Two domains of the tolloid protein contribute to its unusual genetic interaction with decapentaplegic.

The zygotic dorsal-ventral patterning gene tolloid (tld) exhibits a complex structure consisting of an N-terminal domain with sequence similarity to the astacin family of metalloproteases and a C-terminal domain composed of two EGF-like repeats and five copies of the CUB repeat which was first found in human complement proteins C1r and C1s. The overall structure of TLD is similar to the human bone morphogenetic protein, BMP-1. Previous genetic studies (Ferguson and Anderson, 1992) have shown that certain tld alleles behave antagonistically toward decapentaplegic (dpp), a second zygotic dorsal-ventral patterning gene whose product is most closely related to the TGF-beta superfamily members BMP-2 and BMP-4. The common phenotype and genetic interaction data may suggest a direct physical association between these two proteins. In this study, we use an allelic series of haplo-viable dpp mutations to order the strength of the tld antagonistic interaction. In addition, we have identified the lesions associated with 20 tld mutations by DNA sequencing. These mutations consist of six alleles that behave in an antimorphic manner with respect to dpp, five alleles that revert the antagonistic interaction, and 9 loss-of-function alleles with a range of phenotypes from weak to strong. We find that antimorphic mutations and second-site revertants cluster to the protease domain and to a second site located at the junction of the first two CUB repeats. In contrast, loss-of-function mutations are dispersed throughout the length of the protein. Those mutations that map within the CUB repeats invariably affect consensus residues, thereby establishing the functional significance of certain residues in the CUB repeat consensus sequence. Possible molecular models for TLD action are discussed.

Two distinct transmembrane serine/threonine kinases from Drosophila melanogaster form an activin receptor complex.

A transmembrane protein serine/threonine kinase, Atr-I, that is structurally related to receptors for members of the transforming growth factor-beta (TGF-beta) family has been cloned from Drosophila melanogaster. The spacing of extracellular cysteines and the cytoplasmic domain of Atr-I resemble most closely those of the recently described mammalian type I receptors for TGF-beta and activin. When expressed alone in test cells, Atr-I is unable to bind TGF-beta, activin, or bone morphogenetic protein 2. However, Atr-I binds activin efficiently when coexpressed with the distantly related Drosophila activin receptor Atr-II, with which it forms a heteromeric complex. Atr-I can also bind activin in concert with mammalian activin type II receptors. Two alternative forms of Atr-I have been identified that differ in an ectodomain region encompassing the cysteine box motif characteristic of receptors in this family. Comparison of Atr-I with other type I receptors reveals the presence of a characteristic 30-amino-acid domain immediately upstream of the kinase region in all these receptors. This domain, of unknown function, contains a repeated Gly-Ser sequence and is therefore referred to as the GS domain. Maternal Atr-I transcripts are abundant in the oocyte and widespread during embryo development and in the imaginal discs of the larva. The structural properties, binding specificity, and dependence on type II receptors define Atr-I as an activin type I receptor from D. melanogaster. These results indicate that the heteromeric kinase structure is a general feature of this receptor family.

Identification of a Drosophila activin receptor.

Activins are cytokines of the transforming growth factor beta superfamily that control various events during vertebrate embryo development and cell differentiation in the adult, and act through transmembrane receptors that contain a cytoplasmic protein-serine/threonine kinase domain. We describe the identification, deduced primary structure, and expression pattern of Atr-II, a receptor serine/threonine kinase found in Drosophila. With the exception of the spacing of 10 cysteine residues, the extracellular domain of Atr-II is very dissimilar from those of vertebrate activin receptors, yet it binds activin with high affinity and specificity. The kinase domain sequence of Atr-II is 60% identical to those of activin receptors from vertebrates, suggesting similarities in their signaling mechanisms. Maternal Atr-II transcript and its product are abundant in the oocyte. During development, the highest levels of Atr-II transcript and protein are observed in the mesoderm and gut. The possible role of an activin signaling system in Drosophila development is discussed.

The Drosophila dorsal-ventral patterning gene tolloid is related to human bone morphogenetic protein 1.

Mutations in the Drosophila tolloid (tld) gene lead to a partial transformation of dorsal ectoderm into ventral ectoderm. The null phenotype of tld is similar to, but less severe than decapentaplegic (dpp), a TGF-beta family member required for the formation of all dorsal structures. We have cloned the tld locus by P element tagging. At the blastoderm stage, tld RNA is expressed dorsally, similar to that described for dpp. Analysis of a tld cDNA reveals three sequence motifs: an N terminal region of similarity to a metalloprotease, two EGF-like repeats, and five copies of a repeat found in human complement proteins C1r and C1s. tld sequence is 41% identical to human bone morphogenetic protein 1 (BMP-1); the closest members to dpp within the TGF-beta superfamily are BMP-2 and BMP-4, two other bone morphogenetic proteins. These findings suggest that these genes are members of a signal generating pathway that has been conserved between insects and mammals.