Drouin S M, Kiley S C, Carlino J A, Barnum S R
Department of Microbiology, University of Alabama at Birmingham, 35294, USA.
Mol Immunol. 1998 Jan;35(1):1-11. doi: 10.1016/s0161-5890(98)00014-5.
Previously, we reported that TGF-beta2 regulates the C3 gene expression in a dose- and time-dependent manner in monocytes. To extend these studies, we examined the role of PKC in the TGF-beta2-mediated induction of C3 expression by the human monocyte cell line, U937. Treatment of U937 cells with the PKC inhibitors, H7 and calphostin C, suppressed TGF-beta2-mediated induction of C3 protein levels, but not mRNA levels, in a dose-dependent manner. At the highest concentrations of H7 and calphostin C, C3 protein levels were inhibited 50% and 93%, respectively, compared to control levels. Treatment of U937 cells with HA1004, a weak PKC inhibitor used as a control for H7, did not inhibit induction of C3 protein levels. Down-modulating PKC with a prolonged exposure of U937 cells to PMA also suppressed TGF-beta2-mediated C3 protein induction by as much as 82%. Incubating cell extracts isolated from TGF-beta2-treated U937 cells with the PKC substrate, MIBP(4-14), resulted in increased substrate phosphorylation compared to cell extracts isolated from untreated cells. Addition of calphostin C suppressed the increased substrate phosphorylation by TGF-beta2. Furthermore, biosynthetic labeling of U937 cells treated with TGF-beta2 and calphostin C demonstrated an accumulation of C3 protein within cell lysates compared to controls. Collectively, these studies suggest a role for PKC in the secretion of C3 protein during TGF-beta2-mediated regulation of C3 expression in U937 cells.
此前，我们报道过转化生长因子-β2（TGF-β2）在单核细胞中以剂量和时间依赖性方式调节C3基因表达。为扩展这些研究，我们检测了蛋白激酶C（PKC）在人单核细胞系U937中TGF-β2介导的C3表达诱导过程中的作用。用PKC抑制剂H7和钙泊三醇C处理U937细胞，以剂量依赖性方式抑制了TGF-β2介导的C3蛋白水平的诱导，但不影响mRNA水平。在H7和钙泊三醇C的最高浓度下，与对照水平相比，C3蛋白水平分别被抑制了50%和93%。用HA1004（一种用作H7对照的弱PKC抑制剂）处理U937细胞，并未抑制C3蛋白水平的诱导。通过使U937细胞长时间暴露于佛波酯（PMA）来下调PKC，也可将TGF-β2介导的C3蛋白诱导抑制多达82%。将从TGF-β2处理的U937细胞中分离的细胞提取物与PKC底物MIBP(4 - 14)一起孵育，与从未处理细胞中分离的细胞提取物相比，底物磷酸化增加。添加钙泊三醇C可抑制TGF-β2引起的底物磷酸化增加。此外，对用TGF-β2和钙泊三醇C处理的U937细胞进行生物合成标记，结果表明与对照相比，细胞裂解物中C3蛋白有所积累。总体而言，这些研究表明PKC在U937细胞中TGF-β2介导的C3表达调节过程中C3蛋白分泌方面发挥作用。
