Exploring the Distribution and Potential Roles of the Developmental Transcription Factor Zic2 in an Adult Protogynous Teleost, Thalassoma bifasciatum

Abstract

The bluehead wrasse (Thalassoma bifasciatum) is a valuable model for studying neuroendocrine processes because it displays two discrete male phenotypes, initial phase (IP) males and territorial, terminal phase (TP) males, and undergoes socially-controlled protogynous sex change. Previously generated microarray-based comparisons suggested that zic2 was upregulated in the brains of terminal phase males relative to initial phase males. The highly conserved members of the zic family of zinc-finger transcription factors play critical roles in regulating cellular proliferation and differentiation, but their roles as transcription factors in adulthood are poorly understood. Studies aimed at understanding the conserved roles of zic transcription factors during vertebrate brain development have targeted zic2 as one of the few genes that is independent of the hedgehog pathway and linked to the congenital malformation of the forebrain termed holoprosencephaly (HPE). In addition, zic2 mRNA is found in a wide range of human cancers, and several researchers have found that zic family members may serve as a potential marker for abnormal cell growth. This study describes the localization and expression patterns across sexual phenotypes of neural zic2 in an adult teleost and offers insight into the potential role of zic2 as a regulatory factor in the sex-change signaling pathway.

We cloned a 727bp sequence for neural zic2 from field-collected bluehead wrasses. In situ hybridization with [α-35S]CTP-labeled riboprobes was utilized to localize and assess the relative abundance of brain zic2 mRNA across sexual phenotypes. We found that zic2 mRNA expression in the adult bluehead wrasse brain was not only extremely abundant in the granular cells of the cerebellum, but also widespread in other brain areas including in the thalamus, hypothalamus, habenula, torus semicircularis, torus longitudinalis, medial longitudinal fascicle and telencephalic areas. Quantitative autoradiography and phosphor screens showed zic2 mRNA hybridization signal density in the preoptic area of the hypothalamus was significantly higher in terminal phase males relative to both initial phase males and females, and silver grain analysis confirmed this relationship between phenotypes. No significant difference in abundance was found in zic2 signal across phenotypes in the habenula or cerebellum. As zic2 has been shown to regulate tissue specific expression of the dopamine receptor D1 (DRD1) and function as a transcriptional repressor, our study illustrates zic2 expression could potentially be acting on dopaminergic targets in the preoptic area of the hypothalamus, an area associated with sexually-motivated behavior and behavioral sex change.