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小鼠中两种生长激素受体mRNA及初级翻译产物的检测

Detection of two growth hormone receptor mRNAs and primary translation products in the mouse.

作者信息

Smith W C, Linzer D I, Talamantes F

机构信息

Department of Biology, University of California, Santa Cruz 95064.

出版信息

Proc Natl Acad Sci U S A. 1988 Dec;85(24):9576-9. doi: 10.1073/pnas.85.24.9576.

Abstract

Two mouse growth hormone-receptor primary translation products of Mr 95,900 and 31,800 were identified from in vitro-translated late pregnant mouse liver mRNA. RNA isolated from mouse liver was translated in a rabbit reticulocyte lysate system containing [35S]methionine, and the growth hormone receptor primary translation products were identified by immunoprecipitation with anti-mouse growth hormone receptor antiserum followed by sodium dodecyl sulfate/PAGE and fluorography. Detectable amounts of the Mr 95,900 and 31,800 proteins were not present in in vitro-translated nonpregnant mouse liver mRNA. This result is consistent with previous observations of the up-regulation of growth hormone receptors in the liver during pregnancy in the mouse. Northern (RNA) blot analysis of mouse liver and adipose tissue RNA with a rabbit growth hormone receptor cDNA probe revealed two hybridizing mRNAs of approximately 3.9 and 1.2 kilobases. These two RNAs were greatly up-regulated in liver, but not in adipose tissue, during pregnancy. The sizes of these mRNAs closely matched predictions of the sizes of the mRNAs coding for the proteins of Mr 95,900 and 31,800 made by in vitro translation of size-fractionated late-pregnant mouse liver poly(A)+RNA. These results suggest a mechanism for the generation of both the heterogeneous forms of the growth hormone receptor identified in mouse liver membrane preparations and the mouse serum growth hormone-binding protein.

摘要

从小鼠妊娠后期肝脏mRNA体外翻译产物中鉴定出两种分子量分别为95,900和31,800的小鼠生长激素受体初级翻译产物。从小鼠肝脏分离的RNA在含有[35S]甲硫氨酸的兔网织红细胞裂解物系统中进行翻译,然后用抗小鼠生长激素受体抗血清进行免疫沉淀,接着进行十二烷基硫酸钠/聚丙烯酰胺凝胶电泳(SDS/PAGE)和荧光自显影,从而鉴定出生长激素受体初级翻译产物。在非妊娠小鼠肝脏mRNA的体外翻译产物中未检测到可察觉量的分子量为95,900和31,800的蛋白质。这一结果与先前关于小鼠妊娠期间肝脏中生长激素受体上调的观察结果一致。用兔生长激素受体cDNA探针进行的小鼠肝脏和脂肪组织RNA的Northern(RNA)印迹分析显示出两条分别约为3.9和1.2千碱基的杂交mRNA。在妊娠期间,这两种RNA在肝脏中大幅上调,但在脂肪组织中未上调。这些mRNA的大小与通过对妊娠后期小鼠肝脏聚腺苷酸加尾(poly(A)+)RNA进行大小分级体外翻译所预测的编码分子量为95,900和31,800蛋白质的mRNA大小紧密匹配。这些结果提示了一种机制,可用于解释在小鼠肝细胞膜制剂中鉴定出的生长激素受体的异质形式以及小鼠血清生长激素结合蛋白的产生。

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