Chatterjee B, Mancini M A, Roy A K
Department of Cellular, University of Texas Health Science Center, San Antonio 78284.
Biochim Biophys Acta. 1990 May 16;1034(2):162-9. doi: 10.1016/0304-4165(90)90071-4.
In vitro translation of total rat hepatic mRNAs has identified a 31 kilodalton senescence marker protein (SMP-2) which is present in higher amounts in prepubertal and senescent males than in the post-pubertal adult male (more than 10-fold). SMP-2 is an androgen-repressible protein. The negative regulation of the SMP-2 gene activity by androgen accounts for its increased expression during the androgen insensitive states of the prepubertal and senescent livers, and its constitutive expression in the female liver. A combination of separation procedures including salt fractionation, chromatofocusing, ion-exchange chromatography and preparative gel electrophoresis have led to the purification of SMP-2 to apparent homogeneity. The purified protein showed the same electrophoretic mobility as the sex- and age-specific in vitro translation product of hepatic mRNAs. The polyclonal antibody to SMP-2 was produced in the rabbit. The antibody selectively reacted with the 31 kDa sex- and age-specific translation product of hepatic mRNAs. Western blot analysis of the liver cytosol confirms monospecificity of the antiserum, as well as age- and sex-dependent changes in the tissue level of SMP-2. Histochemical staining of liver sections with the antiserum reveals a preferential periportal localization of SMP-2 in the hepatocytes. This finding is in marked contrast to the androgen-inducible alpha 2u globulin which is preferentially synthesized and localized in the pericentral hepatocytes. Thus, the zonal distribution of SMP-2 correlates with polarized androgen sensitivity of the hepatocytes within the liver lobule.
对大鼠肝脏总mRNA进行体外翻译，鉴定出一种31千道尔顿的衰老标记蛋白（SMP-2），该蛋白在青春期前和衰老雄性大鼠中的含量高于青春期后成年雄性大鼠（超过10倍）。SMP-2是一种雄激素可抑制蛋白。雄激素对SMP-2基因活性的负调控解释了其在青春期前和衰老肝脏雄激素不敏感状态下表达增加，以及在雌性肝脏中组成性表达的原因。包括盐分级分离、色谱聚焦、离子交换色谱和制备性凝胶电泳在内的一系列分离程序相结合，已将SMP-2纯化至明显的均一性。纯化后的蛋白与肝脏mRNA的性别和年龄特异性体外翻译产物具有相同的电泳迁移率。用兔制备了抗SMP-2的多克隆抗体。该抗体与肝脏mRNA的31 kDa性别和年龄特异性翻译产物发生选择性反应。对肝脏细胞质进行蛋白质免疫印迹分析证实了抗血清的单特异性，以及SMP-2组织水平的年龄和性别依赖性变化。用该抗血清对肝脏切片进行组织化学染色，显示SMP-2在肝细胞中优先定位于门静脉周围。这一发现与雄激素诱导的α2u球蛋白形成鲜明对比，后者优先在中央周围肝细胞中合成和定位。因此，SMP-2的区域分布与肝小叶内肝细胞的极化雄激素敏感性相关。
