RESEARCH ARTICLE Molecular Reproduction & Development 77:521–532 (2010) Molecular Cloning and Expression Analysis of Fshr and Lhr in Relation to Fshb and Lhb Subunits During the Period of Temperature-Dependent Sex Determination in Pejerrey Odontesthes bonariensis TAKAHIRO SHINODA, 1 LEANDRO A. MIRANDA, 2 KOZUE OKUMA, 1 RICARDO S. HATTORI, 1 JUAN I. FERNANDINO, 2 GORO YOSHIZAKI, 1 GUSTAVO M. SOMOZA, 2 AND CARLOS A. STR € USSMANN 1 * 1 Graduate School of Marine Science and Technology, Tokyo University of Marine Science and Technology, Minato, Tokyo, Japan 2 Laboratorio de Ictiofisiologı ´a y Acuicultura, Instituto de Investigaciones Biotecnol  ogicas, Instituto Tecnol  ogico de Chascom us, Chascom us, Argentina SUMMARY In this study, we cloned and characterized the follicle stimulating hormone receptor (Fshr) and luteinizing hormone receptor (Lhr) cDNAs of pejerrey Odontesthes bonariensis, a species with temperature-dependent sex determination (TSD), and analyzed their expression in relation to Fshb and Lhb subunits during gonadogenesis at temperatures producing only females (17  C, FPT), both sexes (25  C, MixPT), and only males (29  C, MPT). The pejerrey Fshr cDNA had 3,069 bp for a mature protein of 694 amino acids (aa) and a signal peptide of 22 aa; the Lhr cDNA had 2,936 bp for a mature protein of 676 aa and a signal peptide of 25 aa. With the exception of Lhr in fish at the MPT, all genes showed significant increases and/or peaks of expression before histological differentiation of the gonads regardless of temperature. Larvae at the FPT had lower Fshb and Lhb but higher Lhr expression during the TSD period than those at the MPT; a clear pattern could not be ascertained for Fshr. At the MixPT, Fshb, Lhb, and Lhr mRNA increased in approximately half of the fish during TSD and sex differentiation and the sex ratio was 55.2% male. Based on the above results, it is suggested that animals with high Fshb and Lhb and low Lhr values represent putative males. These evidences, together with other studies, suggest that temperature may signal through the pituitary (differential expression of Fshb and Lhb) down to the gonads (differential expression of Lhr), probably affecting the regulation of steroido- genesis during the TSD process of pejerrey. Mol. Reprod. Dev. 77: 521–532, 2010. ß 2010 Wiley-Liss, Inc. Received 25 August 2009; Accepted 10 February 2010 * Corresponding author: Graduate School of Marine Science and Technology Tokyo University of Marine Science and Technology Konan 4-5-7, Minato Tokyo 108-8477, Japan. E-mail: carlos@kaiyodai.ac.jp Published online 30 March 2010 in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/mrd.21179 INTRODUCTION In teleost species, as in other vertebrates, the reproduc- tive cycle is controlled by the brain–pituitary–gonadal axis (Zohar et al., 2010). The pituitary gonadotropins (GtHs), heterodimeric glycoproteins known as follicle stimulating hormone (FSH) and luteinizing hormone (LH), are key Abbreviations: ECD ¼ extracellular domain; FPT ¼ female-producing temper- ature; FSH ¼ follicle-stimulating hormone; GtH ¼ gonadotropin; LH ¼ luteinizing hormone receptor; MixPT ¼ mixed sex-producing temperature; MPT ¼ male- producing temperature; TMD ¼ transmembrane domain; TSD ¼ temperature- dependent/thermolabile sex determination ß 2010 WILEY-LISS, INC.