It is known that the spitz class genes, pointed (pnt), rhomboid (rho), single-minded (sim), spitz (spi) and Star (S) as well as argos (aos), orthodenticle (otd) and Drosophila EGF receptor (DER) genes are required for proper formation of ventral neuroblasts and epidermis derived from ventral neuroectoderm. The expression of the spitz class genes was analyzed to determine the spatial and temporal pattern of each individual spitz class gene expression and to establish a hierarchical relationship of the spitz class genes in cell fate determination of ventral neuroectoderm. This analysis showed that aos, otd, pnt, rho and spi expression is severely reduced in sim mutant and greatly broadened in the HS-sim mutant. In addition, sim expression in the other spitz class, DER and otd mutants, is not affected. These results indicate that the sim gene acts upstream of all the spitz class genes as well as aos, DER and otd. It was also shown that otd and pnt expression in the ventralmost neuroectoderm is almost abolished in rho, S, spi and DER mutants and their expression in the HS-S, -spi, -rho and DER^(ELP) is expanded, indicating that otd and pnt genes act downstream of the most spitz class genes. aos and otd expression in pnt mutant becomes almost abolished and otd expression is widened into 3-4 rows of medial neuroectodermal cells in aos mutant and its expression is reduced into 1 row of cell in the HS-aos mutant, indicating that the aos gene acts downstream of pnt and upstream of the otd gene. These results suggest the following tentative hierarchical relationship among the spitz class and aos, DER and otd genes: sim → rho, S, spi → DFR → pnt → aos → otd. The finding that otd and pnt expression in the medial neuroectoderm of sim mutant is affected indicates the presence of cell-cell interaction between the medial neuroectoderm and CNS midline cells through the EGF signaling pathway.