Abstract
Lysosomal integral membrane protein-2 (LIMP-2/SCARB2) contributes to endosomal and lysosomal function. LIMP-2 deficiency is associated with neurological abnormalities and kidney failure and, as an acid glucocerebrosidase receptor, impacts Gaucher and Parkinson’s diseases. Here we report a crystal structure of a LIMP-2 luminal domain dimer with bound cholesterol and phosphatidylcholine. Binding of these lipids alters LIMP-2 from functioning as a glucocerebrosidase-binding monomer toward a dimeric state that preferentially binds anionic phosphatidylserine over neutral phosphatidylcholine. In cellular uptake experiments, LIMP-2 facilitates transport of phospholipids into murine fibroblasts, with a strong substrate preference for phosphatidylserine. Taken together, these biophysical and cellular studies define the structural basis and functional importance of a form of LIMP-2 for lipid trafficking. We propose a model whereby switching between monomeric and dimeric forms allows LIMP-2 to engage distinct binding partners, a mechanism that may be shared by SR-BI and CD36, scavenger receptor proteins highly homologous to LIMP-2.
Highlights
Lysosomal integral membrane protein-2 (LIMP-2/SCARB2) contributes to endosomal and lysosomal function
Two lines of evidence suggest that, like CD36 and SR-B1, LIMP-2 may function in lipid trafficking: (1) overexpressing LIMP-2 results in cholesterol (CLR) containing enlarged hybrid endosome/lysosome compartments[21]; (2) LIMP-2 knockout (KO) mice have a cellular phenotype in the inner ear and ureter reminiscent of an impaired membrane trafficking and tubular proteinemia[22]
By determining a crystal structure of a lipid-bound dimeric form of LIMP-2 luminal domain, and by testing LIMP-2 binding to liposomes using surface plasmon resonance (SPR), negative stain electron microscopy (NS-EM), and dynamic light scattering (DLS), we show that LIMP-2 binds phospholipids much like SR-BI and CD3617
Summary
Lysosomal integral membrane protein-2 (LIMP-2/SCARB2) contributes to endosomal and lysosomal function. LIMP-2 facilitates transport of phospholipids into murine fibroblasts, with a strong substrate preference for phosphatidylserine Taken together, these biophysical and cellular studies define the structural basis and functional importance of a form of LIMP-2 for lipid trafficking. LIMP-2 loss-of-function mutations result in reduced lysosomal GCase activity with consequent accumulation of the GCase substrate glucosylceramide, glucosylsphingosine and the fibril-forming protein α-synuclein, characteristic pathological markers of Gaucher disease and Parkinson’s disease, respectively[11]. CD36 and SR-B1 are primarily receptors for fatty acids (FAs)[13] and high-density lipoproteins (HDL)[14], respectively, but their known ligands have expanded to include collagen[15], low-density lipoproteins (LDL)[16], and phospholipids[17] These proteins can facilitate selective, endocytosis-independent uptake of lipids[14, 18]. A lipid uptake mechanism involving this tunnel will require large conformational changes since the tunnel is largely blocked in these monomeric structures based on our analysis
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