Mannose 6-phosphate-independent Lysosomal Sorting of LIMP-2

Judith Blanz, Friederike Zunke, Sandra Markmann, Markus Damme, Thomas Braulke, Paul Saftig*, Michael Schwake

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

18 Scopus citations

Abstract

The widely accepted view that β-glucocerebrosidase (GC) is targeted to lysosomes by the lysosomal integral membrane protein type 2 (LIMP-2) independent of the mannose 6-phosphate (M6P) pathway has been recently challenged by the identification of a putative M6P residue in the crystalized LIMP-2 ectodomain. We demonstrate that LIMP-2 and GC reach lysosomes independent of the M6P pathway in fibroblasts and in purified liver lysosomes. Additionally, LIMP-2 could not be affinity-purified using M6P-specific antibodies. Our data prove M6P-independent lysosomal sorting of LIMP-2 and GC. The lysosomal integral membrane protein type 2 (LIMP-2/SCARB2) has been described as a mannose 6-phosphate (M6P)-independent trafficking receptor for β-glucocerebrosidase (GC). Recently, a putative M6P residue in a crystal structure of a recombinantly expressed LIMP-2 ectodomain has been reported. Based on surface plasmon resonance and fluorescence lifetime imaging analyses, it was suggested that the interaction of soluble LIMP-2 with the cation-independent M6P receptor (MPR) results in M6P-dependent targeting of LIMP-2 to lysosomes. As the physiological relevance of this observation was not addressed, we investigated M6P-dependent delivery of LIMP-2 to lysosomes in murine liver and mouse embryonic fibroblasts. We demonstrate that LIMP-2 and GC reach lysosomes independent of the M6P pathway. In fibroblasts lacking either MPRs or the M6P-forming N-acetylglucosamine (GlcNAc)-1-phosphotransferase, LIMP-2 still localizes to lysosomes. Immunoblot analyses also revealed comparable LIMP-2 levels within lysosomes purified from liver of wild-type (wt) and GlcNAc-1-phosphotransferase-defective mice. Heterologous expression of the luminal domain of LIMP-2 in wild-type, LIMP-2-deficient and GlcNAc-1-phosphotransferase-defective cells further established that the M6P modification is dispensable for lysosomal sorting of LIMP-2. Finally, cathepsin Z, a known GlcNAc-1-phosphotransferase substrate, but not LIMP-2, could be precipitated with M6P-specific antibodies. These data prove M6P-independent lysosomal sorting of LIMP-2 and subsequently GC in vivo.

Original languageEnglish (US)
Pages (from-to)1127-1136
Number of pages10
JournalTraffic
Volume16
Issue number10
DOIs
StatePublished - Oct 1 2015

Keywords

  • Gaucher disease
  • LIMP-2/SCARB2
  • Mannose 6-phosphate
  • Mannose 6-phosphate receptor
  • β-glucocerebrosidase

ASJC Scopus subject areas

  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

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