Examination of the contribution of vacuolar proteases to intracellular protein degradation in Chara corallina.
The contribution of proteases in the central vacuole of Chara corallina internodal cells to overall cellular protein degradation was examined. I measured the decrease in the trichloroacetic acid (TCA)-precipitable radioactivity in the cell for a 6-d chase period after labeling cellular proteins with [3H]leucine. The kinetics of [3H]leucine-labeled protein disappearance showed that the half-life of the cellular soluble proteins was 4 to 5 d. This value did not change when cells were treated with (2S,3S)-trans-epoxysuccinyl-L-leucylamido- 3-methyl-butane ethyl ester, a permeant inhibitor of cysteine proteases. This inhibitor mostly inhibited bovine serum albumin-degrading activity in the vacuole. I also measured the release of TCA-soluble radioactivity from the TCA-insoluble fraction in the cell. This experiment showed that 13% of [3H]leucine-labeled cellular proteins were degraded in 1 d. This value agreed well with the half-life obtained for soluble proteins in the above experiment. This value did not change even when both trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane, a cysteine protease inhibitor, and pepstatin A, an aspartic protease inhibitor, were introduced into the vacuole. With this operation, bovine serum albumin-degrading activity in the vacuole was almost completely inhibited. These data suggest that the cytoplasmic but not the vacuolar proteases contribute to cellular protein turnover in Chara internodal cells.
Moriyasu
Y
Y
Department of Biology, Faculty of International Relations, University of Shizuoka, Japan.
D000818:Animals; D002417:Cattle; D000460:Chlorophyta / Q000201:enzymology / Q000378:metabolism*; D002738:Chloroquine / Q000494:pharmacology; D010450:Endopeptidases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007930:Leucine / Q000378:metabolism; D008433:Mathematics; D008962:Models, Theoretical; D010940:Plant Proteins / Q000096:biosynthesis / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology; D012710:Serum Albumin, Bovine / Q000378:metabolism; D014617:Vacuoles / Q000187:drug effects / Q000201:enzymology*
D010940:Plant Proteins; D011480:Protease Inhibitors; D012710:Serum Albumin, Bovine; D002738:Chloroquine; D010450:Endopeptidases; D007930:Leucine