Intracellular phosphatidylserine is essential for retrograde membrane traffic through endosomes

Abstract

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

Coxiella burnetii effector CvpB modulates phosphoinositide metabolism for optimal vacuole development

“…CvpB Modulates the Intracellular Levels of PI(3)P. Host PIs and their metabolism are known targets of a number of bacterial pathogens; we thus investigated whether Coxiella uses CvpB to manipulate pathways controlled by PS and/or PI(3)P. Besides its role as a signaling lipid during apoptosis, it has been reported that PS plays an important role in retrograde traffic from the plasma membrane to the Golgi complex (22). We therefore transfected U2OS cells with mCherry or CvpB-mCherry and monitored retrograde transport by incubating cells with 1 μg/mL FITC-labeled Cholera toxin B subunit (CtxB).…”

Mitosis-Coupled, Microtubule-Dependent Clustering of Endosomal Vesicles around Centrosomes

“…To confirm that the cluster is indeed derived from the ERC, we exploited a recent observation that phosphatidylserine (PS) is enriched in membranes of the ERC (Uchida et al, 2011); PS was detected using a PS-specific probe, the C2 domain of lactadherin (LactC2). As shown in Fig.…”

“…An expression vector for FLAG-tagged dynamitin (Hirose et al, 2004) was provided by Mitsuo Tagaya (Tokyo University of Pharmacy). A LactC2 cDNA fragment (Uchida et al, 2011) (provided by Hiroyuki Arai, University of Tokyo) was initially subcloned into pcDNA3-mRFP (provided by Roger Tsien). The mRFP-LactC2 cDNA fragment was then amplified by polymerase chain reaction from pcDNA3-mRFP-LactC2 and cloned into the pGEX-6P-1 vector (GE Healthcare).…”

Transport of cholera toxin B-subunit from recycling endosomes to the Golgi requires clathrin and AP-1

“…Two recycling-endosome-localized proteins, evectin-2 (also known as PLEKHB2) and SMAP2, have recently been shown to be essential for the retrograde transport of CTxB at the recycling-endosome-toGolgi step (Matsudaira et al, 2013;Uchida et al, 2011). Evectin-2 contains a pleckstrin homology (PH) domain which specifically binds phosphatidylserine (PtdSer) and this interaction is required for its function and localization to PtdSer-enriched recycling endosomes.…”

“…Recycling endosomes are emerging as an essential sorting station for some retrograde cargo proteins, such as MPR300 (Lin et al, 2004;McKenzie et al, 2012), the Shiga toxin B-subunit (STxB) (McKenzie et al, 2009(McKenzie et al, , 2012, and the cholera toxin B-subunit (CTxB) (Uchida et al, 2011); these proteins traffic sequentially from the plasma membrane to early endosomes, then to recycling endosomes and finally to the Golgi (Taguchi, 2013). Two recycling-endosome-localized proteins, evectin-2 (also known as PLEKHB2) and SMAP2, have recently been shown to be essential for the retrograde transport of CTxB at the recycling-endosome-toGolgi step (Matsudaira et al, 2013;Uchida et al, 2011).…”