Chen Alon M, Perrin Marilyn H, Digruccio Michael R, Vaughan Joan M, Brar Bhawanjit K, Arias Carlos M, Lewis Kathy A, Rivier Jean E, Sawchenko Paul E, Vale Wylie W
Clayton Foundation Laboratories for Peptide Biology and Laboratory of Neuronal Structure and Function, The Salk Institute for Biological Studies, La Jolla, CA 92037, USA.
Proc Natl Acad Sci U S A. 2005 Feb 15;102(7):2620-5. doi: 10.1073/pnas.0409583102. Epub 2005 Feb 8.
Peptides of the corticotropin-releasing factor (CRF) family signal through the activation of two receptors, CRF receptor type 1 (CRFR1) and type 2 (CRFR2), both of which exist as multiple splice variants. We have identified a cDNA from mouse brain encoding a splice variant, soluble CRFR2alpha (sCRFR2alpha), in which exon 6 is deleted from the gene encoding CRFR2alpha. Translation of this isoform produces a predicted 143-aa soluble protein. The translated protein includes a majority of the first extracellular domain of the CRFR2alpha followed by a unique 38-aa hydrophilic C terminus resulting from a frame shift produced by deletion of exon 6. By using RT-PCR and Southern hybridization, the relative mRNA expression levels of full-length (seven transmembrane domains) CRFR2alpha and the soluble form (sCRFR2alpha) in the mouse brain were measured with a single reaction. The results demonstrate high levels of expression of sCRFR2alpha in the olfactory bulb, cortex, and midbrain regions. A rabbit antiserum raised against a synthetic peptide fragment encoding the unique C terminus revealed specific sCRFR2alpha immunoreactivity in mouse brain slices by immunohistochemistry and in extracts of brain regions by RIA. Interestingly, the sCRFR2alpha immunoreactivity distribution closely approximated that of CRFR1 expression in rodent brain. A protein corresponding to sCRFR2alpha, expressed and purified from either mammalian or bacterial cell systems, binds several CRF family ligands with low nanomolar affinities. Furthermore, the purified sCRFR2alpha protein inhibits cellular responses to CRF and urocortin 1. These data support a potential role of the sCRFR2alpha protein as a possible biological modulator of CRF family ligands.
促肾上腺皮质激素释放因子（CRF）家族的肽通过激活两种受体发出信号，即1型CRF受体（CRFR1）和2型（CRFR2），这两种受体均以多种剪接变体的形式存在。我们从小鼠脑中鉴定出一种编码剪接变体可溶性CRFR2α（sCRFR2α）的cDNA，其中编码CRFR2α的基因中外显子6被缺失。该异构体的翻译产生一种预测的143个氨基酸的可溶性蛋白。翻译后的蛋白包括CRFR2α的大部分第一个细胞外结构域，随后是一个独特的38个氨基酸的亲水性C末端，这是由于外显子6缺失导致的移码产生的。通过使用RT-PCR和Southern杂交，在单一反应中测量了小鼠脑中全长（七个跨膜结构域）CRFR2α和可溶性形式（sCRFR2α）的相对mRNA表达水平。结果表明，sCRFR2α在嗅球、皮质和中脑区域高表达。针对编码独特C末端的合成肽片段产生的兔抗血清通过免疫组织化学在小鼠脑切片中以及通过放射免疫分析在脑区提取物中显示出特异性的sCRFR2α免疫反应性。有趣的是，sCRFR2α免疫反应性分布与啮齿动物脑中CRFR1的表达分布非常接近。从哺乳动物或细菌细胞系统表达和纯化的与sCRFR2α对应的蛋白以低纳摩尔亲和力结合几种CRF家族配体。此外，纯化的sCRFR2α蛋白抑制细胞对CRF和尿皮质素1的反应。这些数据支持sCRFR2α蛋白作为CRF家族配体可能的生物调节剂的潜在作用。
