RESUMO
Ruk/CIN85 is a receptor-proximal 'signalling' adaptor that possesses three SH3 domains, Pro- and Ser-rich regions and C-terminal coiled-coil domain. It employs distinct domains and motifs to act as a transducer platform in intracellular signaling. Based on cDNA analysis, various isoforms of Ruk/CIN85 with different combination of protein-protein interaction domains as well as additional Ruk/CIN85 forms that are the products of post-translational modifications have been demonstrated. Nevertheless, there is no precise information regarding both the subcellular distribution and the role of Ruk/CIN85 multiple molecular forms in cellular responses. Using MCF-7 human breast adenocarcinoma cells and cell fractionation technique, specific association of Ruk/CIN85 molecular forms with different subcellular compartments was demonstrated. Induction of apoptosis of MCF-7 cells by doxorubicin treatment or by serum deprivation resulted in the system changes of Ruk/CIN85 molecular forms intracellular localization as well as their ratio. The data obtained provide a new insight into potential physiological significance of Ruk/CIN85 molecular forms in the regulation of various cellular functions.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Regulação Neoplásica da Expressão Gênica , Organelas/química , Processamento de Proteína Pós-Traducional , Proteínas Adaptadoras de Transdução de Sinal/genética , Antibióticos Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Compartimento Celular , Fracionamento Celular , Fragmentação do DNA , Doxorrubicina/farmacologia , Humanos , Células MCF-7 , Organelas/efeitos dos fármacos , Organelas/metabolismo , Domínios e Motivos de Interação entre Proteínas , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transdução de SinaisRESUMO
The values of endoplasmic reticulum Ca(2+)-pool and store-operated Ca2+ entry (SOCE) were estimated in rat thymocytes and Jurkat cells loaded with indo-1 and treated with thapsigargin. It was shown that the relative value of SOCE in thymocytes was substantially lower than in Jurkat cells. Significant increase of SOCE in Jurkat cells preincubated with 10(-5) M C60 and exposed to uv/visible light irradiation was detected at 1-3 h after exposure. At this time FCCP-induced Ca(2+)-release from mitochondria was shown to be reduced, while cytochrome c level into the cytoplasm of Jurkat cells, detected by Western blot analysis, to be increased. It is supposed that Ca2+ flux remodulation induced by photoexcited fullerene C60 in Jurkat cells might be involved in the initiation of signalling events leading to cell apoptosis.
Assuntos
Canais de Cálcio/metabolismo , Cálcio/metabolismo , Citocromos c/metabolismo , Fulerenos/farmacologia , Mitocôndrias/efeitos dos fármacos , Fármacos Fotossensibilizantes/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Carbonil Cianeto p-Trifluormetoxifenil Hidrazona/farmacologia , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/efeitos da radiação , Inibidores Enzimáticos/farmacologia , Corantes Fluorescentes , Humanos , Indóis , Células Jurkat , Luz , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Especificidade de Órgãos , Ionóforos de Próton/farmacologia , Ratos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/efeitos da radiação , Tapsigargina/farmacologia , Timócitos/citologia , Timócitos/efeitos dos fármacos , Timócitos/metabolismo , Timócitos/efeitos da radiaçãoRESUMO
The viability of normal (Wistar rat thymocytes) and transformed (human leukemia Jurkat cells) T cells after UV/Vis irradiation in the presence of pristine C60 fullerene was studied. The data obtained have shown that C60 fullerene exhibits cytotoxic effect against transformed T lymphocytes when combined with UV/Vis irradiation using mercury-vapor lamp (320-600 nm). C60 fullerene photocytotoxicity was not detected in thymocytes. C60-dependent photoinduced apoptosis of Jurkat cells was confirmed by DNA fragmentation and caspase-3 activation. No substantial increase of caspase-3 activation was observed in thymocytes treated with C60 fullerene plus irradiation, while antileukemic agent cytosine arabinoside was shown to induce caspase-3 activation both in Jurkat cells and thymocytes. The data obtained may be useful for development of photosensitizers for photodynamic therapy with selective action on leukemia cells.
