Purification of Drosophila hsp 83 and immunoelectron microscopic localization.
Heat-shock protein (hsp) 83 was purified from Drosophila culture cells. Analysis by gel filtration revealed that this hsp exists in a dimeric form under nondenaturing conditions. Monoclonal and polyclonal antibodies produced against this hsp have been used to determine its intracellular localization by indirect immunofluorescence and immunogold electron microscopy in normal cells, after heat shock, during recovery and after a second heat shock. Under normal conditions, hsp 83 is predominantly cytoplasmic. Immunogold labeling reveals that this hsp is associated with vacuole-like structures containing numerous dense bodies. In addition, hsp 83 is detected, albeit at a lower level, in the nucleus where it is found within the network of perichromatin ribonucleoprotein (RNP) fibrils. This distribution changes during heat shock: hsp 83 is then found in increased concentrations at the cell periphery close to the plasma membrane. After a recovery period, hsp 83 appears associated with the nuclear membrane and/or with the neighboring endoplasmic reticulum. Following a second heat shock at 37 degrees C after recovery, a renewed deposition of hsp 83 is observed at the cell periphery. A small population of cells also shows an increased concentration of this protein in the nucleus. This intracellular distribution of hsp 83 is consistent with its reported association with various cellular proteins and suggest that this hsp may be involved in their intracellular transport and/or in the modulation of their activity.
Centre de Recherche du CHUL, Centre Hospitalier Université Laval, Sainte-Foy, Québec, Canada.
M E
ME
Carbajal
J P
JP
Valet
P M
PM
Charest
R M
RM
Tanguay
D000818:Animals; D000911:Antibodies, Monoclonal; D002478:Cells, Cultured; D002850:Chromatography, Gel; D004330:Drosophila / Q000032:analysis*; D005455:Fluorescent Antibody Technique; D006360:Heat-Shock Proteins / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification*; D015151:Immunoblotting; D007150:Immunohistochemistry; D008854:Microscopy, Electron
D000911:Antibodies, Monoclonal; D006360:Heat-Shock Proteins