Multiple domains of Stardust differentially mediate localisation of the Crumbs-Stardust complex during photoreceptor development in<i>Drosophila</i>

The zebrafish MAGUK protein Nagie oko is a member of the evolutionarily conserved Crumbs protein complex and functions as a scaffolding protein involved in the stabilization of multi-protein assemblies at the tight junction. During zebrafish embryogenesis, mutations in nagie oko cause defects in both epithelial polarity and cardiac morphogenesis. We used deletion constructs of Nagie oko in functional rescue experiments to define domains essential for cell polarity, maintenance of epithelial integrity and cardiac morphogenesis. Inability of Nagie oko to interact with Crumbs proteins upon deletion of the PDZ domain recreates all aspects of the nagie oko mutant phenotype. Consistent with this observation, apical localization of Nagie oko within the myocardium and neural tube is dependent on Oko meduzy/Crumbs2a. Disruption of direct interactions with Patj or Lin-7, two other members of the Crumbs protein complex, via the bipartite L27 domains produces only partial nagie oko mutant phenotypes and does not impair correct junctional localization of the truncated Nagie oko deletion protein within myocardial cells. Similarly, loss of the evolutionarily conserved region 1 domain, which mediates binding to Par6, causes only a subset of the nagie oko mutant epithelial phenotypes. Finally, deletion of the C-terminus, including the entire guanylate kinase and the SH3 domains, renders the truncated Nagie oko protein inactive and recreates all features of the nagie oko mutant phenotype when tested in functional complementation assays. Our observations reveal a previously unknown diversity of alternative multi-protein assembly compositions of the Crumbs–Nagie-oko and Par6-aPKC protein complexes that are highly dependent on the developmental context.

Section: Nok M520 Genotypingsupporting

confidence: 87%

Divergent polarization mechanisms during vertebrate epithelial development mediated by the Crumbs complex protein Nagie oko

Bit‐Avragim

Hellwig

Rudolph

et al. 2008

“…In Drosophila, the Hook domain of Discs Large is crucial for plasma membrane targeting, whereas particular PDZ domains promote septate junction localization in epithelia and synapse localization in neurons (Hough et al, 1997;Thomas et al, 2000). Similar 'twostep' localization mechanisms have been described for C. elegans and mammalian Discs large (Wu et al, 1998;Lockwood et al, 2008), mammalian PSD-95 (Arnold and Clapham, 1999;Craven et al, 1999;Nonaka et al, 2006;Sturgill et al, 2009), Drosophila Inscuteable and Pins (Knoblich et al, 1999;Yu et al, 2002) and Drosophila Stardust (Bulgakova et al, 2008). Contrasting these mechanisms, no single site in Baz is essential for membrane recruitment in ectodermal cells.…”

Section: Comparisons With Other Scaffold Proteinsmentioning

confidence: 78%

Assembly of Bazooka polarity landmarks through a multifaceted membrane-association mechanism

McKinley

Harris

2012

SummaryEpithelial cell polarity is essential for animal development. The scaffold protein Bazooka (Baz/PAR-3) forms apical polarity landmarks to organize epithelial cells. However, it is unclear how Baz is recruited to the plasma membrane and how this is coupled with downstream effects. Baz contains an oligomerization domain, three PDZ domains, and binding regions for the protein kinase aPKC and phosphoinositide lipids. With a structure-function approach, we dissected the roles of these domains in the localization and function of Baz in the Drosophila embryonic ectoderm. We found that a multifaceted membrane association mechanism localizes Baz to the apical circumference. Although none of the Baz protein domains are essential for cortical localization, we determined that each contributes to cortical anchorage in a specific manner. We propose that the redundancies involved might provide plasticity and robustness to Baz polarity landmarks. We also identified specific downstream effects, including the promotion of epithelial structure, a positive-feedback loop that recruits aPKC, PAR-6 and Crumbs, and a negative-feedback loop that regulates Baz.

“…The small, 37-amino-acid cytoplasmic domain contains two highly conserved regions, the C-terminal PSD-95/Discs-large/ ZO-1 (PDZ)-binding motif ERLI, and a 4.1/ezrin/radixin/moesin (FERM)-binding domain. Drosophila sdt encodes several protein isoforms that are scaffolding proteins of the membrane-associated guanylate kinase (MAGUK) family, members of which are characterised by the presence of two Lin-2/Lin-7 (L27) domains, a PDZ domain, a Src-homology 3 (SH3) domain and a guanylate kinase (GUK) domain (Bachmann et al, 2001;Berger et al, 2007;Bulgakova et al, 2008;Hong et al, 2001). PATJ, which was first described, erroneously, as Discs lost (Dlt) (Bhat et al, 1999), contains four PDZ domains and a single L27 domain at the N-terminus (Pielage et al, 2003).…”

Section: Core Components Of the Crumbs Complex In Drosophilamentioning

confidence: 99%

“…The central component, Sdt, acts as a 'hub' to organise a plasmamembrane-associated protein scaffold via an interaction between its PDZ domain and the C-terminal ERLI motif of Crb (Bachmann et al, 2001;Hong et al, 2001). The two L27 domains of Sdt bind to the L27 domains of PATJ and Lin-7 (Bachmann et al, 2004;Bulgakova et al, 2008;Roh et al, 2002). These core components always colocalise; this has been observed in embryonic epithelia derived from the ectoderm, in follicle epithelial cells (Bachmann et al, 2008;Horne-Badovinac and Bilder, 2008;Tanentzapf et al, 2000), in wing and eye imaginal discs (Bachmann et al, 2004), and in pupal and adult PRCs (Bachmann et al, 2001;Berger et al, 2007;Izaddoost et al, 2002;Johnson et al, 2002;Nam and Choi, 2003;Richard et al, 2006a).…”

Section: Core Components Of the Crumbs Complex In Drosophilamentioning

confidence: 99%

The Crumbs complex: from epithelial-cell polarity to retinal degeneration

Bulgakova

Knust

2009

Self Cite

223

217

The evolutionarily conserved Crumbs protein complex is a key regulator of cell polarity and cell shape in both invertebrates and vertebrates. The important role of this complex in normal cell function is illustrated by the finding that mutations in one of its components, Crumbs, are associated with retinal degeneration in humans, mice and flies. Recent results suggest that the Crumbs complex plays a role in the development of other disease processes that are based on epithelial dysfunction, such as tumorigenesis or the formation of cystic kidneys. Localisation of the complex is restricted to a distinct region of the apical plasma membrane that abuts the zonula adherens in epithelia and photoreceptor cells of invertebrates and vertebrates, including humans. In addition to the core components, a variety of other proteins can be recruited to the complex, depending on the cell type and/or developmental stage. Together with diverse post-transcriptional and posttranslational mechanisms that regulate the individual components, this provides an enormous functional diversity and flexibility of the complex. In this Commentary, we summarise findings concerning the organisation and modification of the Crumbs complex, and the conservation of its constituents from flies to mammals. In addition, we discuss recent results that suggest its participation in various human diseases, including blindness and tumour formation.