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Several AP-1γ-interacting motifs were predicted in the Nak polypeptide, with one located in the N-terminal kinase domain and others distributed in the C-terminal region (arrows and predicted motifs in Fig. 3A). Similarly assay was performed to test the association between Nak and AP-1γ in S2 cells. AP-1γ indeed co-precipitated with the Myc-Nak immunocomplex (line 4 in Fig. 3B, right panel), but not with the C-terminal truncated Myc-Nak-ΔC immunocomplex (lane 3 in Fig. 3B, right panel), indicating that PR-957 the C-terminal region of Nak that includes eleven putative AP-1γ-interacting motifs is required for the interaction with AP-1γ.
Requirement of AP-1 and AP-2 in Dlg basal localization in SG cells
The associations of Nak with α-Ada and AP-1γ prompted us to study whether Dlg basal localization in SG cells depends on AP-2 and AP-1 complexes. In the transheterozygous mutants for the α subunit of AP-2 complex, α-ada1/3, and for the μ1 subunit of AP-1 complex, AP47SHE11/EP1112, the survival larvae allowed us to examine the Dlg localizations in mutant SG cells. Although the cell growth and morphology were severely affected in both mutants ( Supplemental Table 1), Dlg localizations were strongly biased toward the apical membrane, similar to what was observed in nak mutants ( Supplementary Fig. 3).