Interleukin-3 inhibits cycloheximide induction of C-jun mRNA in human monocytes: possible role for a serine/threonine phosphatase

Cycloheximide is a strong inducer of the c-jun protooncogene mRNA at concentrations (< or = 50 ng/ml) that do not inhibit protein synthesis in human monocytes. This induction is transient lasting 30-60 min in contrast to the sustained induction obtained with concentrations that inhibit protein synthesis. The pluripotent colony stimulating factor interleukin-3 (IL-3) (10 ng/ml) is also a modest inducer of the c-jun gene in these cells; however, in combination with cycloheximide, IL-3 dramatically reduces the c-jun induction below levels induced by cycloheximide alone. This is a true inhibition and is not due to a change in temporal kinetics of induction because the suppression in the presence of IL-3 is observed at both 30 and 60 min after simultaneous addition of both IL-3 and cycloheximide. Preincubation of monocytes with 12.5 nM okadaic acid (a potent inhibitor of protein phosphatases 1 and 2A) and cycloheximide prior to addition of IL-3 restored the level of c-jun induction to that mediated by cycloheximide alone. This concentration of okadaic acid inhibited almost 70% of the phosphorylase phosphatase activity in monocyte lysates. These observations suggest that activation of protein serine/threonine phosphatase(s) underlies the ability of IL-3 to inhibit cycloheximide induction of c-jun in monocytes.     

The effect of cycloheximide on the regulation of proenkephalin and prodynorphin gene expressions induced by kainic acid in rat hippocampus

The effect of cycloheximide (CHX), a protein synthesis inhibitor, on the regulation of proenkephalin (proENK) and prodynorphin (proDYN) mRNA levels, proto-oncogenes, such as c-fos, 35-kDa fra and c-jun mRNA, and the levels of their products induced by kainic acid (KA) in rat hippocampus was studied. The proENK and proDYN mRNA levels were markedly increased 4 and 8 h after KA (10 mg/kg i.p.) administration. However, the intracellular proENK protein level was not affected by KA. The elevations of both proENK and proDYN mRNA levels induced by KA were inhibited by pre-administration of CHX (15 mg/kg i.p.). The increases of proENK and proDYN mRNA levels induced by KA were well-correlated with the increases of c-Fos, 35-kDa Fra and c-Jun protein levels. KA administration increased the hippocampal levels of c-Fos, 35-kDa Fra and c-Jun proteins with the time. The increases of c-Fos, 35-kDa Fra and c-Jun protein levels induced by KA administration were also inhibited by CHX pre-administration. KA administration markedly increased both c-fos and c-jun mRNA levels during 1 and 4 h and the increased levels of these proto-oncogene mRNA were further prolonged by the treatment with CHX. In addition, CHX alone increased both c-fos and c-jun mRNA levels although the onset times of induction were different. In electrophoretic mobility shift-assay, both AP-1 and ENKCRE-2 DNA-binding activities were increased by KA. KA-induced increases of AP-1 and ENKCRE-2 DNA-binding activities were also attenuated by CHX. In addition, KA-induced AP-1 and ENKCRE-2 DNA-binding activities were diminished by the antibodies against Fos and Jun family proteins. Furthermore, the cross-competition studies revealed that AP-1 proteins actively participated in ENKCRE-2 DNA domain. The results suggest that KA-induced proENK and proDYN mRNA expressions may require on-going synthesis of proteins, such as c-Fos, c-Jun and 35-kDa Fra, which may have a possible role in the up-regulation of proENK and proDYN gene expression through the binding with AP-1 and ENKCRE-2 DNA-binding motifs.     

Advantages of a two-chamber culture system to test drug nephrotoxicity: the example of cephaloridine

BACKGROUND: The adult prostate is maintained through an equilibrium between cell growth and death rates. Androgen deprivation induces an increase in intracellular Ca++, AP-1 gene expression of androgen-inducible genes. METHODS: Northern blot analysis, band-shift assays, and transient cotransfection assays were used to study the effects of Ca++ mobilizer A23187 on gene expression in human prostate cancer cells. RESULTS: A23187 repressed androgen-upregulated mRNAs for prostate-specific antigen (PSA) and hKLK2, and rapidly induced mRNA levels for c-fos and c-jun. AP-1 protein-DNA binding activities were elevated after A23187 treatments. Androgen receptor (AR)-mediated induction of chloramphenicol acetyltransferase (CAT) reporter was repressed by AP-1 proteins. CONCLUSIONS: The repression of AR-mediated induction of PSA and hKLK2 genes by Ca++ mobilizers is due to the interference of AR transactivation activity by AP-1 proteins.     

cAMP inhibits linoleic acid-induced growth by antagonizing p27(kip1) depletion, but not interfering with the extracellular signal-regulated kinase or AP-1 activities

To understand the underlying signaling events of polyunsaturated fatty acid-induced growth, we studied the effect of cAMP on early and delayed growth response events induced by linoleic acid in smooth muscle cells (SMC). cAMP significantly inhibited both basal and linoleic acid-induced DNA synthesis. Linoleic acid-induced early growth response events, such as activation of ERKs, induction of expression of c-fos and jun-B and stimulation of AP-1 activity, however, were not affected by cAMP. In contrast, linoleic acid-induced c-jun expression was blocked by cAMP. cAMP alone stimulated ERKs activation, c-fos and jun-B expression and increased AP-1 activity. Linoleic acid induced depletion of p27kip1 and increased CDK2 activity, events required for G1/S transition. In contrast to early growth response events, linoleic acid-induced G1/S transition signals were significantly inhibited by cAMP. These findings suggest that in addition to inducing immediate early growth response events, linoleic acid mimics growth factors in activating cell cycle events that are associated with G1/S transition in SMC and the negative regulation of linoleic acid-induced growth by cAMP is apparently due to its antagonism with linoleic acid-induced p27kip1 depletion and CDK2 activation.