Protein Kinase C Cascade Research Articles

Secreted form of β-amyloid precursor protein activates protein kinase C and phospholipase Cγ1 in cultured embryonic rat neocortical cells

The secreted form of β-amyloid precursor protein (sAPP) has been reported to exert various biological activities in cultured neurons. The signal transduction mechanisms underlying these physiological functions of sAPP remain unclear. We now report that treatment of neural cells with the secreted form of APP695 (sAPP695) leads to dose- and time-dependent increase in phosphorylation of the endogenous substrates with a molecular mass of 80, 57 and 43 kDa. Pretreatment of cells with protein kinase C (PKC) inhibitor H-7 reduced phosphorylation of the 80- and 43-kDa proteins in a dose-dependent manner. The effect of sAPP695 on the phosphorylation is mimicked by phorbol 12-myristate-13-acetate (PMA). Downregulation of PKC by prolonged treatment of cells with PMA abolished sAPP695-enhanced phosphorylation of the 80- and 43-kDa proteins, indicating PKC is involved in the sAPP695-enhanced phosphorylation of these proteins in the cells. We also suggest that the 80- and 43-kDa proteins phosphorylated by sAPP695-stimulation are the major PKC substrates myristoylated alanine-rich C-kinase substrate and growth-associated protein-43. Furthermore, we demonstrate that tyrosine phosphorylation of phospholipase Cγ1 and formation of inositol 1,4,5-trisphosphate were increased by sAPP695-stimulation. These observations suggest that sAPP695 induces the activation of the signaling pathways through a stimulation of phosphoinositide–PKC cascade.

Modulation of a cAMP/Protein Kinase A Cascade by Protein Kinase C in Sensory Neurons ofAplysia

The synaptic connections between the sensory neurons of Aplysia and their follower neurons have been used as a model system for examining the cellular mechanisms contributing to neuronal and synaptic plasticity. Recent studies suggest that at least two protein kinases, protein kinase A (PKA) and protein kinase C (PKC), contribute to serotonin (5-HT)-induced short-term facilitation. The interaction between these two kinase cascades has not been examined, however. Using electrophysiological and biochemical approaches, we examined possible interactions between PKA and PKC cascades. The results indicated that prolonged activation of PKC by preincubation with phorbol esters attenuated PKA-mediated actions of 5-HT, including increases in sensory neuron excitability and spike broadening in the presence of tetraethylammonium (TEA) and nifedipine. Although phorbol esters also attenuated increases in excitability by an analog of cAMP and small cardioactive peptide B (SCPB), the degree of attenuation was smaller. In addition, phorbol esters did not attenuate broadening of TEA spikes by the cAMP analog and SCPB. Thus, phorbol esters appeared specifically to attenuate aspects of the 5-HT activation of the cAMP/PKA cascade. Measurements of cAMP levels with radioimmunoassays revealed that phorbol esters did not attenuate 5-HT-induced cAMP synthesis, however. Finally, the results indicated that phorbol esters themselves induced a small but significant increase in excitability as well as an increase in the level of cAMP. Our results suggest that there is crosstalk between the PKC and PKA cascades. The mechanisms by which phorbol esters specifically attenuate 5-HT-induced activation of the cAMP/PKA cascade are not known, however.

Human T-cell lymphotropic virus type 1 Tax1 activation of NF-kappa B: involvement of the protein kinase C pathway

Human T-cell lymphotropic virus type 1 Tax1 induces the activation and nuclear localization of the cellular transcription factor, NF-kappa B. Treatment of cells with calphostin C, a protein kinase C (PKC) inhibitor, blocked induction of NF-kappa B DNA binding activity in human T-cell lymphotropic virus type 1-transformed C81 cells and Tax1-stimulated murine pre-B cells, suggesting that PKC was an important intermediate in the NF-kappa B induction pathway. We further demonstrate that Tax1 associates with, and activates, PKC. PKC was coimmunoprecipitated with anti- Tax1 sera from Tax1-expressing MT4 extracts and Jurkat extracts in the presence of exogenous Tax1 protein. In addition, the glutathione-S-transferase-Tax1 protein bound specifically to the alpha, delta, and eta PKC isoenzymes synthesized in rabbit reticulocyte lysates. The addition of Tax1 to in vitro kinase reaction mixtures leads to the phosphorylation of Tax1 and an 18-fold increase in the autophosphorylation of PKC. Transfection of Jurkat cells with wild-type Tax1 stimulated membrane translocation of PKC. In contrast, Tax1 mutant M22, which fails to stimulate NF-kappa B-dependent transcription, failed to stimulate membrane translocation of PKC. Tax1 did not directly increase PKC phosphorylation of I kappa B alpha. Our results are consistent with a model in which Tax1 interacts with PKC and stimulates membrane translocation and triggering of the PKC pathway. Subsequent steps in the PKC cascade likely stimulate phosphorylation of I kappa B alpha.

