Determination of cleavage site of Reelin between its sixth and seventh repeat and contribution of meprin metalloproteases to the cleavage

J Biochem. 2016 Mar;159(3):305-12. doi: 10.1093/jb/mvv102. Epub 2015 Oct 21.

Abstract

Reelin is a secreted glycoprotein whose function is regulated by proteolysis. One of the specific cleavage sites of Reelin, called C-t, is located approximately between the sixth and seventh Reelin repeat but its exact site was unknown. We here show that a metalloprotease present in the culture supernatant of cerebellar granular neurons (CGN) cleaves Reelin between Ala2688 and Asp2689. A Reelin mutant in which Asp2689 is replaced by Lys (Reelin-DK) is resistant to C-t cleavage by culture supernatant of CGN. From biochemical characteristics and the cleavage site preference, meprin α and meprin β were suggested candidate proteases and both were confirmed to cleave Reelin at the C-t site. Meprin α cleaved Reelin-DK but meprin β did not. Actinonin, a meprin α and meprin β inhibitor, did not inhibit the Reelin-cleaving activity of CGN and the amount of Reelin fragments in brains of meprin β knock-out mice was not significantly different from that of the wild-type, indicating that meprin β does not play a major role in Reelin cleavage under basal conditions. We propose that meprin α and meprin β join the modulators of Reelin signalling as they cleave Reelin at a specific site and are upregulated under specific pathological conditions.

Keywords: Reelin; meprin; neuron; protease; protein degradation.

MeSH terms

  • Animals
  • COS Cells
  • Cell Adhesion Molecules, Neuronal / metabolism*
  • Cell Culture Techniques
  • Cerebral Cortex / cytology*
  • Chlorocebus aethiops
  • Extracellular Matrix Proteins / metabolism*
  • HEK293 Cells
  • Humans
  • Hydroxamic Acids / pharmacology
  • Metalloendopeptidases / antagonists & inhibitors
  • Metalloendopeptidases / genetics
  • Metalloendopeptidases / metabolism*
  • Mice
  • Mice, Knockout
  • Nerve Tissue Proteins / metabolism*
  • Neurons / enzymology*
  • Proteolysis*
  • Reelin Protein
  • Serine Endopeptidases / metabolism*
  • Signal Transduction

Substances

  • Cell Adhesion Molecules, Neuronal
  • Extracellular Matrix Proteins
  • Hydroxamic Acids
  • Nerve Tissue Proteins
  • Reelin Protein
  • RELN protein, human
  • Reln protein, mouse
  • Serine Endopeptidases
  • Metalloendopeptidases
  • meprin A
  • actinonin