RESUMO
The purpose of the study was to develop a test of the multifunctional protein YB-1 in the intraoperative biopsy specimen to predict the course of breast cancer (BC). Its tasks were to use of real-time reverse-transcription polymerase chain reaction (RT-RCR) to substantiate the data previously obtained by semiquantitative RT-PCR and to clarify whether there was a correlation between the amount of YB-1 mRNA in the BC tissue and the status of steroid hormone receptors of these tumors. The determination of the tumor amount of YB-1 mRNA was shown to predict the course of BC: a statistically significant correlation was found between the higher content of YB-1 mRNA and the aggressive course of BC--the emergence of distant metastases. Comparing the content of YB-1 mRNA and the hormonal status of a tumor (the number of estrogen and progesterone receptors) revealed no correlations. The findings indicate that the determination of YB-1 mRNA by both real-time RT-PCR and semiquantitative RT-PCR may be used to predict BC metastases in distant organs.
Assuntos
Biomarcadores Tumorais/análise , Neoplasias da Mama/química , Neoplasias da Mama/diagnóstico , Proteínas de Ligação a DNA/análise , Proteínas Nucleares/análise , RNA Mensageiro/análise , Biomarcadores Tumorais/genética , Neoplasias da Mama/patologia , Proteínas de Ligação a DNA/genética , Feminino , Humanos , Pessoa de Meia-Idade , Metástase Neoplásica , Proteínas Nucleares/genética , Prognóstico , Proteína 1 de Ligação a Y-BoxRESUMO
The multifunctional mammalian protein YB-1 is involved in multiple DNA- and mRNA-dependent events in the cell and regulates gene expression at different levels. The intracellular localization and relative mRNA content of YB-1 in the breast tumors were studied. The presence of cells with nuclear YB-1 localization in the tumor cell population is a poor predictor that correlates with larger tumors (more than 5 cm). The high YB-1 mRNA content in the breast tumors promotes metastasis of small neoplasms and patients with breast cancer who have high tumor tissue YB-1 mRNA levels may referred to as an early distant metastasis risk group. The findings suggest that combined determination of YB-1 intercellular localization and mRNA levels can ensure a more reliable prognosis of breast cancer.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas Nucleares/metabolismo , RNA Mensageiro/metabolismo , Neoplasias da Mama/patologia , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Citoplasma/metabolismo , Citoplasma/patologia , Feminino , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína 1 de Ligação a Y-BoxRESUMO
The effects of YB-1 gene on the expression level of P-glycoprotein and drug resistance of tumor cells were studied in cultured HCT116 colon cancer cells. Transitory transfection of chimeric YB-1/GFP gene rendered HCT116 cells a selective advantage in a medium with vinblastine, which caused translocation of the chimeric protein into cell nuclei. This was paralleled by an increase in the expression of P-glycoprotein (multiple drug resistance protein).
Assuntos
Resistência a Múltiplos Medicamentos/genética , Resistencia a Medicamentos Antineoplásicos/genética , Proteína 1 de Ligação a Y-Box/fisiologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Microscopia de Fluorescência , Transporte Proteico/efeitos dos fármacos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Vimblastina/farmacologia , Proteína 1 de Ligação a Y-Box/genética , Proteína 1 de Ligação a Y-Box/metabolismoRESUMO
Expression of mRNA for the mdr1 gene, cytochrome P450 isoforms 1A1 and 1B1, Ah receptor, and ARNT protein regulating the concentration of cytochrome P450 mRNA was compared in normal and spontaneously transformed mesothelial cells and mesothelioma cells from rats. Expression of cytochrome P450 1A1 and 1B1 mRNA decreased in transformed mesothelial and mesothelioma cells compared to normal mesothelial cells. mRNA for the mdr1 gene was undetected in normal mesothelial cells. Expression of mRNA for the Ah receptor and ARNT protein did not differ in cultured cells.