Results of the use of vitamin A and retionoids in cutaneous malignancies

Pharmac. Ther. Vol. 40, No. I, pp. 107-122, 1989 Printed in Great Britain. All rights reserved Copyright © 1988 Pergamon Press pie Guest Editor: R. M. MACKIE RESULTS OF THE USE OF VITAMIN A AND RETINOIDS IN CUTANEOUS MALIGNANCIES S. M. LIPPMAN and F. L. MEYSKENS, JR Department of Internal Medicine and Arizona Cancer Center, University o f Arizona, Tucson, Arizona, U.S.A. INTRODUCTION Vitamin A was identified long ago as an essential agent in the normal differentiation and maturation of epithelial tissues (Moore, 1967; Wolbach and Howe, 1925; Wolf, 1984). Laboratory research has demonstrated clearly that vitamin A deficiency enhances the effect of chemical carcinogens and tumor promoters and that replacing vitamin A reverses neoplastic abnormalities in these tissues (Lippman et aL, 1987a; Lotan, 1980; Lotan and Nicolson, 1977; Moon et al., 1983). Clinical study has documented that many preneoplastic and neoplastic disorders respond to vitamin A and its natural and synthetic derivatives (retinoids) (Lippman et aL, 1987b). This chapter will present a brief review of the data from in vitro and animal skin carcinogenesis model studies that unequivocally demonstrate the anticancer activity of retinoids and a detailed discussion of the growing data from human studies employing retinoids to treat cutaneous neoplastic and preneoplastic processes. MECHANISMS A vast amount of laboratory data has accumulated on the potent antipromoter, antiproliferative and differentiation-inducing effects of vitamin A derivatives on many malignant cell types (Lippman et al., 1987a; Lotan, 1980; Moore, 1967; Wolbach and Howe, 1925; Wolf, 1984). Despite extensive investigation, the mechanism(s) of action which causes the diverse cellular changes modulated by retinoids remains incompletely understood. Recently, however, it has become possible to hypothesize a basic mechanism underlying these other actions and to present a unifying picture of retinoid effects. This involves the protein kinase C/phosphoinositol cascade system. Phorbol esters have been identified as primary promoters of carcinogenesis. Therefore, protein kinase C (PK-C), the phorbol ester receptor, plays a critical role in the promotion phase of the carcinogenic process (Nishizuka, 1986). Recently, PK-C has been implicated in the mediation of many phorbol ester-promoted actions, such as ornithine decarboxylase induction and epidermal growth factor receptor down regulation (Jetten and Shirley, 1986). Membrane-bound (activated) PK-C may transmit signals to the nucleus to regulate gene (or oncogene) transcription by phosphorylating proteins and may act by nontranscriptional actions as well, such as by phosphorylating the EGF receptor. Recent work by Cope and others proposes the theory that retinoids modulate PK-C activity (Cope et al., 1986; Jetten and Shirley, 1986; Lockyer et al., 1984). Cope et al. (1986) have shown through their study of mouse brain PK-C that, although unbound retinoid binding proteins (Apo-cRABP and Apo-cRBP) are substrates for PK-C, the holo-forms (e.g. retinoic acid-cRABP) inhibit PK-C activity. Although the molecular details of retinoids' inhibition of the PK-C cascade are unclear, they do not appear to block phorbol ester binding to PK-C. Cope (1986) also showed that the regulation of PK-C alters phosphorylation of the retinoid binding proteins and other specific cytosolic and