Cell cycle-specific effects of sodium arsenite and hyperthermic exposure on incorporation of radioactive leucine and phosphate by stress proteins from mouse lymphoma cell nuclei.
Cultured mouse lymphoma cells incorporated [3H]leucine and [32P]phosphate into nuclear stress proteins within 3 h after exposure to either elevated temperature (45 degrees C) or sodium arsenite. Radiolabeled proteins were detected by autoradiography after two-dimensional polyacrylamide gel electrophoresis. To determine the cell cycle stage specificity of labeling, nuclei were isolated and sorted into two cell cycle phases using a fluorescent activated cell sorter. After either heat shock or sodium arsenite treatment, the majority of [3H]leucine incorporation into stress proteins occurred during the G0 + G1 phase with minimal labeling in the G2 phase. On the other hand, 32P labeling of stress proteins occurred in both the G0 + G1 and G2 phases after exposure to sodium arsenite, while incorporation of 32P was limited after heat stress. Following sodium arsenite treatment, a distinct set of four stress proteins (80-84 kDa) was detected with [3H]leucine only in G0 + G1 phase, but with [32P]phosphate these stress proteins were labeled in both G0 + G1 and G2. There was differential [32P]phosphate labeling between proteins of the 80-84 kDa set during cell cycling. Individual proteins of this set were isolated from gel plugs after sodium arsenite or heat-shock treatment. Coelectrophoresis of proteins from the two treatment groups showed that they had similar electrophoretic mobilities. All four proteins of the 80-84 kDa set (sodium arsenite induced) possessed similar polypeptide maps after digestion with V8 protease. Cytofluorometric analysis demonstrated a reduction in the number of nuclei in both S and G2 phases of the cell cycle two h after heat shock, but not following sodium arsenite treatment. However, there was a significant depression in the number of nuclei in S and G2 4 h after exposure to sodium arsenite and very modest labeling with 32P of stress proteins was observed at this time.
D000818:Animals; D001151:Arsenic / Q000494:pharmacology*; D018053:Arsenites; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D006358:Hot Temperature; D007930:Leucine / Q000378:metabolism*; D008223:Lymphoma / Q000378:metabolism*; D051379:Mice; D008970:Molecular Weight; D010710:Phosphates / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D017670:Sodium Compounds; D013439:Sulfhydryl Reagents / Q000494:pharmacology*; D014316:Tritium
D018053:Arsenites; D006360:Heat-Shock Proteins; D010710:Phosphates; D010761:Phosphorus Radioisotopes; D017670:Sodium Compounds; D013439:Sulfhydryl Reagents; D014316:Tritium; C017947:sodium arsenite; D007930:Leucine; D001151:Arsenic