|Budget Amount *help
¥7,200,000 (Direct Cost : ¥7,200,000)
Fiscal Year 1996 : ¥1,400,000 (Direct Cost : ¥1,400,000)
Fiscal Year 1995 : ¥1,300,000 (Direct Cost : ¥1,300,000)
Fiscal Year 1994 : ¥4,500,000 (Direct Cost : ¥4,500,000)
1) We found a new member of the family of alpha crystallin small hsps, which could be copurified with alphaB crystallin and hsp27 from extracts of rat or human skeletal muscle. The primary structure of the new protein, p20, determined by the use of a peptide sequenser, was highly homologous to alphaB crystallin and hsp27 consisting of 162 amino acid residues with the alpha crystallin domain in the carboxy-terminal region. p20 was present at high livels in soleus muscle, smooth muscle and heart, as is hsp27, but p20 was not stress-inducible. A cDNA clone encoding p20 was isolated from a rat soleus cDNA library.
2) We found taht hsp27 exists in extracts of cells and tissues in two forms : an aggregated form (L-hsp27,300-400 kDa) and a dissociated form (S-hsp27, <70 kDa). Dissociation of L-hsp27 is enhanced in cells that were exposed to a phorbol ester, okadaic acid, cytokines, or arsenite, all of which enhance or mimic the phosphorylation of hsp27. These results indicate that the molecular configulation of hsp27 in cells is determined in part by phosphorylationn and dephosphorylation of this protein, We also found that phosphorylation of alphaB crystallin is stimulated under the conditions described above. The physiological significance of phosphorylation of alpha crystallin small hsps is under investigation.
3) We found several facotrs that modeulate the stress-induced expression of hsp27, alphaB crustallin, and hsp70. Stress responses is modulated by the metabolic activity of arachidonic acid cascade, by the activities of protein kinase C and protein kinase A,and by prostaglandins. Expression of alphaB crystallin in C6 glioma cells is selectively enhanced by agents that promote the disassembly of microtubules.