Recurrent copy number gains of ACVR1 and corresponding transcript overexpression are associated with survival in head and neck squamous cell carcinomas

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Copy number of P elements, KP/full-sized P element ratio and their relationships with environmental factors in Brazilian Drosophila melanogaster populations

The P transposable element copy numbers and the KP/full-sized P element ratios were determined in eight Brazilian strains of Drosophila melanogaster. Strains from tropical regions showed lower overall P element copy numbers than did strains from temperate regions. Variable numbers of full-sized and defective elements were detected, but the full-sized P and KP elements were the predominant classes of elements in all strains. The full-sized P and KP element ratios were calculated and compared with latitude. The northernmost and southernmost Brazilian strains showed fewer full-sized elements than KP elements per genome, and the strains from less extreme latitudes had many more full-sized P than KP elements. However, no clinal variation was observed. Strains from different localities, previously classified as having P cytotype, displayed a higher or a lower proportion of KP elements than of full-sized P elements, as well as an equal number of the two element types, showing that the same phenotype may be produced by different underlying genomic components of the P-M system.

Characterization of hobo element copy number and integrity in Brasilian populations of Drosophila melanogaster

We have determined the copy number and the presence of full-size hobo transposable elements in eight Brasilian strains of Drosophila melanogaster. Genomic DNA was digested with AvaII and XhoI restriction enzymes, respectively, and probed with a 963 bp sequence of the hobo element. Variable numbers of full-sized and defective elements were detected in all strains. The range of the copy number was 22.13 +/- 4.52. Blots showed the presence of a 2.6 kb fragment, corresponding to the complete element, in all strains exception of one and the 1.0 kb sequence, correponding to the Th1 and Th2 repressor elements. There was neither association among copy numbers of hobo elements and latitude nor the mean annual temperatures in the original geographical region of each strain.

Copy number variation of individual cattle genomes using next-generation sequencing

Copy number variations (CNVs) affect a wide range of phenotypic traits; however, CNVs in or near segmental duplication regions are often intractable. Using a read depth approach based on next-generation sequencing, we examined genome-wide copy number differences among five taurine (three Angus, one Holstein, and one Hereford) and one indicine (Nelore) cattle. Within mapped chromosomal sequence, we identified 1265 CNV regions comprising similar to 55.6-Mbp sequence-476 of which (similar to 38%) have not previously been reported. We validated this sequence-based CNV call set with array comparative genomic hybridization (aCGH), quantitative PCR (qPCR), and fluorescent in situ hybridization (FISH), achieving a validation rate of 82% and a false positive rate of 8%. We further estimated absolute copy numbers for genomic segments and annotated genes in each individual. Surveys of the top 25 most variable genes revealed that the Nelore individual had the lowest copy numbers in 13 cases (similar to 52%, chi(2) test; P-value <0.05). In contrast, genes related to pathogen- and parasite-resistance, such as CATHL4 and ULBP17, were highly duplicated in the Nelore individual relative to the taurine cattle, while genes involved in lipid transport and metabolism, including APOL3 and FABP2, were highly duplicated in the beef breeds. These CNV regions also harbor genes like BPIFA2A (BSP30A) and WC1, suggesting that some CNVs may be associated with breed-specific differences in adaptation, health, and production traits. By providing the first individualized cattle CNV and segmental duplication maps and genome-wide gene copy number estimates, we enable future CNV studies into highly duplicated regions in the cattle genome.

Análise do gene TUBB3 e de Copy Number Variations em pacientes com Polimicrogiria

A polimicrogiria (PMG) é uma malformação do córtex cerebral causada por falhas no seu desenvolvimento, caracterizando-se por um número excessivo de pequenos giros e laminação anormal, dando à superfície cortical uma aparência irregular e grosseira. A gravidade de suas manifestações clínicas se relaciona diretamente com a extensão da malformação e das regiões cerebrais afetadas, sendo que a presença de lesões bilaterais ou unilaterais extensas indica um pior prognóstico. Uma das síndromes de polimicrogiria mais freqüentes e, conseqüentemente, mais bem descritas clinicamente, é a polimicrogiria perisylviana bilateral (PPB). Essa forma de PMG atinge a região que tange a fenda Sylviana, podendo apresentar-se tanto unilateralmente quanto em ambos os hemisférios. Vários genes têm sido relacionados a diferentes formas de polimicrogiria, são eles AFF2,TUBA1A, TUBB2B e TUBA8, SRPX2 e WDR62. Estes genes já foram estudados pelo nosso grupo de pesquisa em um grupo de pacientes compostos de casos familiares e esporádicos, acometidos em sua maioria pela forma perisylviana de PMG. Nenhuma variante deletéria foi identificada nestes genes. Recentemente um novo gene foi implicado na etiologia molecular das PMG, o TUBB3. O gene em questão pertence à mesma família de TUBA1A, TUBB2B e TUBA8 e codifica uma proteína de ligação aos microtúbulos, tendo importante papel na formação do fuso. Além deste gene, também tem sido descritas alterações genômicas, denominadas de Copy Number Variations (CNV), estas variações estruturais tem sido associadas com diversos distúrbios neurológicos, que vão desde transtornos psiquiátricos até malformações do córtex cerebral como a PMG. Desta forma, o objetivo deste trabalho foi analisar a existência de alterações de ponto deletérias no gene TUBB3 em pacientes com PMG e também, o envolvimento de CNVsna etiologia deste tipo de malformação ...

