Polarity in the Drosophila eye is manifested as a dorsoventral reflection of two chiral forms of the individual unit eyes, or ommatidia. These forms fall on opposite sides of a dorsoventral midline of mirror symmetry known as the equator. Polarity is established in the eye imaginal disc as cells adopt their fates and as the ommatidial precursors undergo coordinated rotation within the epithelium; the mechanisms that coordinate these early patterning events remain poorly understood. We have identified a novel gene, strabismus (stbm), which is required to establish polarity in the eye, legs and bristles of Drosophila. Many stbm ommatidia are reversed anteroposteriorly and/or dorsoventrally. In stbm eye discs, ommatidial rotation is delayed and some ommatidial precursors initiate rotation in the wrong direction. Mosaic analysis indicates that stbm is ommatidium autonomous and required in most, if not all, photoreceptors within an ommatidium to establish normal polarity. stbm also appears to play an instructive role during the establishment of the fates of photoreceptors R3 and R4. stbm encodes a novel protein with a potential PDZ domain-binding motif and two possible transmembrane domains. Sequence analysis of both vertebrate and invertebrate homologs indicates that stbm has been highly conserved throughout evolution.