Decreases in Raf-1 levels in galactosaemic lens epithelial cells are partially reversed by myo-inositol.
Changes in the amounts and localization of protein kinase C (PKC) isoforms occur in galactosaemic lens epithelial cells. A link between PKC changes and myo-inositol depletion has been suggested. Raf-1, a component of a Ras pathway, is a substrate for PKC. Raf-1 levels were measured in galactosaemic lens epithelial cells grown with or without myo-inositol. Raf-1 levels were measured by densitometric scanning of Western blots from cells grown with or without 40 mmol/l galactose or 40 mmol/l galactose plus 1.0 micromol/l myo-inositol for 1, 3, 5 or 7 days. Scans were compared to those for PKCalpha, an isoform of PKC and to 14-3-3, a protein which binds to Raf-1. Cell growth was quantitated by thymidine incorporation. Raf-1 levels were decreased in bovine lens epithelial cells after 3, 5 or 7 days (33% of control) of growth in 40 mmol/l galactose. Addition of 1 micromol/l myo-inositol reversed this decrease at day 3, but not after 5 or 7 days of growth in 40 mmol/l galactose. PKCalpha and 14-3-3 levels were not affected by galactose. The decrease in Raf-1 was not a result of cell growth as measured by thymidine incorporation. These results suggest that Raf-1 levels are decreased during galactosaemia. This was only partially reversed by the addition of myoinositol.
Takemoto
D J
DJ
Department of Biochemistry, Kansas State University, Manhattan 66506, USA.
D000818:Animals; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005690:Galactose / Q000494:pharmacology*; D005693:Galactosemias / Q000378:metabolism*; D007294:Inositol / Q000494:pharmacology*; D007527:Isoenzymes / Q000378:metabolism*; D007908:Lens, Crystalline / Q000166:cytology / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*
D007527:Isoenzymes; D007294:Inositol; D004247:DNA; D019908:Proto-Oncogene Proteins c-raf; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D005690:Galactose