Kim Jayoung, Lin Jianqing, Adam Rosalyn M, Lamb Carolyn, Shively Sharon Baughman, Freeman Michael R
The Urological Diseases Research Center, Department of Urology, Children's Hospital Boston, Massachusetts 02115, USA.
J Cell Biochem. 2005 Jan 1;94(1):39-49. doi: 10.1002/jcb.20276.
Regulated shedding of cell surface proteins is a mechanism for rapid activation of autocrine and paracrine signaling. Here we report that chelerythrine, a protein kinase C (PKC) inhibitor that possesses a variety of biological functions, is a potent inducer of heparin-binding epidermal growth factor-like growth factor (HB-EGF) shedding from the cell surface. Chelerythrine induced a time- and dose-dependent shedding of an HB-EGF-alkaline phosphatase (HB-EGF-AP) fusion protein expressed in MC2 rat prostate epithelial cells. The soluble form of HB-EGF-AP bound to heparin and exhibited potent biological activity as measured by DNA synthesis assay. Chelerythrine-induced HB-EGF shedding was metalloproteinase-(MMP-) mediated because specific MMP antagonists inhibited shedding by > or =60%. Chelerythrine stimulated production of reactive oxygen species, and antioxidants prevented chelerythrine-induced HB-EGF shedding, suggesting that the production of intracellular peroxides is necessary for this event. Consistent with this possibility, antioxidant- and MMP-inhibitable shedding was also demonstrated when hydrogen peroxide was used as an inducer. Although JNK/SAPK and p38 MAPK pathways were activated by chelerythine, these signaling mechanisms were not required to mediate the shedding event. However, JNK signaling was involved in chelerythrine-stimulated apoptosis. Our results suggest that HB-EGF shedding induced by chelerythrine is mediated predominantly via the production of reactive oxygen species.
细胞表面蛋白的调控性脱落是自分泌和旁分泌信号快速激活的一种机制。在此我们报告，白屈菜红碱是一种具有多种生物学功能的蛋白激酶C（PKC）抑制剂，是细胞表面肝素结合表皮生长因子样生长因子（HB-EGF）脱落的强效诱导剂。白屈菜红碱诱导了在MC2大鼠前列腺上皮细胞中表达的HB-EGF-碱性磷酸酶（HB-EGF-AP）融合蛋白呈时间和剂量依赖性的脱落。HB-EGF-AP的可溶性形式与肝素结合，并通过DNA合成测定显示出强效生物活性。白屈菜红碱诱导的HB-EGF脱落是由金属蛋白酶（MMP）介导的，因为特异性MMP拮抗剂抑制脱落达60%或更高。白屈菜红碱刺激活性氧的产生，抗氧化剂可阻止白屈菜红碱诱导的HB-EGF脱落，这表明细胞内过氧化物的产生对此过程是必需的。与此可能性一致的是，当用过氧化氢作为诱导剂时，也证明了抗氧化剂和MMP可抑制的脱落。虽然JNK/SAPK和p38 MAPK途径被白屈菜红碱激活，但这些信号机制并非介导脱落事件所必需。然而，JNK信号传导参与了白屈菜红碱刺激的细胞凋亡。我们的结果表明，白屈菜红碱诱导的HB-EGF脱落主要通过活性氧的产生来介导。
