title: Overexpression of H- and L-ferritin subunits in lens epithelial cells: Fe metabolism and cellular response to UVB irradiation.
abstract: OBJECTIVE
  To determine the effect of changes in ferritin subunit makeup on Fe metabolism and the resistance of lens epithelial cells (LEC) to photo-oxidative stress.
  
  
  METHODS
  Cultured canine LEC were transiently transfected with pTargeT mammalian expression vector containing the whole coding sequence of H- or L-chain cDNA. The subunit composition of newly synthesized ferritin was analyzed by metabolic labeling and SDS-PAGE electrophoresis. Total ferritin concentration was measured by ELISA: Fe uptake and incorporation into ferritin was determined by incubating transfected cells with (59)Fe-labeled transferrin followed by native PAGE electrophoresis. The effect of UV irradiation was assessed by cell count after exposure of transfected cells to UVB radiation.
  
  
  RESULTS
  Transfected cells differentially expressed H- and L-ferritin chains from cDNA under the control of CMV promoter; overexpression of L-chain was much greater than that of H-chain. The expressed chains assembled into ferritin molecules under in vitro and in vivo condition. The ferritin of H-transfectants incorporated significantly more Fe than those of L-transfectants. The UVB irradiation reduced cell number of L-transfectants by half, whereas H-chain transfectants were protected.
  
  
  CONCLUSIONS
  Post-transfectional expression of ferritin H- and L-chains in LEC appears to be regulated differentially. Overexpression of L-chain ferritin did not have a major effect on cellular Fe distribution and did not protect LEC against UV irradiation, whereas overexpression of H-chain resulted in increased storage of Fe in ferritin and protected cells from UV damage.
authors: Department of Anatomy, Physiology, and Radiology, College of Veterinary Medicine, North Carolina State University, 4700 Hillsborough Street, Raleigh, NC 27606, USA.
authors: M
authors: M
authors: Goralska
authors: B L
authors: BL
authors: Holley
authors: M C
authors: MC
authors: McGahan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001052:Apoferritins; D001483:Base Sequence; D002452:Cell Count; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000378:metabolism; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000378:metabolism / Q000528:radiation effects*; D005293:Ferritins / Q000096:biosynthesis* / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D007501:Iron / Q000378:metabolism*; D007908:Lens, Crystalline / Q000378:metabolism / Q000528:radiation effects*; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D014162:Transfection; D014466:Ultraviolet Rays
chemical_list: D018076:DNA, Complementary; D005293:Ferritins; D001052:Apoferritins; D007501:Iron