Assuntos
Hidrocarboneto de Aril Hidroxilases/genética , Citocromo P-450 CYP1A1/genética , Epitélio/enzimologia , Mesotelioma/enzimologia , RNA Mensageiro/genética , Animais , Sequência de Bases , Células Cultivadas , Citocromo P-450 CYP1B1 , Primers do DNA , Epitélio/patologia , Inativação Metabólica , Mesotelioma/patologia , RatosRESUMO
The multifunctional mammalian protein YB-1 is a member of the large DNA- and RNA-binding protein family with an evolutionarily ancient cold-shock domain. YB-1 is involved in multiple DNA- and mRNA-dependent events and regulates gene expression at various levels. It can be found both in the nucleus and the cytoplasm. Bound to DNA in the cell nucleus, YB-1 functions as a transcription factor interacting with inverted CCAAT-box (Y-box) in promoters and enhancers of multiple genes. In particular, YB-1 regulates activity of the multidrug resistance (MDR) genes MDR1 and LRP. In tumors, YB-1 has been suggested to be an early and global marker of MDR. In this study, we compared amounts of YB-1 mRNAs and intracellular localization of YB-1 protein in six pairs of drug sensitive and drug resistant sublines of diverse tumors. We have shown that neither great increase in the level of YB-1 mRNA nor substantial increase in the number of cells with nuclear localization of YB-1 are obligatory traits of drug resistant tumor cell populations. However, the cells with highest amounts of YB-1 mRNA also demonstrated increased quantities of MDR1, MRP1, BCRP, and LRP mRNAs encoding different MDR proteins. Transfection of two different populations of drug-sensitive cells with YB-1 cDNA led to increase in the amount of YB-1 mRNA. The quantities of MRP1 and LRP mRNAs increased in both populations. Introduction of YB-1 small hairpin RNA (shRNA) resulted in decreased amounts of YB-1 mRNA, as well as MRP1, LRP, and MDR1 mRNAs (in three different cell lines). Our data suggest that although YB-1 regulates several MDR genes, it could not be regarded as a global marker of already formed drug resistant tumor cell populations. It is most likely that at the first steps of MDR development YB-1 activity is necessary for propagation of resistant cell populations rather than for maintenance of drug resistance.
Assuntos
Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Líquido Intracelular/metabolismo , Proteínas de Neoplasias/metabolismo , Sequência de Aminoácidos , Linhagem Celular Tumoral , DNA Complementar/genética , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/genética , Células HCT116 , Humanos , Líquido Intracelular/química , Células K562 , Células KB , Dados de Sequência Molecular , Proteínas Associadas à Resistência a Múltiplos Medicamentos/biossíntese , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/química , Proteínas de Neoplasias/genética , Proteínas Nucleares , Interferência de RNA , RNA Polimerase III/genética , RNA Mensageiro/metabolismo , RNA Viral/genética , Partículas de Ribonucleoproteínas em Forma de Abóbada/biossíntese , Partículas de Ribonucleoproteínas em Forma de Abóbada/genética , Proteína 1 de Ligação a Y-BoxRESUMO
We have studied the effect of polycyclic aromatic hydrocarbons (PAH) on gap junction intercellular communications (GJIC) in culture of hepatoma cells Hep G2 and G27. Carcinogenic PAH inhibited GJIC in both cultures in contrast to non-carcinogenic PAH. We showed that both constitutive and inducible expressions of mRNAs of Ah receptor and cytochrome P4501A1 (the main isoform involved in PAH metabolism) were absent in hepatoma G27 cells. We concluded that the initial, non-metabolized molecules of carcinogenic PAH are responsible for changes in GJIC through interaction with an unknown factor in the cellular membrane.
Assuntos
Comunicação Celular/efeitos dos fármacos , Junções Comunicantes/efeitos dos fármacos , Hidrocarbonetos Policíclicos Aromáticos/farmacologia , Sequência de Bases , Benzo(a)pireno/metabolismo , Benzo(a)pireno/farmacologia , Linhagem Celular Tumoral , Células Cultivadas , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Humanos , Dados de Sequência Molecular , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Relação Quantitativa Estrutura-Atividade , RNA Mensageiro/metabolismo , Receptores de Hidrocarboneto Arílico/genética , Receptores de Hidrocarboneto Arílico/metabolismoRESUMO
We studied comparative expression and activity of cytochrome P450 family 1 (CYP1) isoforms in rat embryo cells, both primary and immortalized by Rausher leukemia virus (RLV). In RLV-infected embryonal cells compared with the initial ones the expression levels of CYP1A1 and 1B1 mRNAs and benzo[a]pyrene (BP) hydroxylase activity were higher, regardless of their treatment with the CYP1 inducer 2,3,7,8-tetrachlorodibenzo-p-dioxin. The sensitivity to BP and 7,12-dimethylbenzo[a]anthracene was higher in the cells immortalized with RLV. The expression level of mRNAs of induction-mediating proteins aryl hydrocarbon receptor and aryl hydrocarbon receptor nuclear translocator was the same in both cell cultures tested. Higher sensitivity of cells immortalized with RLV compared with the initial embryo cells to transforming effect of BP, which was described previously, is possibly associated with elevated expression of CYP1 isoforms.
