The homeodomain transcription factor EMX2 is critical for central nervous system and urogenital development. In addition, EMX2 maps to a region of allelic deletion corresponding to a putative endometrial tumor suppressor at 10q26. We now report another polyadenylated transcript that is transcribed on the strand opposite to EMX2 and overlaps with the EMX2 transcript. This transcript was designated EMX2OS (OS, opposite strand), and an orthologous transcript present at the murine Emx2 locus was designated Emx2os. Alternative splicing to generate transcripts with varying 5' sequences was detected in the human but not the mouse. Neither ortholog contains a significant open reading frame, nor is primary sequence conserved between the two species. The sense and antisense transcripts display coordinate expression in that EMX2 and EMX2OS are abundant in normal postmenopausal endometrium, reduced in premenopausal endometrium, and absent or reduced in a majority of primary endometrial tumors. EMX2, EMX2OS, Emx2, and Emx2os are abundant in the uterine endometrium, with sense and antisense transcripts exhibiting identical expression patterns. Conservation of functional human and murine EMX2 antisense genes, of overlap between the sense and the antisense transcripts, and of identical cellular expression patterns suggests a biological function for EMX2OS, presumably to regulate EMX2.