHEK2 1100delC mutation. Seven (25%) pathogenic mutations in BRCA1/2 were firstly \ud described, including a splice-site BRCA1 mutation for which pathogenicity was confirmed by \ud the presence of an aberrant transcript showing the loss of the last 62 bp of exon 7. \ud Microdeletions of exon 4 in ATM and exon 2 in PTEN were identified in BRCA2-mutated and \ud BRCA1/2-negative patients, respectively. \ud Conclusions \ud In summary, our results showed a high frequency of BRCA1/2 mutations and a higher \ud prevalence of BRCA1 (64.5%) gene. Moreover, the detection of the TP53 R337H variant in \ud our series and the fact that this variant has a founder effect in our population prompted us to \ud suggest that all female breast cancer patients with clinical criteria for HBOC and negative for \ud BRCA1/2 genes should be tested for the TP53 R337H variant. Furthermore, the presence of \ud genomic structural rearrangement resulting in CNVs in other genes that predispose breast \ud cancer in conjunction with BRCA2 point mutations demonstrated a highly complex genetic \ud etiology in Brazilian breast cancer families.Fundação de Amparo à Pesquisa do Estado de São Paulo (2008/57887- 9).Conselho Nacional de Desenvolvimento Científico e Tecnológico (408833/2006-8

    Transcriptional Alterations Related to Neuropathology and Clinical Manifestation of Alzheimer's Disease

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    Alzheimer's disease (AD) is the most common cause of dementia in the human population, characterized by a spectrum of neuropathological abnormalities that results in memory impairment and loss of other cognitive processes as well as the presence of non-cognitive symptoms. Transcriptomic analyses provide an important approach to elucidating the pathogenesis of complex diseases like AD, helping to figure out both pre-clinical markers to identify susceptible patients and the early pathogenic mechanisms to serve as therapeutic targets. This study provides the gene expression profile of postmortem brain tissue from subjects with clinic-pathological AD (Braak IV, V, or V and CERAD B or C; and CDR >= 1), preclinical AD (Braak IV, V, or VI and CERAD B or C; and CDR = 0), and healthy older individuals (Braak <= II and CERAD 0 or A; and CDR = 0) in order to establish genes related to both AD neuropathology and clinical emergence of dementia. Based on differential gene expression, hierarchical clustering and network analysis, genes involved in energy metabolism, oxidative stress, DNA damage/repair, senescence, and transcriptional regulation were implicated with the neuropathology of AD; a transcriptional profile related to clinical manifestation of AD could not be detected with reliability using differential gene expression analysis, although genes involved in synaptic plasticity, and cell cycle seems to have a role revealed by gene classifier. In conclusion, the present data suggest gene expression profile changes secondary to the development of AD-related pathology and some genes that appear to be related to the clinical manifestation of dementia in subjects with significant AD pathology, making necessary further investigations to better understand these transcriptional findings on the pathogenesis and clinical emergence of AD.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP) [2005/04151-7]Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP

    Li-Fraumeni-like syndrome associated with a large BRCA1 intragenic deletion

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    Background: Li-Fraumeni (LFS) and Li-Fraumeni-like (LFL) syndromes are associated to germline TP53 mutations, and are characterized by the development of central nervous system tumors, sarcomas, adrenocortical carcinomas, and other early-onset tumors. Due to the high frequency of breast cancer in LFS/LFL families, these syndromes clinically overlap with hereditary breast cancer (HBC). Germline point mutations in BRCA1, BRCA2, and TP53 genes are associated with high risk of breast cancer. Large rearrangements involving these genes are also implicated in the HBC phenotype. Methods: We have screened DNA copy number changes by MLPA on BRCA1, BRCA2, and TP53 genes in 23 breast cancer patients with a clinical diagnosis consistent with LFS/LFL; most of these families also met the clinical criteria for other HBC syndromes. Results: We found no DNA copy number alterations in the BRCA2 and TP53 genes, but we detected in one patient a 36.4 Kb BRCA1 microdeletion, confirmed and further mapped by array-CGH, encompassing exons 9-19. Breakpoints sequencing analysis suggests that this rearrangement was mediated by flanking Alu sequences. Conclusion: This is the first description of a germline intragenic BRCA1 deletion in a breast cancer patient with a family history consistent with both LFL and HBC syndromes. Our results show that large rearrangements in these known cancer predisposition genes occur, but are not a frequent cause of cancer susceptibility.Brazilian National Institute of Science and Technology in OncogenomicsBrazilian National Institute of Science and Technology in Oncogenomics [FAPESP 2008/57887-9, CNPq 573589/08-9]Fundo de Incentivo a Pesquisa (FIP)Fundo de Incentivo a Pesquisa (FIPE)Hospital de Clinicas de Porto Alegre [04-081, 09-115]Hospital de Clinicas de Porto AlegreFAPERGS (Brazil)FAPERGS, BrazilCAPES [Process: 2317/10-9]CAPE

