Discs Lost, a Novel Multi-PDZ Domain Protein, Establishes and Maintains Epithelial Polarity

Abstract

we reported that the relevant locus for the dre1(discs lost) complementation group is CG12021, based on mapping and complementation data shown in Figure 4 of the paper. CG12021 encodes a homolog of the mammalian PDZ protein Patj. However, based on the most current annotation in FlyBase, the 9-10 kb deficiency (Df(3L)MY10) that was generated to remove the CG12021 transcript affects not just two transcripts (Cdc37 and CG12021) as originally documented, but rather seven genes including vanaso (CG32315) located approximately 5 kb upstream to CG12021 (Figure 1 (below), Fly-Base). Recently, it was indicated to us by Dr. Christian Klambt and his colleagues (Pielage et al., 2003) that dre1, the mutation that we claimed to be responsible for the documented phenotype, is allelic to CG32315, rather than CG12021. We therefore re-evaluated our data and discovered the following: sequencing of the CG12021 gene identified a threonine to serine transition at amino acid 516 as well as a stop codon in CG32315 at nucleotide position 865. Since the mutation in CG12021 is a conservative amino acid change, it may not underlie the loss of imaginal discs observed in dre1. Rather, the dre1 phenotypes we described are likely due to the mutation in CG32315. This is supported by the observation that a critical genomic rescue construct named dlt EHHS (Figure 4 of Bhat et al.) now failed to rescue the dre1 mutation in our assays. This result combined with the other rescue data, the stop codon mutation in CG32315, and the failure of dre1 mutation to complement a vanaso allele (van 04276 ) obtained from the Bloomington Stock Center, clearly implicates CG32315 as causing the lethality associated with dre1. However, we do not exclude the possibility that dre1 phenotypes documented in Figure 6 of Bhat et al. (1999), which we confirmed, may result from a combination of mutations in both genes. Indeed, we also confirmed that overexpression of CG12021 causes a severe loss of epithelial polarity as shown in Figure 7 (Bhat et al., 1999). Hence, based on these observations, the phenotype of CG12021-specific mutations remains to be determined. We also repeated the biochemical interaction between Crb and the protein encoded by CG12021 shown in Figure 3N, lane 2 (Bhat et al., 1999). Although we confirmed that these proteins form a complex in vitro and in vivo (data not shown), the complex formation appears to be mediated mainly by Stardust, as shown by Roh et al. (2002). We have not attempted to repeat the dsRNA interference experiments included in Figure 5 of Bhat et al. (1999) in light of the new data on this genomic region, indicating that the selected dsRNAs affect at least one other gene than CG12021.We regret any confusion that our previous conclusions and interpretations might have created.

The apical transmembrane protein Crumbs functions as a tumor suppressor that regulates Hippo signaling by binding to Expanded

“…The Crb-Ex interactions revealed in the current study is clearly distinct from the Crb-Yurt interaction as the former represents positive interactions. Besides the inhibitory Crb-Yurt interaction, Crb also positively regulates the Crb-Sdt-Patj complex via its PBM (14)(15)(16). How Crb coordinately interacts with the growth-regulatory Hippo pathway and the apical-basal polarity pathway remains to be determined.…”

“…It contains 28 EGF-like and four laminin AG-like repeats in its extracellular domain and a short intracellular domain including a juxtamembrane FERM-binding motif (FBM) and a C-terminal PDZbinding motif (PBM). Through its PBM, Crb forms a complex with the PDZ domain proteins Stardust (Sdt) and Patj (14)(15)(16). In regulating apical-basal polarity in Drosophila embryos, the Crb-Sdt-Patj complex functions together with another apical protein complex containing Bazooka (Baz), Par-6, and aPKC, and these apical complexes are antagonized by a basolateral protein complex containing Scribble (Scrib), Discs large (Dlg), and Lethal giant larvae (Lgl) (17,18).…”

Axonal Ensheathment and Septate Junction Formation in the Peripheral Nervous System ofDrosophila

“…Antibody staining of embryos was performed as described previously (Bhat et al, 1999). Primary antibodies used were rabbit anti-Nrx IV (Baumgartner et al, 1996), guinea pig anti-Cont (Faivre-Sarrailh et al, 2004), rabbit anti-GFP (Invitrogen, San Diego, CA), mouse anti-FAS II (Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA), rabbit anti-␤3 tubulin (Dettman et al, 2001), rat anti-Nrg (our study), and guinea pig anti-Repo (our study).…”

“…5B, green) (Dettman et al, 2001), scolopale cell lumen, which is labeled with anti-Crb (Fig. 5B, red) (Bhat et al, 1999), and the sensory neuron marked with anti-22C10 (Fig. 5B, blue) (Fujita et al, 1982).…”

Rôle des protéines Crumbs dans le contrôle de la morphogenèse des cellules épithéliales et des photorécepteurs

“…Les protéines Sdt et D-PATJ sont des protéines d'échafaudage de complexes moléculaires sousmembranaires. Sdt est une MAGUK (membrane associated guanylate kinase), comme les protéines PSD95 (importante pour l'organisation des synapses neuronales) ou ZO-1 (essentielle pour l'organisation des jonctions serrées des cellules épithéliales) [16]: elle interagit avec Crb par son domaine PDZ (module d'interaction commun aux trois protéines PSD95, Discs large et ZO-1), tandis que D-PATJ, qui possède quatre domaines PDZ mais n'a pas de domaine catalytique [17], interagit avec Sdt par sa partie aminoterminale. Crumbs interagit également avec le cytosquelette apical, nécessaire pour l'architecture et le maintien des zonula adherens [8].…”