Department of Biology, Tufts University, Medford, MA 02155, USA.
Gen Comp Endocrinol. 2013 Nov 1;193:27-36. doi: 10.1016/j.ygcen.2013.07.008. Epub 2013 Jul 24.
The stress response represents an animal's attempt to cope with a noxious stimulus through a rapid release of corticosterone or cortisol (CORT) into the bloodstream, resulting in a suite of physiological and behavioral changes. These changes are mediated in large part through CORT's binding to two different intracellular receptors, the high-affinity mineralocorticoid receptor (MR) and the lower-affinity glucocorticoid receptor (GR). We tested the hypothesis that GR and MR mRNA expression would correlate with functional protein expression in neuronal tissue of wild-caught house sparrows (Passer domesticus). To test this hypothesis, we performed a parallel procedure in which protein concentrations were quantified in one half of house sparrow brains (n=16) using radioligand binding assays, and mRNA levels were quantified in the other brain half using reverse-transcriptase quantitative PCR (RT-qPCR). Two reference genes, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and TATA-box binding protein (TBP), were used for relative quantification of GR and MR mRNA. Quantifications showed that these two reference genes were not correlated with each other. Furthermore, there was no correlation between mRNA and protein levels for GR or MR using either reference gene, suggesting that regulation of mRNA and protein levels for MR and GR is not tightly linked. This study provides insight into the importance of regulatory steps between mRNA expression and the creation and stability of a functional protein. The overall conclusion is that mRNA expression cannot be used as a proxy for GR or MR binding in house sparrows.
应激反应代表动物通过将皮质酮或皮质醇(CORT)迅速释放到血液中,尝试应对有害刺激,从而导致一系列生理和行为变化。这些变化在很大程度上是通过 CORT 与两种不同的细胞内受体结合介导的,即高亲和力盐皮质激素受体(MR)和低亲和力糖皮质激素受体(GR)。我们测试了这样一个假设,即 GR 和 MR mRNA 表达与野生捕获的家麻雀(Passer domesticus)神经元组织中的功能性蛋白表达相关。为了验证这一假设,我们进行了平行程序,使用放射性配体结合测定法在一半家麻雀大脑(n=16)中定量蛋白质浓度,并使用逆转录定量 PCR(RT-qPCR)在另一半大脑中定量 mRNA 水平。使用甘油醛-3-磷酸脱氢酶(GAPDH)和 TATA 框结合蛋白(TBP)这两个参考基因,对 GR 和 MR mRNA 进行相对定量。定量结果表明,这两个参考基因彼此之间没有相关性。此外,使用任一参考基因,GR 或 MR 的 mRNA 和蛋白水平之间均没有相关性,这表明 MR 和 GR 的 mRNA 和蛋白水平的调节并不紧密相关。本研究深入了解了 mRNA 表达与功能性蛋白的产生和稳定性之间的调节步骤的重要性。总的结论是,在麻雀中,mRNA 表达不能作为 GR 或 MR 结合的替代物。