Novel Member of the Otx Family 1 Otx 3 : identification , tissue expression and functional characterization of a novel member of the Otx family Running title : Otx 3 , a novel member of the Otx family

Zhang et al. (Revised version) Otx3, a Novel Member of the Otx Family 2 SUMMARY Transcription factors containing a homeodomain (HD) play an important role in the organogenesis of vertebrates. We have isolated a novel HD transcription factor, Otx3, which is structurally and functionally related to Otx1 and Otx2, transcription factors that are critical in brain morphogenesis. Mouse Otx3 is a protein composed of 376 amino acids. Otx3 mRNA is expressed in mouse embryos from 10.5 days post coitum (dpc) to 13.5 dpc and in adult cerebellum, as assessed by Northern blotting. Whole-mount in situ hybridization of mouse embryos from 9.5 dpc to 11.5 dpc revealed strong expression of Otx3 mRNA in the diencephalon, mesencephalon, metencephalon, myelencephalon, and developing eye, indicating an expression pattern largely overlapping but distinct from those of Otx1 and Otx2. In addition, Otx3 is shown by electrophoretic mobility shift assay (EMSA) to bind to the TAATCC motif, the consensus binding sequence for Otx1, Otx2, and Crx. Transcription reporter assay suggests that Otx3 functions as a transcription repressor by binding to this motif. These results suggest that Otx3 is a novel member of the Otx family and may be involved in the development of the central nervous system (CNS).