Phosphoinositides and Membrane Targeting in Cell Polarity

Abstract

Selective enrichment of the polyphosphoinositides (PPIn), such as PtdIns(4,5)P₂ and PtdIns4P, helps to determine the identity of the plasma membrane (PM) and regulates many aspects of cell biology through a vast number of protein effectors. Polarity proteins had long been assumed to be non-PPIn-binding proteins that mainly associate with PM/cell cortex through their extensive protein-protein interaction network. However, recent studies began to reveal that several key polarity proteins electrostatically bind to PPIn through their positively charged protein domains or structures and such PPIn-binding property is essential for their direct and specific attachment to PM. Although the physical nature of the charge-based PPIn binding appears to be simple and nonspecific, it serves as an elegant mechanism that can be efficiently and specifically regulated for achieving polarized PM targeting of polarity proteins. As an unexpected consequence, subcellular localization of PPIn-binding polarity proteins are also subject to regulations by physiological conditions such as hypoxia and ischemia that acutely and reversibly depletes PPIn from PM.

Figure 1.

Generation of plasma membrane (PM) phosphoinositides. The lipid phosphatidylinositol (PtdIns) may be phosphorylated at any of three hydroxyl groups on its cytosol-exposed six-membered polyol headgroup; the major plasma membrane phosphoinositides and their synthetic pathways are indicated.

Figure 2.

Polyphosphoinositides (PPIn)-binding polarity proteins. (A) Subcellular localization of key polarity proteins in epithelial cells and in dividing neuroblasts. Both cells were drawn apical side up. Only polarity proteins relevant to this review were drawn. In Drosophila epithelial cells, aPKC/Par-6 complex and Crb/Sdt(Pals1) complex localize to the apical membrane, and Bazooka (Baz) mostly localizes to apical adherens junction (AJ). Lgl and Par-1 localize to the basolateral plasma membrane (PM). In dividing neuroblasts, aPKC, Par-6, and Baz localize exclusively at the apical PM, with Numb and Miranda (Mir) at basal PM. (B) The conserved polybasic domain in Lgl. Shown at bottom right is the structure of Sro7, the yeast homolog of Lgl, which contains 14 WD40 blades (β-sheets are in green) organized in two β-propellers (Hattendorf et al. 2007). Blades 10 and 11 are enlarged at bottom left. The loop is in magenta. Shown at the top is the alignment of the loop regions between WD40 10D and 11A β-sheets in Lgl/Sro7. Positively charged Lys and Arg residues within the loop sequences are in red and polybasic domains are boxed, Trp/Phe/Leu residues are in green and aPKC-phosphorylatable serines are in blue. Ser residues were numbered based on Drosophila Lgl (dLgl) isoform A (gi24464586). Sequence alignments are modified from Hattendorf et al. (2007). (C) PPIn-binding domains in Numb, Miranda, Baz, and Par-1. PPIn-binding domains are in blue, except that in Baz the carboxyl-terminus regions that bind PPIn are indicated by blue lines. Blue box in Baz indicates a conserved motif required for Baz PM localization in the absence of the amino terminus (Krahn et al. 2010b). Published phosphorylations sites are indicated by red vertical bars. Phosphorylation sites that regulate the polybasic domain-mediated PM targeting are in blue and those are conserved between Drosophila and mammalian Numb proteins are in bold. AJ, Adherens junction; PB, polybasic domain; PTB, phosphotyrosine-binding domain; UBA, ubiquitin-association domain; CLD, cortical localization domain; CargoBD, cargo-binding domain (Atwood and Prehoda 2009); OLG, oligomerization domain (Benton and St Johnston 2003a).

Figure 3.

Regulation of polyphosphoinositides (PPIn) binding in polarity proteins. (A) Three potential mechanisms that may regulate the plasma membrane (PM)-targeting of PPIn-binding polarity proteins. Events depicted in allosteric regulation and structural masking of PPIn-binding domain are hypothetical. (B) Hypoxia induces ATP reduction, loss of PPIn in PM and loss of PM binding of Lgl and Numb. Lgl and Numb are the only two that are experimentally confirmed with loss of PM targeting under hypoxia. Baz remains on PM under hypoxia, likely through its interaction with protein complexes such as adherence junctions (AJs). PB, Polybasic domain.