McManaman J L, Hanson L, Neville M C, Wright R M
Department of Biochemistry and Molecular Genetics, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA.
Arch Biochem Biophys. 2000 Jan 15;373(2):318-27. doi: 10.1006/abbi.1999.1573.
Xanthine oxidoreductase (XOR) is a prominent component of the milk lipid globule, whose concentration is selectively increased in mammary epithelial cells during the transition from pregnancy to lactation. To understand how XOR expression is controlled in the mammary gland, we investigated its properties and regulation by lactogenic hormones in cultured HC11 mammary epithelial cells. XOR was purified as the NAD(+)-dependent dehydrogenase by benzamidine-Sepharose chromatography and was shown to be intact and to have biochemical properties similar to those of enzyme from other sources. Treating confluent HC11 cells with prolactin and cortisol produced a progressive, four- to fivefold, increase in XOR activity, while XOR activity in control cells remained constant. Elevated cellular XOR activity was correlated with increased XOR protein and was due to both increased synthesis and decreased degradation of XOR. Prolactin and cortisol increased XOR protein and mRNA in the presence of epidermal growth factor, which blocked the stimulation of beta-casein synthesis by these hormones. Further, hormonal stimulation of XOR was inhibited by genistein (a protein tyrosine kinase inhibitor) and by PD 98059 (a specific inhibitor of the MAP kinase cascade). These findings indicate that lactogenic hormones stimulate XOR and beta-casein expression via distinct pathways and suggest that a MAP kinase pathway mediates their effects on XOR. Our results provide evidence that lactogenic hormones regulate milk protein synthesis by multiple signaling pathways.
黄嘌呤氧化还原酶（XOR）是乳脂肪球的一个重要组成部分，在从妊娠到泌乳的转变过程中，其浓度在乳腺上皮细胞中选择性增加。为了了解乳腺中XOR表达是如何被调控的，我们在培养的HC11乳腺上皮细胞中研究了其特性以及催乳激素对其的调控作用。通过苯甲脒-琼脂糖凝胶层析法将XOR纯化为依赖NAD(+)的脱氢酶，结果表明其结构完整，并且具有与其他来源的酶相似的生化特性。用催乳素和皮质醇处理汇合的HC11细胞，可使XOR活性逐渐增加4到5倍，而对照细胞中的XOR活性保持不变。细胞内XOR活性的升高与XOR蛋白的增加相关，这是由于XOR的合成增加和降解减少所致。在表皮生长因子存在的情况下，催乳素和皮质醇可增加XOR蛋白和mRNA的表达，而表皮生长因子会阻断这些激素对β-酪蛋白合成的刺激作用。此外，染料木黄酮（一种蛋白酪氨酸激酶抑制剂）和PD 98059（MAP激酶级联反应的特异性抑制剂）可抑制激素对XOR的刺激作用。这些发现表明，催乳激素通过不同途径刺激XOR和β-酪蛋白的表达，并提示MAP激酶途径介导了它们对XOR的作用。我们的结果提供了证据，证明催乳激素通过多种信号通路调节乳蛋白的合成。
