Jun localization in cytosolic and nuclear compartments in brain–pituitary system of the frog, Rana esculenta: an analysis carried out in parallel with GnRH molecular forms during the annual reproductive cycle q R. Meccariello, a M. Mathieu, b G. Cobellis, a M. Vallarino, b F. Bruzzone, b G. Fienga, a R. Pierantoni, a, * and S. Fasano a a Dipartimento di Medicina Sperimentale, II Universit  a di Napoli, Via Costantinopoli 16, 80138 Napoli, Italy b Dipartimento di Biologia Sperimentale, Universit  a di Genova, Italy Received 16 July 2003; revised 18 September 2003; accepted 30 September 2003 Abstract The presence of c-jun like mRNA was assessed in the brain of the frog, Rana esculenta, during the annual sexual cycle. In parallel, Jun protein and GnRH molecular form (mammalian and chicken II also indicated as GnRH1 and GnRH2, respectively) activity was studied in order to establish possible relationships. Northern blot analysis of total RNA reveals the presence of a 2.7 kb c-jun- like mRNA. Western blots, carried out on cytoplasmic and nuclear protein extracts, show the presence of Jun immunoreactive band of 39 kDa in brain and pituitary. Fluctuations of c-jun-like mRNA and Jun immunoreactive protein (cytoplasmic and nuclear) levels in brains during the year indicate relationships among transcription, translation, and nuclear activity. In particular, mRNA levels increase gradually from September until November when Jun protein concentration peaks in cytosolic extracts. Conversely, the nuclear protein reaches highest concentration in July when the cytosolic level shows low values. Immunocytochemical studies confirm the presence of Jun immunoreactivity in both cytoplasmic and nuclear compartments of several brain areas, including those primarily involved in gonadotropin discharge (e.g., anterior preoptic area and preoptic nucleus). GnRH molecular forms and Jun are colocalized in anterior preoptic area and preoptic nucleus. Moreover, during the period characterized by GnRH release, Jun levels strongly decrease in nuclei. Finally, we show that treatments with a GnRH analog (buserelin, Hoechst, Frankfurt) increase Jun levels in brain nuclear extracts. Ó 2003 Elsevier Inc. All rights reserved. Keywords: Proto-oncogenes; Jun; GnRH; Brain; Amphibian 1. Introduction The dimeric transcription factor AP-1 (activator protein 1) is generally composed of members of Jun (c- Jun, Jun B, and Jun D) and Fos (c-Fos, Fos B, Fra-1, and Fra-2) families (Angel and Karin, 1991). Only Jun proteins form both homodimers and heterodimers, whereas Fos family members require heterodimerization to be functional. AP-1 dimers bind to a DNA regulatory element (AP-1) that is commonly found in a wide range of promoter and enhancer regions (Curran and Franza, 1988; Nakabeppu et al., 1988; Rauscher et al., 1988). Besides Fos heterodimerization and Jun homo/hetero- dimerization, Fos and Jun can form complexes with Maf (v-Maf, c-Maf, and Nrl) and ATF proteins (ATF2, LRF1/ATF3, and B-ATF) (Chinenov and Kerppola, 2001; vanDam and Castellazzi, 2001). More than 20 complexes can be formed and, depending on their composition, homo/heterodimers differentially modulate transcriptional activity. In the nervous system, Jun and Fos proteins are involved in numerous cellular events following physiological and pathophysiological stimuli, showing a temporospatial expression (Dragunow, 1996; Ferrer q This paper is dedicated to the memory of Aubrey Gorbman. * Corresponding author. Fax: +39-081-5667536. E-mail address: riccardo.pierantoni@unina2.it (R. Pierantoni). 0016-6480/$ - see front matter Ó 2003 Elsevier Inc. All rights reserved. doi:10.1016/j.ygcen.2003.09.017 www.elsevier.com/locate/ygcen General and Comparative Endocrinology 135 (2004) 310–323 GENERAL AND COMPARATIVE ENDOCRINOLOGY