Real time PCR quantification of GFRα-2 alternatively spliced isoforms in murine brain and peripheral tissues

Abstract

The neurotrophic factor neurturin (NTN) is structurally related to the glial-derived neurotrophic factor (GDNF) and has been shown to prevent the degeneration of dopaminergic neurons both in vitro and in vivo. The preferred receptor for NTN is the GDNF family receptor alpha 2 (GFRα-2). To date, three protein-coding alternatively spliced GFRα-2 isoforms (GFRα-2a, GFRα-2b, GFRα-2c) have been identified in mammalian tissues. An accurate quantification of the expression levels is necessary when determining the contributions of these isoforms to NTN signaling in tissues. In this report, sequence independent real time RT-PCR is used to determine the expression levels of GFRα-2 isoforms at different developmental stages of the murine embryos, and in various adult tissues. In the adult murine brain, GFRα-2a was found to be the most abundant, GFRα-2c was slightly less and GFRα-2b was 10-fold lower. The testis did not appear to express significant levels of GFRα-2a, 2b or 2c, compared to the brain. A novel finding in this study is that in some tissues, including the adult brain, the expression levels of GFRα-2, as quantified by the amplification of the 3′ sequences encoding the putative glycosyl-phosphatidylinositol anchor signal sequence, were significantly higher than the combined levels of GFRα-2a, GFRα-2b and GFRα-2c. This indicates the existence of yet to be identified forms of GFRα-2 in some tissues that may be of physiological significance. © 2003 Elsevier Science B.V. All rights reserved.