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Role of Protein Kinases in the Prolactin‐Induced Intracellular Calcium Rise in Chinese Hamster Ovary Cells Expressing the Prolactin Receptor
Author(s) -
Sorin B.,
Vacher A. M.,
Djiane J.,
Vacher P.
Publication year - 2000
Publication title -
journal of neuroendocrinology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.062
H-Index - 116
eISSN - 1365-2826
pISSN - 0953-8194
DOI - 10.1046/j.1365-2826.2000.00546.x
Subject(s) - protein kinase c , chinese hamster ovary cell , prolactin , medicine , endocrinology , protein kinase a , biology , signal transduction , kinase , tyrosine kinase , prolactin receptor , microbiology and biotechnology , chemistry , receptor , biochemistry , hormone
There is still only limited understanding of the early steps of prolactin signal transduction in target cells. It has been shown that prolactin actions are associated with cell protein phosphorylation, Ca 2+ increases, and so on. However, the link between the activation of kinases and calcium influx or intracellular Ca 2+ mobilization has not yet been clearly established. Chinese hamster ovary (CHO) cells, stably transfected with the long form of rabbit mammary gland prolactin receptor (PRL‐R) cDNA were used for PRL‐R signal transduction studies. Spectrofluorimetric techniques were used to measure intracellular calcium ([Ca 2+ ] i ) in cell populations with Indo1 as a calcium fluorescent probe. We demonstrate that, although protein kinase C activation (PMA or DiC8) caused a calcium influx in CHO cells, prolactin‐induced PKC activation was not responsible for the early effect of prolactin on [Ca 2+ ] i . Activation of protein kinase A (PKA) or protein kinase G did not modify [Ca 2+ ] i and inhibition of PKA pathway did not affect the prolactin response. In the same way, phosphatidylinositol‐3 kinaseinhibition had no effect on the prolactin‐induced Ca 2+ increase. On the other hand, tyrosine kinase inhibitors (herbimycin A, lavendustin A, and genistein) completely blocked the effect of prolactin on [Ca 2+ ] i (influx and release). W7, a calmodulin‐antagonist, and a specific inhibitor of calmodulin kinases (KN‐62), only blocked prolactin‐induced Ca 2+ influx but had no significant effect on Ca 2+ release. Using pharmacological agents, we present new data concerning the involvement of protein phosphorylations in the early effects of prolactin on ionic channels in CHO cells expressing the long form of PRL‐R. Our results suggest that, at least in the very early steps of prolactin signal transduction, serine‐threonine phosphorylation does not participate in the prolactin‐induced calcium increase. On the other hand, tyrosine phosphorylation is a crucial, very early step, since it controls K + channel activation, calcium influx, and intracellular calcium mobilization. Calmodulin acts later, since its inhibition only blocks the prolactin‐induced Ca 2+ influx.