Phosphatidylinositol-(4,5)-Bisphosphate Regulates Sorting Signal Recognition by the Clathrin-Associated Adaptor Complex AP2

Höning, Stefan; Ricotta, Doris; Krauß, Michael; Späte, Kira; Spolaore, Barbara; Motley, Alison M.; Robinson, Margaret S.; Robinson, Carol V.; Haucke, Volker; Owen, David J.

doi:10.1016/j.molcel.2005.08.001

Cited by 43 publications

(67 citation statements)

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“…For SPR measurements, liposomes were immobilized on a Pioneer® L1 sensor chip (Biacore), presenting alkyl chains of appropriate length (C 8 – 12 ) that inserted into the lipid bilayer. It has been shown by electron microscopy [41], fluorescence microscopy [42], or by analysing the release of a fluorescent marker from liposomes while loading the chip [43,44], that the sensor chip captures mostly intact liposomes. All SPR measurements were performed at room temperature.…”

Section: Resultsmentioning

confidence: 99%

Saposin B mobilizes lipids from cholesterol‐poor and bis(monoacylglycero)phosphate‐rich membranes at acidic pH

et al. 2007

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Sphingolipid activator proteins (SAPs), GM2 activator protein (GM2AP) and saposins (Saps) A-D are small, enzymatically inactive glycoproteins of the lysosome. Despite of their sequence homology, these lipid-binding and -transfer proteins show different specificities and varying modes of action. Water-soluble SAPs facilitate the degradation of membrane-bound glycosphingolipids with short oligosaccharide chains by exohydrolases at the membrane-water interface. There is strong evidence that degradation of endocytosed components of the cell membrane takes place at intraendosomal and intralysosomal membranes. The inner membranes of the lysosome differ from the limiting membrane of the organelle in some typical ways: the inner vesicular membranes lack a protecting glycocalix, and they are almost free of cholesterol, but rich in bis(monoacylglycero)phosphate (BMP), the anionic marker lipid of lysosomes. In this study, we prepared glycosylated Sap-B free of other Saps by taking advantage of the Pichia pastoris expression system. We used immobilized liposomes as a model for intralysosomal vesicular membranes to probe their interaction with recombinantly expressed Sap-B. We monitored this interaction using SPR spectroscopy and an independent method based on the release of radioactively labelled lipids from liposomal membranes. We show that, after initial binding, Sap-B disturbs the membrane structure and mobilizes the lipids from it. Lipid mobilization is dependent on an acidic pH and the presence of anionic lipids, whereas cholesterol is able to stabilize the liposomes. We also show for the first time that glycosylation of Sap-B is essential to achieve its full lipid-extraction activity. Removal of the carbohydrate moiety of Sap-B reduces its membrane-destabilizing quality. An unglycosylated Sap-B variant, Asn215His, which causes a fatal sphingolipid storage disease, lost the ability to extract membrane lipids at acidic pH in the presence of BMP.Abbreviations BMP, bis(monoacylglycero)phosphate; GM2AP, ganglioside GM2 activator protein; PtdCho, phosphatidylcholine; pSap, Sap precursor; RU, response unit; SAP, sphingolipid activator protein; Sap, saposin.

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Section: Resultsmentioning

confidence: 99%

Saposin B mobilizes lipids from cholesterol‐poor and bis(monoacylglycero)phosphate‐rich membranes at acidic pH

et al. 2007

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“…Phosphatidylinositides (PIPs) are known to confer, at least in part, identities to membrane domains, by regulating the activities of ARF effectors and thus local ARF activation necessary for subsequent coat binding (De Matteis and Godi, 2004;Di Paolo and De Camilli, 2006). For example, phosphatidylinositol 4-phosphate (PI-4P) is required for AP-1 binding (Heldwein et al, 2004;Baust et al, 2006), whereas phosphatidylinositol 4,5-bisphosphate is required for AP-2dependent sorting (Honing et al, 2005). PIPs also provide additional binding sites for APs.…”

Section: Introductionmentioning

confidence: 99%

Protein Networks Supporting AP-3 Function in Targeting Lysosomal Membrane Proteins

Baust

Anitei

Czupalla

et al. 2008

MBoC

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The AP-3 adaptor complex targets selected transmembrane proteins to lysosomes and lysosome-related organelles. We reconstituted its preferred interaction with liposomes containing the ADP ribosylation factor (ARF)-1 guanosine triphosphatase (GTPase), specific cargo tails, and phosphatidylinositol-3 phosphate, and then we performed a proteomic screen to identify new proteins supporting its sorting function. We identified approximately 30 proteins belonging to three networks regulating either AP-3 coat assembly or septin polymerization or Rab7-dependent lysosomal transport. RNA interference shows that, among these proteins, the ARF-1 exchange factor brefeldin A-inhibited exchange factor 1, the ARF-1 GTPase-activating protein 1, the Cdc42-interacting Cdc42 effector protein 4, an effector of septin-polymerizing GTPases, and the phosphatidylinositol-3 kinase IIIC3 are key components regulating the targeting of lysosomal membrane proteins to lysosomes in vivo. This analysis reveals that these proteins, together with AP-3, play an essential role in protein sorting at early endosomes, thereby regulating the integrity of these organelles.

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“…In this still somewhat hypothetical scenario the presence of PI(4,5)P 2 in the plasma membrane appears to shift the equilibrium towards the ‘open’ conformation thereby increasing the affinity for cargo membrane proteins (compare also Fig. 3 , further discussed below) [24]. It is likely that similar mechanisms facilitate cargo recognition by AP‐1 [25], AP‐3, or AP‐4 adaptor complexes at TGN or endosomal membranes.…”

Section: Binding To Phosphoinositides Can Be Associated With Conformamentioning

confidence: 96%

“…Efficient cargo recognition strictly depends on the presentation of endocytic sorting motifs in the context of a PI(4,5)P 2 ‐rich membrane, thus restricting AP‐2 function to the plasmalemma (Fig. 3, center left) [24]. High‐affinity membrane binding of AP‐2 therefore depends on the simultaneous presence of both PI(4,5)P 2 and cargo membrane proteins as parts of a coincidence detection mechanism.…”

Section: Conferring Specificity To the System: Coincidence Detection mentioning

confidence: 99%

Phosphoinositides: Regulators of membrane traffic and protein function

Krauß

Haucke

2007

FEBS Letters

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Phosphoinositides serve as important spatio-temporal regulators of intracellular trafficking and cell signalling events. In addition to their recognition by specific phosphoinositide binding domains present within cytoplasmic adaptor proteins or membrane integral channels and transporters phosphoinositides may affect membrane transport by eliciting conformational changes within proteins or by regulating enzymatic activities. During adaptor-mediated membrane traffic phosphoinositides form part of coincidence detection systems that aid in targeting pools of specific phosphoinositides to select intracellular transport pathways. In this review, we discuss potential mechanisms for conferring selectivity onto the phosphoinositide code as well as possible avenues for future research.

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Phosphatidylinositol-(4,5)-Bisphosphate Regulates Sorting Signal Recognition by the Clathrin-Associated Adaptor Complex AP2

Cited by 43 publications

References 0 publications

Saposin B mobilizes lipids from cholesterol‐poor and bis(monoacylglycero)phosphate‐rich membranes at acidic pH

Saposin B mobilizes lipids from cholesterol‐poor and bis(monoacylglycero)phosphate‐rich membranes at acidic pH

Protein Networks Supporting AP-3 Function in Targeting Lysosomal Membrane Proteins

Phosphoinositides: Regulators of membrane traffic and protein function

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