Phosphatidylserine (PS) is a relatively minor constituent of biological membranes. Despite its low abundance, PS in the plasma membrane (PM) plays key roles in various phenomena such as the coagulation cascade, clearance of apoptotic cells, and recruitment of signaling molecules. PS also localizes in endocytic organelles, but how this relates to its cellular functions remains unknown. Here we report that PS is essential for retrograde membrane traffic at recycling endosomes (REs). PS was most concentrated in REs among intracellular organelles, and evectin-2 (evt-2), a protein of previously unknown function, was targeted to REs by the binding of its pleckstrin homology (PH) domain to PS. X-ray analysis supported the specificity of the binding of PS to the PH domain. Depletion of evt-2 or masking of intracellular PS suppressed membrane traffic from REs to the Golgi. These findings uncover the molecular basis that controls the RE-to-Golgi transport and identify a unique PH domain that specifically recognizes PS but not polyphosphoinositides. cholera toxin | endocytosis

The newly identified SYLF lipid-binding domain of SH3YL1 mediates phosphoinositide binding during dorsal membrane morphogenesis.

In most eukaryotes, phosphoinositides (PIs) have crucial roles in multiple cellular functions. Although the cellular levels of phosphatidylinositol 5-phosphate (PI5P) and phosphatidylinositol 3,5-bisphosphate (PI(3,5)P2) are extremely low relative to some other PIs, emerging evidence demonstrates that both lipids are crucial for the endocytic pathway, intracellular signaling, and adaptation to stress. Mutations that causes defects in the biosynthesis of PI5P and PI(3,5)P2 are linked to human diseases including neurodegenerative disorders. Here, we review recent findings on cellular roles of PI5P and PI(3,5)P2, as well as the pathophysiological importance of these lipids.

Autophagy is a multistep membrane traffic pathway. In contrast to autophagosome formation, the mechanisms underlying autophagosome-lysosome fusion remain largely unknown. Here, we describe a novel autophagy regulator, inositol polyphosphate-5-phosphatase E (INPP5E), involved in autophagosome-lysosome fusion process. In neuronal cells, INPP5E knockdown strongly inhibited autophagy by impairing the fusion step. A fraction of INPP5E is localized to lysosomes, and its membrane anchoring and enzymatic activity are necessary for autophagy. INPP5E decreases lysosomal phosphatidylinositol 3,5-bisphosphate (PI(3,5)P 2 ), one of the substrates of the phosphatase, that counteracts cortactinmediated actin filament stabilization on lysosomes. Lysosomes require actin filaments on their surface for fusing with autophagosomes. INPP5E is one of the genes responsible for Joubert syndrome, a rare brain abnormality, and mutations found in patients with this disease caused defects in autophagy. Taken together, our data reveal a novel role of phosphoinositide on lysosomes and an association between autophagy and neuronal disease.

Downregulation of cell-cell adhesion and upregulation of cell migration play critical roles in the conversion of benign tumors to aggressive invasive cancers. In this study, we show that changes in cell-cell adhesion and cancer cell migration/invasion capacity depend on the level of phosphatidylinositol 4-phosphate [PI(4)P] in the Golgi apparatus in breast cancer cells. Attenuating SAC1, a PI(4)P phosphatase localized in the Golgi apparatus, resulted in decreased cell-cell adhesion and increased cell migration in weakly invasive cells. In contrast, silencing phosphatidylinositol 4-kinase IIIb, which generates PI(4)P in the Golgi apparatus, increased cell-cell adhesion and decreased invasion in highly invasive cells. Furthermore, a PI(4)P effector, Golgi phosphoprotein 3, was found to be involved in the generation of these phenotypes in a manner that depends on its PI(4)P-binding ability. Our results provide a new model for breast cancer cell progression in which progression is controlled by PI(4)P levels in the Golgi apparatus. Cancer Res; 74(11); 3054-66. Ó2014 AACR.

Phosphatidic acid (PA), which can be produced by phospholipase D (PLD), is involved in various signaling events, such as cell proliferation, survival, and migration. However, the molecular mechanisms that link PA to cell migration are largely unknown. Here, we show that PA binds to the tyrosine kinase Fer and enhances its ability to phosphorylate cortactin, a protein that promotes actin polymerization. We found that a previously unknown lipid-binding module in Fer adjacent to the F-BAR [Fes-Cdc42-interacting protein 4 (CIP4) homology (FCH) and bin-amphiphysin-Rvs] domain mediated PA binding. We refer to this lipid-binding domain as the FX (F-BAR extension) domain. Overexpression of Fer enhanced lamellipodia formation and cell migration in a manner dependent on PLD activity and the PA-FX interaction. Thus, the PLD-PA pathway promotes cell migration through Fer-induced enhancement of actin polymerization.

The discovery of the causative gene for Huntington’s disease (HD) has promoted numerous efforts to uncover cellular pathways that lower levels of mutant huntingtin protein (mHtt) and potentially forestall the appearance of HD-related neurological defects. Using a cell-based model of pathogenic huntingtin expression, we identified a class of compounds that protect cells through selective inhibition of a lipid kinase, PIP4Kγ. Pharmacological inhibition or knock-down of PIP4Kγ modulates the equilibrium between phosphatidylinositide (PI) species within the cell and increases basal autophagy, reducing the total amount of mHtt protein in human patient fibroblasts and aggregates in neurons. In two Drosophila models of Huntington’s disease, genetic knockdown of PIP4K ameliorated neuronal dysfunction and degeneration as assessed using motor performance and retinal degeneration assays respectively. Together, these results suggest that PIP4Kγ is a druggable target whose inhibition enhances productive autophagy and mHtt proteolysis, revealing a useful pharmacological point of intervention for the treatment of Huntington’s disease, and potentially for other neurodegenerative disorders.

Growth factor stimulations induce dynamic changes in the cytoskeleton beneath the plasma membrane. Among them is the formation of membrane ruffles organized in a circular array, called 'circular dorsal ruffles' (CDRs). Physiological functions of CDRs include downregulation of cell growth by desensitizing the signalling from growth factor receptors as well as rearrangement of adhesion sites at the onset of cell migration. For the formation of CDRs, not only the activators of actin polymerization, such as N-WASP and the Arp2/3-complex, but also membrane deforming proteins with BAR/F-BAR domains are necessary. Small GTPases are also involved in the formation of CDRs by controlling intracellular trafficking through endosomes. Moreover, recent analyses of another circular cytoskeletal structure, podosome rosettes, have revealed common molecular features shared with CDRs. Among them, the roles of PI3-kinase and phosphoinositide 5-phosphatase may hold the key to the induction of these circular structures.