Dual Effect of Prolactin on Protein Kinase C Activity in CHO Cells Expressing Functional Prolactin Receptors

Abstract

We investigated the effects of prolactin (PRL) on the protein kinase C (PKC) activity in Chinese hamster ovary (CHO-E32) cells stably transfected with rabbit mammary gland PRL receptor cDNA. These cells express a functional long form of PRL-R. A 10-min to 2-hour treatment with 5 nM PRL resulted in the translocation of PKC activity from the cytosol to the membrane. Longer treatment ( 10-24 h) with the same concentration of PRL decreased the PKC activity in both particulate and cytoplasmic fractions. The PRL effect was dose dependent: maximal action was obtained with 1-10 nM. The PRLinduced activation of PKC was blocked by 20 nM staurosporine, a PKC inhibitor. Two inhibitors of tyrosine kinase, herbimycin A (1.75 μM) and genistein (100 μM). had no effect on PRL-induced activation of PKC.

Role of Protein Kinases in the Prolactin‐Induced Intracellular Calcium Rise in Chinese Hamster Ovary Cells Expressing the Prolactin Receptor

“…Although PMA‐induced PKC activation was able to increase [Ca 2+ ] i , PKC inhibition, due to long‐term treatment with PMA or various pharmacological agents, did not reduce the prolactin‐induced Ca 2+ influx. We have shown ( 19) that, in CHO cells, prolactin‐induced PKC activation was maximal after 2 h. As previously shown ( 16), the effects of prolactin on [Ca 2+ ] i were observed within 2–3 min. Other arguments against the involvement of PKC in prolactin‐induced Ca 2+ influx are that prolactin response, but not PMA or DiC8 response, is blocked by Nifedipine or KN‐62 treatment.…”

“…This influx is probably due to Ca 2+ ‐induced Ca 2+ release ( 16). In addition, we confirmed, in our cell model, that a short treatment (2 h) with a low concentration (5 n M ) of prolactin resulted in the translocation of protein kinase C (PKC) activity from the cytosol to the membrane ( 19, 20).…”

“…We have recently shown that prolactin has a time‐and dose‐dependent effect on PKC activity ( 19) and [Ca 2+ ] i ( 16, 18, 25) in CHO cells.…”

“…Agreement has now been reached concerning several steps in the transduction pathway, including receptor dimerization ( 38), JAK2 activation ( 8), and cell protein or lipid phosphorylations. The first phosphorylation reactions to be described concerned serine‐threonine phosphorylation via PKC activation ( 5, 19, 39) or via the G‐protein/PKA pathway ( 40–42). More recently, tyrosine phosphorylation ( 2) of several proteins (prolactin receptor itself, JAK2 autophosphorylation, signal transducers and activators of transcription proteins, and mitogen‐activated protein kinase) and lipid phosphorylation ( 35, 43) were associated with prolactin receptor activation ( 44, 45).…”