    Germline DNA copy number variation in familial and early-onset breast cancer

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    Introduction: Genetic factors predisposing individuals to cancer remain elusive in the majority of patients with a familial or clinical history suggestive of hereditary breast cancer. Germline DNA copy number variation (CNV) has recently been implicated in predisposition to cancers such as neuroblastomas as well as prostate and colorectal cancer. We evaluated the role of germline CNVs in breast cancer susceptibility, in particular those with low population frequencies (rare CNVs), which are more likely to cause disease." Methods: Using whole-genome comparative genomic hybridization on microarrays, we screened a cohort of women fulfilling criteria for hereditary breast cancer who did not carry BRCA1/BRCA2 mutations. Results: The median numbers of total and rare CNVs per genome were not different between controls and patients. A total of 26 rare germline CNVs were identified in 68 cancer patients, however, a proportion that was significantly different (P = 0.0311) from the control group (23 rare CNVs in 100 individuals). Several of the genes affected by CNV in patients and controls had already been implicated in cancer. Conclusions: This study is the first to explore the contribution of germline CNVs to BRCA1/2-negative familial and early-onset breast cancer. The data suggest that rare CNVs may contribute to cancer predisposition in this small cohort of patients, and this trend needs to be confirmed in larger population samples.Brazilian National Institute of Science and Technology in Oncogenomics [FAPESP 2008/57887-9, CNPq 573589/08-9, FAPESP (2009/00898-1)]Brazilian National Institute of Science and Technology in Oncogenomic

    The Transcriptomic Portrait of Locally Advanced Breast Cancer and Its Prognostic Value in a Multi-Country Cohort of Latin American Patients

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    Purposes: Most molecular-based published studies on breast cancer do not adequately represent the unique and diverse genetic admixture of the Latin American population. Searching for similarities and differences in molecular pathways associated with these tumors and evaluating its impact on prognosis may help to select better therapeutic approaches. Patients and Methods: We collected clinical, pathological, and transcriptomic data of a multi-country Latin American cohort of 1,071 stage II-III breast cancer patients of the Molecular Profile of Breast Cancer Study (MPBCS) cohort. The 5-year prognostic ability of intrinsic (transcriptomic-based) PAM50 and immunohistochemical classifications, both at the cancer-specific (OSC) and disease-free survival (DFS) stages, was compared. Pathway analyses (GSEA, GSVA and MetaCore) were performed to explore differences among intrinsic subtypes. Results: PAM50 classification of the MPBCS cohort defined 42·6% of tumors as LumA, 21·3% as LumB, 13·3% as HER2E and 16·6% as Basal. Both OSC and DFS for LumA tumors were significantly better than for other subtypes, while Basal tumors had the worst prognosis. While the prognostic power of traditional subtypes calculated with hormone receptors (HR), HER2 and Ki67 determinations showed an acceptable performance, PAM50-derived risk of recurrence best discriminated low, intermediate and high-risk groups. Transcriptomic pathway analysis showed high proliferation (i.e. cell cycle control and DNA damage repair) associated with LumB, HER2E and Basal tumors, and a strong dependency on the estrogen pathway for LumA. Terms related to both innate and adaptive immune responses were seen predominantly upregulated in Basal tumors, and, to a lesser extent, in HER2E, with respect to LumA and B tumors. Conclusions: This is the first study that assesses molecular features at the transcriptomic level in a multicountry Latin American breast cancer patient cohort. Hormone-related and proliferation pathways that predominate in PAM50 and other breast cancer molecular classifications are also the main tumor-driving mechanisms in this cohort and have prognostic power. The immune-related features seen in the most aggressive subtypes may pave the way for therapeutic approaches not yet disseminated in Latin America. Clinical Trial Registration: ClinicalTrials.gov (Identifier: NCT02326857).Fil: Llera, Andrea Sabina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Abdelhay, Eliana Saul Furquim Werneck. Instituto Nacional de Cancer; BrasilFil: Artagaveytia, Nora. Universidad de la Republica; UruguayFil: Daneri Navarro, Adrián. Universidad de Guadalajara; MéxicoFil: Müller, Bettina. Instituto Nacional del Cáncer; ChileFil: Velazquez, Carlos. Universidad de Sonora; MéxicoFil: Alcoba, Elsa B.. Hospital Maria Curie; ArgentinaFil: Alonso, Isabel. Centro Hospitalario Pereira Rossell; UruguayFil: Alves Da Quinta, Daniela Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentina. Universidad Argentina de la Empresa; ArgentinaFil: Binato, Renata. Instituto Nacional de Cancer; BrasilFil: Bravo, Alicia Inés. Hospital Regional de Agudos Eva Perón; ArgentinaFil: Camejo, Natalia. Universidad de la Republica; UruguayFil: Carraro, Dirce Maria. Centro Internacional de Pesquisa; BrasilFil: Castro, Mónica. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Castro Cervantes, Juan M.. Umae Hospital de Especialidades Centro Medico Nacional Siglo XXI; MéxicoFil: Cataldi, Sandra. Instituto Nacional del Cáncer; UruguayFil: Cayota, Alfonso. Instituto Pasteur de Montevideo; UruguayFil: Cerda, Mauricio. Universidad de Chile; ChileFil: Colombo, Alicia. Universidad de Chile; ChileFil: Crocamo, Susanne. National Cancer Institute; Estados UnidosFil: Del Toro Arreola, Alicia. Universidad de Guadalajara; MéxicoFil: Delgadillo Cisterna, Raúl. Umae Hospital de Especialidades Centro Medico Nacional Siglo Xxi; MéxicoFil: Delgado, Lucía. Universidad de la Republica; UruguayFil: Fernandez, Elmer Andres. Area de Cs. Agrarias, Ingeniería, Cs. Biológicas y de la Salud de la Universidad Catollica de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Fejerman, Laura. University of California at Davis; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Trinchero, Alejandra. Hospital Regional de Agudos Eva Perón; ArgentinaFil: Valenzuela, Olivia. Universidad de Sonora; MéxicoFil: Vedham, Vidya. National Cancer Institute; Estados UnidosFil: Zagame, Livia. Instituto Jalisciense de Cancerología; MéxicoFil: Podhajcer, Osvaldo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentin
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