Diego

Diego (Dgo), a cytoplamsic protein, contains six ankyrin repeat domains.

Cytological map position: 41C1-2. Localises to distal cell junctions in the Drosophila pupal wing.

The discovery of Diego

Diego was first identified in an overexpression screen designed to identify genes whose over-expression resulted in planar polarity defects (Feiguin et. al. 2001). This screen made use of the GAl4/UAS expression system (Rorth, 1996/link). Gal4 was expressed under the control of the apterous promoter (expressed in the thorax and dorsal wing blade). Cells expressing the gene apterous were therefore also expressing Gal4. These flies were crossed to flies containing an 'EP' P element construct, many EP lines were created at different points in the Drosophila genome, and they each contain a UAS sequence that is specifically activated by Gal4. Therfore, when Gal4 is expressed then UAS is activated and this also results in expression of genomic sequence that is immediately downstream of the UAS sequence.

2300 EP linnes were crossed to the Apterous-Gal4 line. Only one was found to result hair defects when overepresed (EP(2)2619). The genomic sequence downstream of that UAS line was found to be homologous to an EST corresponding to clone LD08259 (CG12342)

Feiguin et al. 2001: Expression from EP(2)2619 resulting in planar poalrity defects in the thorax and adult wing (A) Thorax of Ap:GAL4/CyO. (B) Wing of Ap:GAL4/CyO (C) Thorax of Ap:GAL4/CyO; EP(2)2619 (D) Wing of Ap:GAL/CyO;EP(2)2619.

What Diego does

Diego when overexpressed results in the accumulation of the core proteins into large puncta (Feiguin et al. 2001).

Diego protein structure and localiation

Protein struture

diego cDNA predicts a 106 kDa protein containing six ankyrin repeats at its N terminus. BLAST search (in 2001) suggest that diego's closest relative is KIAA0957, a human cDNA of unknown function derived from a brain library, both share sequence inside and outside of the ankyrin repeat regions.

Localisation

In the Drosophila pupal wing, Diego protein localises to distal cell membranes (Das et. al. 2004) co localising with Frizzled and Dishevelled. Diego asymmetry in pupal wing cells is most pronounced at 28-31h after puparium formation. Diego localisation requires the presence of Flamingo and Frizzled at the junctions (Feiguin et al. 2001).

Tools for working with Diego

Diego mutants: The diego380 null mutant (Feiguin 2001) deletes the ATG start codon and the first 2 exons containing the six ankyrin repeats and part of the 3rd exon. The end points of diego380 were additionally mapped by sequencing of PCR products. There is no Diego protein recognised on a western using pupal wings and the Diego antibody was made from a GST-Diego fusion protein comprising 25 kb C-terminal to the ankyrin repeats (Feiguin et al. 2001).