Navegando por Palavras-chave "Phosphoproteoma"

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    Modulação do eixo PLCγ1, Ca2+/CAMKII e PKC em células de leucemia mieloide crônica tratadas com violaceína
    (Universidade Federal de São Paulo (UNIFESP), 2020-07-30) Miura, Natalia Tamagusko [UNIFESP]; Justo, Giselle Zenker [UNIFESP]; Universidade Federal de São Paulo
    Violacein is an indole derivative isolated from Chromobacterium violaceum with potential antitumor activity and specific mechanisms of action in vitro and in vivo. This compound was evaluated in vitro in cultures of chronic myeloid leukemia (CML), which is a myeloproliferative disease associated with a genetic translocation that leads to the formation of the BCR-ABL oncoprotein. BCR-ABL is capable of inducing multiple signal transduction pathways responsible for the leukemogenic process. Moreover, BCR-ABL acts with additional mechanisms, dependent or independent of BCR-ABL, to induce resistance, leading to an unfavorable prognosis in the treatment of the disease and the search for alternative drug therapies. One of the factors that lead to the phenotype of multiple drug resistance (MDR) is related to the increase in the expression of efflux pumps, among them, the P glycoproteins (Pgp), which is one of the most studied mechanisms of chemoresistance in CML. Hence, we compared the phosphoproteomic profile of K562 cells and their variant that overexpresses the Pgp and, therefore, presents the MDR phenotype, the Lucena 1 cell line, using a microarray approach of peptides that allows to analyze the activity of different cell kinases. Data analysis showed that the exposure of K562 cells to violacein significantly modulated 13 kinases, whereas the activity of 6 kinases in Lucena 1 cells showed significant changes after treatment. The results showed a reduction in the activity of Axl and c-Met in K562 cells, while p38MAPK and JNK stimulation were observed in Lucena 1 cells. Of particular importance, the quinoma analysis indicated significant modulation in protein kinase C (PKC) activity after treatment of cells of both strains with violacein. In this sense, a reduction in phosphorylation of PKCα(Ser657) by immunoblotting was also observed. On the other hand, the pretreatment of cells with the specific classic PKC inhibitor, chelerythrine, inhibited about 80% of cell death induced by violacein, corroborating the increase in intracellular Ca2+ concentration. In addition, a significant increase in CAMKII(Thr286) activity was observed, suggesting that stimulation of the Ca2+/CAMKII axis and PKCs is important for cell death induced by violacein. In addition, the results also demonstrated the inhibition of PLCγ1(Tyr783), suggesting that this effect is also involved in inducing cell death. Together, the results indicate an important role for Ca2+ signaling and PLCγ1 inhibition in violacein-induced CML cell death. They also open perspectives for investigating new targets of action of this naturally derived compound.