Multiple positive cis elements regulate the asymmetric expression of the SpHE gene along the sea urchin embryo animal-vegetal axis.

The mechanism that establishes the maternally determined animal-vegetal axis of sea urchin embryos is unknown. We have analyzed the cis-regulatory elements of the SpHE gene of Strongylocentrotus purpuratus, which is asymmetrically expressed along this axis, in an effort to identify components of maternal positional information. Previously, we defined a regulatory region that is sufficient to provide correct nonvegetal expression of a beta-galactosidase reporter gene (Wei, Z., Angerer, L. M., Gagnon, M. L., and Angerer, R. C., Dev. Biol. 171, 195-211, 1995). We have now analyzed this region intensively in order to determine if the spatial pattern is controlled by nonvegetal-positive activities or by vegetal-negative activities. The regulatory sequences, except the basal promoter, were mutated by either deletion or sequence replacement. None of these mutations resulted in ectopic beta-gal expression in vegetal cells, showing that no single negative cis element is responsible for the lack of vegetal SpHE transcription. Surprisingly, even short segments of the regulatory region containing only several identified cis elements also direct nonvegetal expression. Furthermore, the SpHE basal promoter functions effectively in vegetal cells in combination with cis-acting elements derived from the PMC-specific gene, SM50. We conclude that the spatial pattern of SpHE transcription is achieved by multiple positive activities concentrated in nonvegetal cells. The vegetal expression of SM50 also is regulated only by positive activities (Makabe, K. W., Kirchhamer, C. V., Britten, R. J., and Davidson, E. H., Development 121, 1957-1970, 1995). A chimeric promoter containing both SpHE and SM50 regulatory sequences is active ubiquitously, suggesting that these regulators are not reciprocally repressive. These observations suggest a model in which the SpHE and SM50 genes are activated by separate sets of positive maternal activities concentrated, respectively, in nonvegetal and vegetal domains of the early embryo.