Interactions Between Wingless and Frizzled Molecules in Drosophila 3
another cell line, clone-8 derived from Drosophila imaginal discs (Van
Leeuwen et al. 1994).
S2 cells themselves are unable to respond to Wg (Yanagawa et al.
1995). This observation proved to be the key in identifying Wg receptors, as we found that a Drosophila frizzled-related gene, Dfz2, identified in the laboratory of our collaborator Dr. Jeremy Nathans, is expressed in clone-8 cells but not in nonresponding S2 cells. When we
transfected the Dfz2 gene into S2 cells, these cells became active in W g
signal transduction. In addition, the S2 cells could bind W g protein on
their cell surface. Transfection of cells with Dfz2 constructs lacking
either the extracellular or intracellular domain of the protein demonstrated that the extracellular domain was required for binding (Bhanot et
al. 1996). We tested several other members ofthe Fz family in the same
assays, finding that the original Fz protein, which genetically had not
been implicated in Wg signaling, can confer wg responsiveness to S2
cells, as well as Wg binding. A more distant relative, the Smoothened
(Smo) protein, does not bind, showing that these assays can discriminate
between family members.
These in vitro results suggest that Dfz2 and Fz could have overlapping functions, a hypothesis that we could strengthen recently from
genetic data described below. Although these findings solved a longstanding problem in the field, we could not provide evidence that Dfz2
is required for W g signaling in the fly, because there were no mutants in
the gene.
1.4 W g Signaling
These and other experiments have led to the following current model of
Wntlwg signaling, which is based on both genetics in Drosophila and
cell biological data (Fig. 1). In the absence of W g signaling, the protein
kinase Zw3 (or GSK-3) inactivates Arm, possibly by direct phosphorylation of Arm followed by proteolytic breakdown. This effect of Zw3 on
Arm is then relieved by W g, resulting in stabilization and upregulation
of the Arm protein. The proteins Axin and APC are both involved in
downregulating Arm, by forming a complex with Arm and Zw3. After
Arm has escaped from these negative regulators, it can act in the nucleus
another cell line, clone-8 derived from Drosophila imaginal discs (Van
Leeuwen et al. 1994).
S2 cells themselves are unable to respond to Wg (Yanagawa et al.
1995). This observation proved to be the key in identifying Wg receptors, as we found that a Drosophila frizzled-related gene, Dfz2, identified in the laboratory of our collaborator Dr. Jeremy Nathans, is expressed in clone-8 cells but not in nonresponding S2 cells. When we
transfected the Dfz2 gene into S2 cells, these cells became active in W g
signal transduction. In addition, the S2 cells could bind W g protein on
their cell surface. Transfection of cells with Dfz2 constructs lacking
either the extracellular or intracellular domain of the protein demonstrated that the extracellular domain was required for binding (Bhanot et
al. 1996). We tested several other members ofthe Fz family in the same
assays, finding that the original Fz protein, which genetically had not
been implicated in Wg signaling, can confer wg responsiveness to S2
cells, as well as Wg binding. A more distant relative, the Smoothened
(Smo) protein, does not bind, showing that these assays can discriminate
between family members.
These in vitro results suggest that Dfz2 and Fz could have overlapping functions, a hypothesis that we could strengthen recently from
genetic data described below. Although these findings solved a longstanding problem in the field, we could not provide evidence that Dfz2
is required for W g signaling in the fly, because there were no mutants in
the gene.
1.4 W g Signaling
These and other experiments have led to the following current model of
Wntlwg signaling, which is based on both genetics in Drosophila and
cell biological data (Fig. 1). In the absence of W g signaling, the protein
kinase Zw3 (or GSK-3) inactivates Arm, possibly by direct phosphorylation of Arm followed by proteolytic breakdown. This effect of Zw3 on
Arm is then relieved by W g, resulting in stabilization and upregulation
of the Arm protein. The proteins Axin and APC are both involved in
downregulating Arm, by forming a complex with Arm and Zw3. After
Arm has escaped from these negative regulators, it can act in the nucleus