Dlg5 maintains apical polarity by promoting membrane localization of Crumbs during Drosophila oogenesis

Abstract

Apical-basal polarity plays critical roles in the functions of epithelial tissues. However, the mechanisms of epithelial polarity establishment and maintenance remain to be fully elucidated. Here we show that the membrane-associated guanylate kinase (MAGUK) family protein Dlg5 is required for the maintenance of apical polarity of follicle epithelium during Drosophila oogenesis. Dlg5 localizes at the apical membrane and adherens junction (AJ) of follicle epithelium in early stage egg chambers. Specifically, we demonstrate that the major function of Dlg5 is to promote apical membrane localization of Crumbs, since overexpression of Crumbs but not other major apical or AJ components could rescue epithelial polarity defects resulted from loss of Dlg5. Furthermore, we performed a structure-function analysis of Dlg5 and found that the C-terminal PDZ3 and PDZ4 domains are required for all Dlg5’s functions as well as its ability to localize to apical membrane. The N-terminal coiled-coil motif could be individually targeted to the apical membrane, while the central linker region could be targeted to AJ. Lastly, the MAGUK core domains of PDZ4-SH3-GUK could be individually targeted to apical, AJ and basolateral membranes.

A modular toolset of phiC31-based fluorescent protein tagging vectors forDrosophila

“…EGFP, Ubi-Dlg5.TagRFP-T and Ubi-Dlg5.mCherry transgenes can also rescue the lethality of dlg5 mutants ( Table 2), revealing that the FP fused Dlg5 proteins are functional. Dlg5 proteins expressed by these constructs localizes in apical domains of follicle cells and nurse cell membranes in early stage egg chambers (Figure 2(d,h,i)), which is consistent with the previous report [33]. Ubi-Dlg5.EGFP shows ubiquitous expression of Dlg5 in egg chamber, eye disc, wing disc and embryo (Figure 2(d-g)), indicating that the Ubi promoter is functional.…”

“…Dlg5 is an evolutionarily conserved membrane-associated guanylate kinase (MAGUK) family protein and is required for epithelial structure maintenance [32]. We have previously shown Drosophila Dlg5 is required for apical polarity maintenance [33]. The genomic construct Dlg5-TagRFP-T is based on pB2GW vector, containing 9.6kb genomic sequence that spans the whole dlg5 locus (Figure 2(a)).…”

Multiple functions of the scaffold protein Discs large 5 in the control of growth, cell polarity and cell adhesion in Drosophila melanogaster

“…These defects have been detected for instance in the kidney or the lung, where a partial mislocalization of the apical determinant aPKC, a key component of the apical PAR complex, has been observed [10,11]. Moreover, knock-down of Dlg5 in follicle cells also give similar phenotypes [13]. In follicular cells mutant for null mutant Dlg5 Ex13 we saw a semipenetrant reduction of the apical level of aPKC, the apical domain of these cells being inwards, at the contact with the germline (12/19 clones) ( Fig 4A).…”

Multiple functions of the scaffold protein Discs large 5 in the control of growth, cell polarity and cell adhesion in Drosophila melanogaster

“…Moreover, knock-down of Dlg5 in follicle cells also give similar phenotypes [13]. In follicular cells mutant for null mutant Dlg5 Ex13 we saw a semi-penetrant reduction of the apical level of aPKC, the apical domain of these cells being inwards, at the contact with the germline (12/19 clones) ( Fig 4A).…”