Phosphorylation and supramolecular organization of murine small heat shock protein HSP25 abolish its actin polymerization-inhibiting activity.
Characteristic features of mammalian small heat shock proteins are their rapid phosphorylation in response to stress and mitogenic signals and their ability to form multimeric particles of 200-700 kDa and large aggregates up to 5000 kDa. Recently, a chaperoning function and an actin polymerization-inhibiting activity were demonstrated for the recombinant murine and turkey small heat shock protein, respectively. In this paper, we demonstrate that the actin polymerization-inhibiting activity of the murine small heat shock protein HSP25 is dependent on the degree of its phosphorylation and structural organization. Non-phosphorylated and phosphorylated HSP25 monomers, as well as non-phosphorylated multimeric HSP25 particles, were isolated from Ehrlich ascites tumor cells by ammonium sulfate precipitation, column chromatography, and ultracentrifugation and tested for their actin polymerization-inhibiting activity. Fluorescence spectroscopy and electron microscopy were used to monitor actin polymerization. Non-phosphorylated HSP25 monomers were active in inhibiting actin polymerization with about 90% inhibition at a 1:1 ratio of actin to HSP25, while phosphorylated HSP25 monomers and non-phosphorylated multimeric HSP25 particles were inactive. Furthermore, we present electron microscopic data on the structure of HSP25 particles.
Benndorf
R
R
Max Delbrück Center for Molecular Medicine, Berlin-Buch, Federal Republic of Germany.
Hayess
K
K
Ryazantsev
S
S
Wieske
M
M
Behlke
J
J
Lutsch
G
G
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002286:Carcinoma, Ehrlich Tumor; D002474:Cell-Free System; D006360:Heat-Shock Proteins / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D008854:Microscopy, Electron; D010766:Phosphorylation; D011108:Polymers; D013050:Spectrometry, Fluorescence; D014407:Tumor Cells, Cultured
D000199:Actins; D006360:Heat-Shock Proteins; D011108:Polymers