CYP1A2*1C, CYP2E1*5B, and GSTM1 polymorphisms are predictors of risk and poor outcome in head and neck squamous cell carcinoma patients

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evolutionary History of the PER3 Variable Number of Tandem Repeats (VNTR): Idiosyncratic Aspect of Primate Molecular Circadian Clock

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Organização genômima e análise da expressão do gene hnRNP Q-like (Heterogeneous nuclear ribonucleoprotein A-like) retroinserido no cromossomo B de Astatotilapia latifasciata

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Conditions affecting production of functional muscle recombinant alpha-tropomyosin in Saccharomyces cerevisiae

Yeasts are attractive hosts for heterologous protein production as they follow the general eukaryotic post-translational modification pattern. The well-known Saccharomyces cerevisiae has been used to produce a large variety of foreign proteins. The proper function of muscle tropomyosin depends on a specific modification at its N-terminus. Although tropomyosin has been produced in different expression systems, only the recombinant protein produced in the yeast Pichia pastoris has native-like functional properties. In this paper we describe the production of functional skeletal muscle tropomyosin in the yeast S. cerevisiae. The recombinant protein was produced in high amounts and production was strongly affected by genetic and environmental factors, including plasmid copy number, promoter strength, and growth media composition. (C) 2003 Elsevier B.V. (USA). All rights reserved.

Cloning and molecular characterization of Trypanosoma cruzi U2, U4, U5, and U6 small nuclear RNAs

Small nuclear RNAs (snRNAs) are important factors in the functioning of eukaryotic cells that form several small complexes with proteins; these ribonucleoprotein particles (U snRNPs) have an essential role in the pre-mRNA processing, particularly in splicing, catalyzed by spliceosomes, large RNA-protein complexes composed of various snRNPs. Even though they are well defined in mammals, snRNPs are still not totally characterized in certain trypanosomatids as Trypanosoma cruzi. For this reason we subjected snRNAs (U2, U4, U5, and U6) from T. cruzi epimastigotes to molecular characterization by polymerase chain reaction (PCR) and reverse transcription-PCR. These amplified sequences were cloned, sequenced, and compared with those other of trypanosomatids. Among these snRNAs, U5 was less conserved and U6 the most conserved. Their respective secondary structures were predicted and compared with known T. brucei structures. In addition, the copy number of each snRNA in the T. cruzi genome was characterized by Southern blotting.

Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

Human epidermal growth factor receptor 2 (HER2) has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC). HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH) in moderate immunoexpression (IHC 2+) cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH), which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH) and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer.

Evolutionary dynamics of heterochromatin in the genome of Dichotomius beetles based on chromosomal analysis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The evolutionary dynamics of the Helena retrotransposon revealed by sequenced Drosophila genomes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Comparative genomic hybridization analysis detects frequent over-representation of DNA sequences at 3q, 7p, and 8q in head and neck carcinomas

Comparative genomic hybridization (CGH) was used to identify chromosomal imbalances in 19 samples of squamous cell carcinoma of the head and neck (HNSCC). The chromosome arms most often or er-represented were 3q (48%), 8q (42%), and 7p (32%); in many cases, these changes were observed at high copy number. Other commonly over-represented sites were 1q, 2q, 6p, 6q, and 18q. The most frequently under-represented segments were 3p and 22q. Loss of heterozygosity of two polymorphic microsatellite loci from chromosome 22 was observed in two tongue tumors, in agreement with the CGH analysis. Gains of 1q and 2q material were detected in patients exhibiting a clinical history of recurrence and/or metastasis followed by terminal disease. This association suggests that gain of 1q and 2q map be a new marker of head and neck tumors with a refractory clinical response. (C) 2000 Elsevier B.V. All rights reserved.

Quantitative real-time PCR identifies a critical region of deletion on 22q13 related to prognosis in oral cancer

Quantitative real time PCR was performed on genomic DNA from 40 primary oral carcinomas and the normal adjacent tissues. The target genes ECGFB, DIA1, BIK, and PDGFB and the microsatellite markers D22S274 and D22S277, mapped on 22q13, were selected according to our previous loss of heterozygosity findings in head and neck tumors. Quantitative PCR relies on the comparison of the amount of product generated from a target gene and that generated from a disomic reference gene (GAPDH-housekeeping gene). Reactions have been performed with normal control in triplicates, using the 7700 Sequence Detection System (PE Applied Biosystems). Losses in the sequences D22S274 (22q13.31) and in the DIA1 (22q13.2-13.31) gene were detected in 10 out of 40 cases (25%) each. Statistically significant correlations were observed for patients with relative copy number loss of the marker D22S274 and stages T3-T4 of disease (P=0.025), family history of cancer (P = 0.001), and death (P = 0.021). Relative copy number loss involving the DIA1 gene was correlated to family history of cancer (P<0.001), death (P=0.002), and consumption of alcohol (P=0.026). Log-rank test revealed a significant decrease in survival (P=0.0018) for patients with DIA1 gene loss. Relative copy number losses detected in these sequences may be related to disease progression and a worse prognosis in patients with oral cancer.