Deletion of cysteine cathepsins B or L yields differential impacts on murine skin proteome and degradome

Mol Cell Proteomics. 2013 Mar;12(3):611-25. doi: 10.1074/mcp.M112.017962. Epub 2012 Dec 10.

Abstract

Numerous studies highlight the fact that concerted proteolysis is essential for skin morphology and function. The cysteine protease cathepsin L (Ctsl) has been implicated in epidermal proliferation and desquamation, as well as in hair cycle regulation. In stark contrast, mice deficient in cathepsin B (Ctsb) do not display an overt skin phenotype. To understand the systematic consequences of deleting Ctsb or Ctsl, we determined the protein abundances of >1300 proteins and proteolytic cleavage events in skin samples of wild-type, Ctsb(-/-), and Ctsl(-/-) mice via mass-spectrometry-based proteomics. Both protease deficiencies revealed distinct quantitative changes in proteome composition. Ctsl(-/-) skin revealed increased levels of the cysteine protease inhibitors cystatin B and cystatin M/E, increased cathepsin D, and an accumulation of the extracellular glycoprotein periostin. Immunohistochemistry located periostin predominantly in the hypodermal connective tissue of Ctsl(-/-) skin. The proteomic identification of proteolytic cleavage sites within skin proteins revealed numerous processing sites that are underrepresented in Ctsl(-/-) or Ctsb(-/-) samples. Notably, few of the affected cleavage sites shared the canonical Ctsl or Ctsb specificity, providing further evidence of a complex proteolytic network in the skin. Novel processing sites in proteins such as dermokine and Notch-1 were detected. Simultaneous analysis of acetylated protein N termini showed prototypical mammalian N-alpha acetylation. These results illustrate an influence of both Ctsb and Ctsl on the murine skin proteome and degradome, with the phenotypic consequences of the absence of either protease differing considerably.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blotting, Western
  • Cathepsin B / deficiency*
  • Cathepsin B / genetics
  • Cathepsin L / deficiency*
  • Cathepsin L / genetics
  • Cell Adhesion Molecules / metabolism
  • Cells, Cultured
  • Chromatography, Liquid
  • Cystatin B / metabolism
  • Cystatin M / metabolism
  • Immunohistochemistry
  • Mice
  • Mice, Knockout
  • Peptides / metabolism
  • Proteolysis
  • Proteome / metabolism*
  • Proteomics / methods*
  • Receptor, Notch1 / metabolism
  • Serpins / metabolism
  • Skin / metabolism*
  • Tandem Mass Spectrometry

Substances

  • Cell Adhesion Molecules
  • Cystatin M
  • Peptides
  • Postn protein, mouse
  • Proteome
  • Receptor, Notch1
  • Serpins
  • Cystatin B
  • Cathepsin B
  • Ctsb protein, mouse
  • Cathepsin L
  • Ctsl protein, mouse