Two isoforms of gonadotropin-releasing hormone are coexpressed in neuronal cell lines.

Abstract

GnRH-I serves as the neuropeptide that regulates mammalian reproduction. Recently, several groups have identified in the brain of rodents, monkeys, and humans a second isoform of GnRH (GnRH-II) whose structure is 70% identical to that of GnRH-I. In this study we demonstrate for the first time human and mouse neuronal cell lines that express both GnRH-I and GnRH-II. Following the screening of several human neuronal cell lines by RT-PCR and Southern hybridization, we demonstrated that two cell lines, TE-671 medulloblastoma and LAN-1 neuroblastoma cells, coexpress messenger RNA encoding the two isoforms of GnRH. Nucleotide sequencing indicated that the complementary DNA fragments are identical to those of the known human GnRH-I and GnRH-II sequences. Extracts obtained from the TE-671 and LAN-1 cell lines as well as from the immortalized mouse hypothalamic GT1-7 neuronal cell line were found to contain the two isoforms of GnRH, which exhibited identical chromatographic properties as synthetic GnRH-I and GnRH-II, in HPLC followed by specific RIAs. Furthermore, double immunofluorescence studies demonstrated the two GnRH isoforms in LAN-1, TE-671, and GT1-7 cells. The identification of neuronal cell lines expressing both GnRH-I and GnRH-II provides tools for studying the differential regulation of gene expression and secretion and for studying the interaction between the two isoforms. Such studies may contribute to elucidation of the physiological functions of GnRH-II, which are still unknown.