Cellular processing of myocilin

PLoS One. 2014 Apr 14;9(4):e92845. doi: 10.1371/journal.pone.0092845. eCollection 2014.

Abstract

Background: Myocilin (MYOC) is a gene linked directly to juvenile- and adult-onset open angle glaucoma. Mutations including Pro370Leu (P370L) and Gln368stop (Q368X) have been identified in patients. In the present study, we investigated the processing of myocilin in human trabecular meshwork (TM) cells as well as in inducible, stable RGC5 cell lines.

Methodology/principal findings: The turnover and photoactivation experiments revealed that the endogenous myocilin in human trabecular meshwork (TM) cells was a short-lived protein. It was found that the endogenous myocilin level in TM cells was increased by treatment of lysosomal and proteasomal inhibitors, but not by autophagic inhibitor. Multiple bands immunoreactive to anti-ubiquitin were seen in the myocilin pull down, indicating that myocilin was ubiquitinated. In inducible cell lines, the turnover rate of overexpressed wild-type and mutant P370L and Q368X myocilin-GFP fusion proteins was much prolonged. The proteasome function was compromised and autophagy was induced. A decreased PSMB5 level and an increased level of autophagic marker, LC3, were demonstrated.

Conclusions/significance: The current study provided evidence that in normal homeostatic situation, the turnover of endogenous myocilin involves ubiquitin-proteasome and lysosomal pathways. When myocilin was upregulated or mutated, the ubiquitin-proteasome function is compromised and autophagy is induced. Knowledge of the degradation pathways acting on myocilin can help in design of novel therapeutic strategies for myocilin-related glaucoma.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Autophagy / radiation effects
  • Cell Line
  • Cytoskeletal Proteins / antagonists & inhibitors
  • Cytoskeletal Proteins / metabolism*
  • Eye Proteins / antagonists & inhibitors
  • Eye Proteins / metabolism*
  • Glycoproteins / antagonists & inhibitors
  • Glycoproteins / metabolism*
  • Green Fluorescent Proteins / metabolism
  • Humans
  • Light
  • Microtubule-Associated Proteins / metabolism
  • Mutation / genetics
  • Proteasome Endopeptidase Complex / metabolism
  • Protein Processing, Post-Translational* / radiation effects
  • Recombinant Fusion Proteins / metabolism
  • Trabecular Meshwork / cytology*
  • Trabecular Meshwork / metabolism*
  • Trabecular Meshwork / radiation effects
  • Ubiquitination / radiation effects
  • Young Adult
  • beta Catenin / metabolism

Substances

  • Cytoskeletal Proteins
  • Eye Proteins
  • Glycoproteins
  • MAP1LC3A protein, human
  • Microtubule-Associated Proteins
  • Recombinant Fusion Proteins
  • beta Catenin
  • trabecular meshwork-induced glucocorticoid response protein
  • Green Fluorescent Proteins
  • PSMB5 protein, human
  • Proteasome Endopeptidase Complex