Calmodulin antagonists increase the amount of mRNA for the low-density-lipoprotein receptor in skin fibroblasts

Abstract

The effects of calmodulin antagonists on the amount of LDL receptor (LDL-R) mRNA in cultured human fibroblasts was examined by hybridization with a fragment of LDL-R cDNA. In a 'Northern' blot the fragment hybridized to a 5.3-kilobase RNA, as expected for LDL-R mRNA. The concentration of this RNA was increased in preparations from cells that were treated with trifluoperazine or W-7 [N-(6-aminohexyl)-5-chloronaphthalene-1-sulphonamide). The selectivity of the increase was established by using a probe for /3-actin mRNA. In dot-blot hybridization it was observed that the calmodulin antagonists cause 2-4-fold relative increase in the amount of LDL-R mRNA. INTRODUCTIONThe low-density-lipoprotein (LDL) receptor, a transmembrane cell-surface glycoprotein, mediates endocytosis of macromolecular lipoprotein ligands. The regulation of the activity of the LDL receptor has been studied extensively in both cultured cells and experimental animals. Although the primary structure (Yamamoto et al., 1984) and synthesis of the LDL receptor (Goldstein et al., 1985) and the organization of the gene encoding this glycoprotein (Suidhof et al., 1987) have been elucidated, little is known about the mechanisms of the regulation of the synthesis of the LDL receptor. In human skin fibroblasts, depletion of cholesterol enhances, and loading with cholesterol lowers, the activity of the LDL receptor . The activity of the LDL receptor Chao et al., 1979) and the amount of its mRNA (Ma et al., 1986a,b) are enhanced in livers of rats, hamsters and rabbits treated with mevinolin (Ma et al., 1986b) or with pharmacological doses of

Ca(2+)-channel blockers modulate expression of 3-hydroxy-3-methylglutaryl-coenzyme A reductase and low density lipoprotein receptor genes stimulated by platelet-derived growth factor.

“…Although the latter effect was channeled apparently through PKC, the former occurred independently of the enzyme. This agrees with the findings of Eckardt et al (17), who showed that synthesis of LDL-R is unaffected by the PKC agonist phorbol 12-myristate 13-acetate using skin fibroblasts; moreover, participation of Ca2+ in LDL-R gene expression was excluded by the following: modulation of extracellular concentration of Ca2+ was without an effect, and addition ofthe Ca2+-ionophore A23187 or the Ca2+-antagonist TMB-8, thought to block intracellular Ca2+ mobilization, failed to alter the rate of LDL-R synthesis. This suggests that the established effects of Ca2+-channel blockers-i.e., inhibition of Ca2+ influx via potentialoperated channels-may not be relevant to their stimulatory influence on the LDL-R gene.…”

Evidence for sterol-independent regulation of low-density lipoprotein receptor activity in Hep-G2 cells

“…The expression of LDL receptors by cells in culture is also affected by a variety of incubation conditions. Receptor activity is increased by incubation of cells with insulin [18][19][20], plateletderived growth factor (PDGF) [21][22][23][24], fibroblast-derived growth factor [21], thyroid hormone [20,25], oestrogen [26], Ca2l channel blockers [27][28][29] and calmodulin inhibitors [30][31], and by factors present in serum [32]. It is not clear whether the effects of these growth-related factors on LDL receptor activity in the various cell types are secondary to changes in cellular cholesterol content or whether they act as primary effectors.…”

Transcriptional activation of low density lipoprotein receptor gene by angiotensin-converting enzyme inhibitors and Ca(2+)-channel blockers involves protein kinase C isoforms.

“…Interestingly, the HMG-CoA reductase inhibitor mevinolin, known to lower elevated cholesterol level in plasma, stimulated the expression of LDLR gene (24), which led to an increase in binding and uptake of cholesterol ester. A similar effect was reported for Ca2+-channel blockers (25,26) corresponding to the recent observation that Ca2+-channel blockers are capable of slightly reducing the concentrations of cholesterol in plasma (27). Although the cholesterol-lowering effect of Ca2+-channel blockers in the plasma may only be marginal, the drugs may correct disturbances of cholesterol metabolism at the cellular level, as has been suggested (28).…”