Wiese R J, Uhland-Smith A, Ross T K, Prahl J M, DeLuca H F
College of Agricultural and Life Sciences, University of Wisconsin-Madison, Department of Biochemistry 53706.
J Biol Chem. 1992 Oct 5;267(28):20082-6.
Several studies have shown that the 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) receptor protein levels increase in response to 1,25-(OH)2D3. We have studied the mechanism of this regulation in both mouse fibroblasts and rat intestinal epithelial cells. Cell extracts and total RNA were prepared at varying times after addition of 1,25-(OH)2D3. The 1,25-(OH)2D3 receptor protein levels, measured using an immunoradiometric assay, rose significantly 2-3 h posttreatment and had risen 3-fold at 8 h. Concurrently, the 1,25-(OH)2D3 receptor mRNA content, measured using a ribonuclease protection assay, was not altered by 1,25-(OH)2D3 during this time. In cycloheximide-blocked cells, the administration of 1,25-(OH)2D3 markedly reduced the degradation rate of previously formed receptor. The 1,25-(OH)2D3 receptor protein half-life was determined as 4 h in the absence of 1,25-(OH)2D3 and increased to at least 8 h in the presence of 1,25-(OH)2D3. We also measured the 1,25-(OH)2D3 receptor mRNA levels in the duodena and kidney of vitamin D-deficient rats after a single 150-pmol injection of 1,25-(OH)2D3. Again, we found that 1,25-(OH)2D3 receptor mRNA levels were not changed in these tissues after 1,25-(OH)2D3 treatment. Therefore, the elevation of the 1,25-(OH)2D3 receptor protein following 1,25-(OH)2D3 administration is apparently the result of increased receptor protein lifetime and not increased transcription.
多项研究表明，1,25 - 二羟基维生素D3（1,25-(OH)2D3）受体蛋白水平会因1,25-(OH)2D3而升高。我们已在小鼠成纤维细胞和大鼠肠上皮细胞中研究了这种调节机制。在添加1,25-(OH)2D3后的不同时间制备细胞提取物和总RNA。使用免疫放射分析测定的1,25-(OH)2D3受体蛋白水平在处理后2 - 3小时显著升高，8小时时升高了3倍。同时，在此期间，使用核糖核酸酶保护分析测定的1,25-(OH)2D3受体mRNA含量并未因1,25-(OH)2D3而改变。在环己酰亚胺阻断的细胞中，给予1,25-(OH)2D3显著降低了先前形成的受体的降解速率。在不存在1,25-(OH)2D3的情况下，1,25-(OH)2D3受体蛋白的半衰期确定为4小时，在存在1,25-(OH)2D3的情况下增加到至少8小时。我们还在单次注射150 pmol的1,25-(OH)2D3后，测量了维生素D缺乏大鼠十二指肠和肾脏中的1,25-(OH)2D3受体mRNA水平。同样，我们发现1,25-(OH)2D3处理后这些组织中的1,25-(OH)2D3受体mRNA水平没有变化。因此，给予1,25-(OH)2D3后1,25-(OH)2D3受体蛋白的升高显然是受体蛋白寿命增加的结果，而非转录增加的结果。
