title: Differential degradation of ferritin H- and L-chains: accumulation of L-chain-rich ferritin in lens epithelial cells.
abstract: OBJECTIVE
  The storage of iron by ferritin is determined by tissue-specific composition of its 24 subunits, which are designated as either heavy (H) or light (L). For a better understanding of how lens epithelial cells regulate their ferritin subunit makeup, the degradation pattern of each subunit type was analyzed. In addition, age-related changes in ferritin concentration and subunit makeup were determined.
  
  
  METHODS
  Ferritin turnover in primary cultures of canine lens epithelial cells was determined by metabolic labeling with [(35)S]-methionine. Transient transfection with vectors containing coding sequences for either H- or L-chains were used to modify ferritin subunit makeup. Ferritin concentration was measured by ELISA. Immunodetection and fluorescence immunocytochemistry were used to study age-related changes in ferritin chain concentration.
  
  
  RESULTS
  Inhibition of the proteasomal protein degradation pathway by clastolactacystin-beta-lactone had no effect on ferritin degradation, whereas inhibition of lysosomal degradation markedly increased ferritin levels, confirming that this system is involved in ferritin turnover. H-chain ferritin degraded at a faster rate than the L-chain. L-chain-rich ferritin in L-chain-transfected cells formed inclusion bodies that were localized to the cytosol. Similar inclusion bodies were found in older lens cells kept in cell culture for more than 8 days.
  
  
  CONCLUSIONS
  Steady degradation of H-chain ferritin contributed to the maintenance of a constant level of this chain within the lens epithelial cells. In contrast, slower turnover of the L-chain resulted in accumulation of L-chain-enriched ferritin associated with cytoplasmic inclusion bodies. These L-chain-containing inclusion bodies were found in the cytosol of cells overexpressing L-ferritin chain and in nontransfected cells maintained in culture for 8 to 35 days. Overexpression of the L-chain has been associated with the formation of premature cataracts in humans with hereditary hyperferritinemia cataract syndrome. The formation of inclusion bodies in older lens epithelial cells, as demonstrated in the current investigation, is intriguing and could point to possible involvement of cytoplasmic L-chain-enriched ferritin aggregates in the formation of age-related cataract.
authors.affiliation: Department of Molecular Biomedical Sciences, North Carolina State University, Raleigh, 27606, USA. margaret_goralska@ncsu.edu
authors.forename: Malgorzata
authors.initials: M
authors.lastname: Goralska
authors.forename: Steven
authors.initials: S
authors.lastname: Nagar
authors.forename: Lloyd N
authors.initials: LN
authors.lastname: Fleisher
authors.forename: M Christine
authors.initials: MC
authors.lastname: McGahan
mesh_terms: D000375:Aging / Q000502:physiology; D000818:Animals; D001052:Apoferritins; D002478:Cells, Cultured; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D005293:Ferritins / Q000378:metabolism*; D007501:Iron / Q000378:metabolism; D007908:Lens, Crystalline / Q000187:drug effects / Q000378:metabolism*; D008247:Lysosomes / Q000378:metabolism; D008715:Methionine / Q000378:metabolism; D010957:Plasmids; D014162:Transfection
chemical_list: D015853:Cysteine Proteinase Inhibitors; D005293:Ferritins; D001052:Apoferritins; D008715:Methionine; D007501:Iron