The Human Doublesex-Related Gene, DM RT2, Is Homologous to a Gene Involved in Somitogenesisand Encodes a Potential Bicistronic Transcript ChrisOttolenghi,* , † ,1 Reiner Veitia,* , ‡ ,1 Marcello Barbieri,† Marc Fellous,* and Ken McElreavey* ,2 * Unite ´ d’Immunoge ´ne ´tique humaine, INSERM U276, Institut Pasteur, 25 rue du Docteur Roux, 75724 Paris, France; †Dipartimento di Morfologia ed Embriologia, Universita ` di Ferrara, 64/B via Fossato di Mortara, 44100 Ferrara, Italy; and ‡Department of Biochemistry, University of Havana, Havana, Cuba Received October 14, 1999; accepted December 21, 1999 Intense efforts are currently being pursued to iden- tify autosomal genes associated with 46,XY male-to- female sex reversal. The genes DMRT1 and 2 are lo- cated on distal 9p, a region deleted in 46,XY sex- reversed patients. They are considered excellent candidates because of their homology to regulators of sex development in invertebrates. We present the genomic structure of DMRT2, showing that it gener- ates several transcripts with distinct coding potential. In addition to the previously reported 226-amino-acid protein-encoding transcript, we describe other mRNA isoforms that are potentially bicistronic and are pre- dicted to encode an additional 328-amino-acid polypeptide. Finally, a stop codon-containing exon (exon 4) can be skipped by alternative splicing and can generate a transcript that is predicted to encode a fusion protein. The latter shares 58% amino acid iden- tity with a gene recently described in fish, termed terra. Differences in expression pattern exist for DMRT2 mRNA isoforms among the human adult tis- sues tested, between adult tissues and human em- bryos, and between DMRT2 and DMRT1 during embry- onic development. We failed to detect mutations by sequencing of DMRT2 in a sample of 46,XY female pa- tients. The interesting structure of DMRT2 coupled to preliminary functional studies in fish showing that terra is involved in somitogenesis suggests that vali- dation or exclusion of this gene as a cause of sex re- versal will require more in-depth investigations. © 2000 Academic Press INTRODUCTION Our molecular understanding of the physiopathology of human sex determination began with the identifica- tion of SRY in 1990. However, while several genes associated with syndromes comprising genital and ex- tragenital anomalies have been recognized, most cases of isolated sex reversal are not accounted for by SRY mutations and remain unexplained (McElreavey et al., 1993; Nordenskjold et al., 1995; Kwok et al., 1996; Veitia et al., 1997). Deletions of the tip of chromosome 9 have been associated with an increasing number of cases of male-to-female sex reversal, and some patients with 9p deletions have anomalies restricted to sexual development (Alfi et al., 1976; Bennett et al., 1993). The critical deleted region is contained in 9p24.3 (Flejter et al., 1998; Veitia et al., 1998; Guioli et al., 1998). Inter- estingly, this region is homologous to a large syntenic region of the avian Z heterochromosome, which is present as a double dose in males and as a single dose in females (Nanda et al., 1999). Taken together, these results suggest that genes located on distal human 9p may cause male-specific developmental anomalies when hemizygous. Consistently, gene dosage effects have been frequently linked to vertebrate and inverte- brate sex reversal (reviewed in Hodgkin, 1990; Swain and Lovell-Badge, 1999). Positional cloning efforts are being pursued to isolate the gene(s) on 9p responsible for sex reversal. Two positional candidate genes have elicited particular in- terest because of their partial homology with the genes doublesex (dsx) of Drosophila and mab3 of Caenorhab- ditis, both involved in the sexual development of these organisms. One candidate, DMRT1, is specifically ex- pressed in adult testis (Raymond et al., 1998). A cDNA corresponding to a second DM domain gene, termed DMRT2, has also been mapped to the 9p critical region, although structural and functional studies of this gene have not been performed (Raymond et al., 1999). All deletions of chromosome 9 associated with sex reversal lead to loss of both DMRT1 and 2. This has prompted Raymond et al. (1999) to claim that one or both genes could be involved in male urogenital development, even though intensive searches by these authors have 1 These authors contributed equally to this work. 2 To whom correspondence should be addressed. Telephone: 00-33- 1-45688920. Fax: 00-33-1-40613153. E-mail: kenmce@pasteur.fr. Genomics 64, 179 –186 (2000) doi:10.1006/geno.2000.6120, available online at http://www.idealibrary.com on 179 0888-7543/00 $35.00 Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.