The 5-HT 2C receptor is one of 14 different serotonin (5-HT) receptors that control neural function and behavior. Here, we present the entire sequence of a zebrafish 5-HT 2C receptor cDNA including the 3' untranslated region and the previously unknown 5' untranslated region. The cloned 5-HT 2C receptor gene is located on chromosome 7, is approximately 202kbp long, and contains six exons. The coding region of the gene is 1557bp long and flanked by a 504bp 5' UTR and a 1474bp 3' UTR. The deduced protein sequence of 518 amino acids aligns with orthologs of other vertebrates and is 54% identical to the human and mouse 5-HT 2C receptor protein sequences. The region of the cDNA that encodes the 2nd cytoplasmic loop of the protein shows a 66% identity with vertebrate orthologs and clearly identifies the gene as a 5-HT 2C receptor gene. Coupling sites for beta-arrestin and calmodulin are conserved in zebrafish. In-situ hybridization shows that the receptor is expressed in the brain and spinal cord including areas such as the olfactory bulb, the dorsal thalamus, the posterior tuberculum, the hypothalamus and the medulla oblongata. Reverse Transcriptase-PCR experiments indicate that the receptor gene can also be active in other tissues such as skin, ovaries, and axial muscle of adult zebrafish. Expression of the 5-HT 2C receptor during ontogeny was found as early as 2.5hpf. Five edited adenines in the region of the human, rat and mouse mRNA that encodes the 2nd cytoplasmic loop are conserved in the zebrafish transcript. However, RNA editing was not detected in the zebrafish. The results characterize the zebrafish 5-HT 2C receptor gene and gene expression pattern for the first time. The similarities to mammalian 5-HT 2C receptor genes suggest the use of zebrafish for the study of 5-HT 2C receptor function in behavior, development and drug discovery.
Bibliographical noteFunding Information:
The project was supported by a grant from the National Science Foundation (NSF) to H.S. ( IOS-0438404 ) and funding from DePauw University (DPU) and William Paterson University of New Jersey (WPUNJ) . We thank Pascal Lafontant and Stephen C. Ekker for helpful comments on the manuscript, Tammy Greenwood and Victoria Bedell for their help with in-situ hybridization experiments, Paula Evans for her help using the DNA sequencer at DPU, and Jane Voos for her support at the beginning of the project.
- In-situ hybridization