A Novel Function for the PAR Complex in Subcellular Morphogenesis of Tracheal Terminal Cells in Drosophila melanogaster

Abstract

The processes that generate cellular morphology are not well understood. To investigate this problem, we use Drosophila melanogaster tracheal terminal cells, which undergo two distinct morphogenetic processes: subcellular branching morphogenesis and subcellular apical lumen formation. Here we show these processes are regulated by components of the PAR-polarity complex. This complex, composed of the proteins Par-6, Bazooka (Par-3), aPKC, and Cdc42, is best known for roles in asymmetric cell division and apical/basal polarity. We find Par-6, Bazooka, and aPKC, as well as known interactions between them, are required for subcellular branch initiation, but not for branch outgrowth. By analysis of single and double mutants, and isolation of two novel alleles of Par-6, one of which specifically truncates the Par-6 PDZ domain, we conclude that dynamic interactions between apical PAR-complex members control the branching pattern of terminal cells. These data suggest that canonical apical PAR-complex activity is required for subcellular branching morphogenesis. In addition, we find the PAR proteins are downstream of the FGF pathway that controls terminal cell branching. In contrast, we find that while Par-6 and aPKC are both required for subcellular lumen formation, neither Bazooka nor a direct interaction between Par-6 and aPKC is needed for this process. Thus a novel, noncanonical role for the polarity proteins Par-6 and aPKC is used in formation of this subcellular apical compartment. Our results demonstrate that proteins from the PAR complex can be deployed independently within a single cell to control two different morphogenetic processes. FOR most cell types, morphology is key to cell function. A dramatic example of this association is seen in cells that undergo subcellular branching morphogenesis. In this process, cells send out extensions from their plasma membranes, which grow and undergo bifurcation events to form complex, branched networks. Examples of subcellular branching morphogenesis are seen in glial oligodendrocytes (Bauer et al. 2009) and in dendritic cells of the mammalian immune system (Makala and Nagasawa 2002), but by far the best studied examples of this process are in neurons (reviewed by Gibson and Ma 2011;Jan and Jan 2010). Indeed, neurons are frequently categorized entirely by differences in their branching morphologies (see Puelles 2009). However, despite the importance of subcellular branching morphogenesis, little is known about the molecular mechanisms that organize distinctive subcellular branching patterns.We are studying the process of subcellular branching morphogenesis in Drosophila tracheal terminal cells, a component of the insect respiratory system. Terminal cells reside at the ends of a network of cellular tubes that functions in delivering air to internal tissues (Guillemin et al. 1996). The cells are specified during embryogenesis, primarily through a process of competitive FGF signaling and lateral inhibition among tracheal precursors (Llimargas 1999;Ghabrial and Krasno...

Exocyst-mediated membrane trafficking is required for branch outgrowth in Drosophila tracheal terminal cells

“…Genetic screens have identified a number of genes required for terminal cell branching morphogenesis and lumen formation (Baer et al, 2007; Ghabrial et al, 2011; Jones and Metzstein, 2011; Levi et al, 2006). One mechanism identified in these screens involves the activity of the PAR-polarity complex (Par-6, Baz, aPKC, and Cdc42).…”

“…One mechanism identified in these screens involves the activity of the PAR-polarity complex (Par-6, Baz, aPKC, and Cdc42). In terminal cells the PAR complex is required for terminal cell branching but not outgrowth, demonstrating that these two processes can be decoupled (Jones and Metzstein, 2011). Here, we focus on the molecular machinery required for branch outgrowth in terminal cells and identify a role for the exocyst complex in subcellular branch outgrowth.…”

Tube fusion: Making connections in branched tubular networks

“…Apical proteins including Crumbs and Podocalyxin start to accumulate at the PAP, and aPKC and Bazooka are pushed from the PAP to the junctional region [84,87]. Notably, a similar distribution of Bazooka and apical proteins was observed during lumen formation both in tracheal FCs and TCs [22,23,62].…”

“…The spatial distribution of the two different tip cell types is stereotyped and essential for the final architecture and function of the tracheal system. Intriguingly, FCs and TCs form intracellular (seamless) tubes, unlike all other tracheal cells, which enclose extracellular lumina sealed by cell-cell junctions [22][23][24].…”

Examination of <em>Drosophila</em> Larval Tracheal Terminal Cells by Light Microscopy

“…Using this system unbiased, forward genetic screens for cell morphogenesis mutants have been performed, yielding insights into mechanisms controlling cell shape 5,6 . For instance, these screens have revealed that a specific RabGAP is required for cytoskeletal polarity and vesicle trafficking in lumen formation and positioning 7 ; that integrin-mediated adhesion is required for branch stability 8 ; and that epithelial PAR-polarity proteins regulate polarized membrane trafficking required for both branching and lumen formation 9 .…”