RESUMO
T-kininogen (T-KG) is a precursor of T-kinin, the most abundant kinin in rat serum, and also acts as a strong and specific cysteine proteinase inhibitor. Its expression is strongly induced during aging in rats, and expression of T-KG in Balb/c 3T3 fibroblasts results in inhibition of cell proliferation. However, T-KG is a serum protein produced primarily in the liver, and thus, most cells are only exposed to the protein from the outside. To test the effect of T-KG on fibroblasts exposed to exogenous T-KG, we purified the protein from the serum of K-kininogen-deficient Katholiek rats. In contrast to the results obtained by transfection, exposure of Balb/c 3T3 fibroblasts to exogenously added T-KG leads to a dose-dependent increase in [3H]-thymidine incorporation. This response does not require kinin receptors, but it is clearly mediated by activation of the ERK pathway. As a control, we repeated the transfection experiments, using a different promoter. The results are consistent with our published data showing that, under these circumstances, T-KG inhibits cell proliferation. We conclude that T-KG exerts opposite effects on fibroblast proliferation, depending exclusively on the way that it is administered to the cells (transfection versus exogenous addition).
Assuntos
Proliferação de Células/efeitos dos fármacos , Inibidores de Cisteína Proteinase/farmacologia , Cininogênios/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Envelhecimento/metabolismo , Animais , Células 3T3 BALB , Inibidores de Cisteína Proteinase/genética , Inibidores de Cisteína Proteinase/metabolismo , Relação Dose-Resposta a Droga , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Cininogênios/genética , Cininogênios/metabolismo , Camundongos , Ratos , TransfecçãoRESUMO
Histones, the basic proteins which compact DNA into the nucleosomal and solenoidal fibers are synthesized in correlation with DNA replication during the S-phase of the cell cycle. This behavior is controlled both at transcriptional and postranscriptional levels in higher eukaryotes and yeasts. We have found that histone synthesis in synchronized trypanosomes is controlled by fluctuations on the levels of their mRNAs. Though we cannot preclude the existence of a transcriptional regulatory mechanism, our results point to the participation of changes in the stability of histone mRNAs as a regulatory mechanism of their levels during the cell cycle in Trypanosoma. We have also found a postranscriptional regulatory mechanism which could be acting at the translational level. These results show both similarities and differences between Trypanosoma and higher eukaryotes regarding the expression of their histone genes.
Assuntos
Ciclo Celular , Histonas/genética , Histonas/metabolismo , Trypanosoma cruzi/metabolismo , Animais , Northern Blotting , DNA/metabolismo , Biossíntese de Proteínas , RNA Mensageiro/metabolismo , Fase S , Transcrição GênicaRESUMO
Trypanosoma cruzi is an ancient, parasitic eukaryote which does not undergo chromatin condensation during cell division. This behavior may be explained if one considers the strong amino acid sequence divergence of Trypanosoma histones compared to higher eukaryotes. In the latter organisms histone synthesis is coupled to DNA replication. Considering the nonconserved amino acid sequence of T. cruzi histones, as well as the absence of chromatin condensation in this organism, we have studied histone synthesis in relation to DNA replication in this parasite. We have found that core histones and a fraction of histone H1 are synthesized concomitantly to DNA replication. However, another fraction of histone H1 is constitutively synthesized.