OLIGOMERIZATION OF THE EXTRACELLULAR DOMAIN OF BOSS ENHANCES ITS BINDING TO THE SEVENLESS RECEPTOR AND ITS ANTAGONISTIC EFFECT ON R7 INDUCTION

In the developing compound eye of Drosophila, neuronal differentiation of the R7 photoreceptor cell is induced by the interaction of the rec eptor tyrosine kinase Sevenless with its ligand Bride of sevenless (Bo ss), which is expressed on the neighboring R8 cell. Boss is an unusual ligand of a receptor tyrosine kinase: it is composed of a large extra cellular domain, a transmembrane domain with seven membrane-spanning s egments and a cytoplasmic tail, Expression of a monomeric, secreted fo rm of the extracellular domain of Boss is not sufficient for Sevenless activation, and instead acts as a weak antagonist. Because oligomeriz ation appears to be a critical step in the activation of receptor tyro sine kinases, we used oligomerized forms of the Boss extracellular dom ain to test their ability to bind to Sevenless in vivo and restore R7 induction in vivo, Oligomerization was achieved by fusion to the leuci ne zipper of the yeast transcription factor GCN4 or to the tetrameriza tion helix of Lac repressor. Binding of these multivalent proteins to Sevenless could be detected in vitro by immunoprecipitation of cross-l inked ligand/receptor complexes and in vivo by receptor-dependent liga nd localization. However, neither RS-specific or ubiquitous expression of multivalent Exboss ligands rescued the boss phenotype, Instead, th ese ligands acted as competitive inhibitors for wild-type Boss protein and thereby suppressed R7 induction. Therefore the role of the transm embrane or cytoplasmic domains of Boss in the activation of the Sev re ceptor cannot be replaced by oligomerization.