No abstract

Drosophila Crumbs (Crb) is required for apical-basal polarity and is an apical determinant in embryonic epithelia. Here, we describe properties of Crb that control the position and integrity of the photoreceptor adherens junction and photosensitive organ, or rhabdomere. In contrast to normal photoreceptor adherens junctions and rhabdomeres, which span the depth of the retina, adherens junctions and rhabdomeres of Crb-deficient photoreceptors initially accumulate at the top of the retina and fail to maintain their integrity as they stretch to the retinal floor. We show that Crb controls localization of the adherens junction through its intracellular domain containing a putative binding site for a protein 4.1 superfamily protein (FERM). Although loss of Crb or overexpression of the FERM binding domain causes mislocalization of adherens junctions, they do not result in a significant loss of photoreceptor polarity. Mutations in CRB1, a human homologue of crb, are associated with photoreceptor degeneration in retinitis pigmentosa 12 (RP12) and Leber congenital amaurosis (LCA). The intracellular domain of CRB1 behaves similarly to its Drosophila counterpart when overexpressed in the fly eye. Our studies may provide clues for mechanisms of photoreceptor degeneration in RP12 and LCA.

Polarization of epithelial cells depends on a hierarchical process whereby specific membrane-associated proteins become targeted to specialized membrane domains. Here, we describe a novel Drosophila protein, Discs Lost (DLT), that plays a crucial role in the polarization of embryonic epithelia during cellular blastoderm formation. At subsequent stages of development, DLT interacts with the apical determinant Crumbs (CRB) and the laterally localized protein Neurexin IV (NRX IV). Mutations in dlt or double-stranded RNA interference lead to aberrant localization of CRB and NRX IV and cause a concomitant loss of epithelial cell polarity. Hence, DLT is required to establish and maintain cell polarity and participates in different molecular complexes that define apical and lateral membrane domains.

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.

Dorsoventral (DV) patterning is essential for growth of the Drosophila eye. Recent studies suggest that ventral is the default state of the early eye, which depends on Lobe (L) function, and that the dorsal fate is established later by the expression of the dorsal selector gene pannier (pnr). However, the mechanisms of regulatory interactions between L and dorsal genes are not well understood. For studying the mechanisms of DV patterning in the early eye disc, we performed a dominant modifier screen to identify additional genes that interact with L. The criterion of the dominant interaction was either enhancement or suppression of the L ventral eye loss phenotype. We identified 48 modifiers that correspond to 16 genes, which include fringe ( fng), a gene involved in ventral eye patterning, and members of both Hedgehog (Hh) and Decapentaplegic (Dpp) signaling pathways, which promote L function in the ventral eye. Interestingly, 29% of the modifiers (6 enhancers and 9 suppressors) identified either are known to interact genetically with pnr or are members of the Wingless (Wg) pathway, which acts downstream from pnr. The detailed analysis of genetic interactions revealed that pnr and L mutually antagonize each other during second instar of larval development to restrict their functional domains in the eye. This time window coincides with the emergence of pnr expression in the eye. Our results suggest that L function is regulated by multiple signaling pathways and that the mutual antagonism between L and dorsal genes is crucial for balanced eye growth.