Phosphorylation of heat shock protein-90 by TSH in FRTL-5 thyroid cells.
BACKGROUND
Although it is well established that thyrotropin (TSH) initiates signal transduction systems resulting in protein kinase(s) activation, the phosphorylated targets have not been fully characterized.


OBJECTIVE
In FRTL-5 thyroid cells, we used two-dimensional (2D) gel images of silver-stained proteins isolated from FRTL- 5 thyroid cells following TSH stimulation to identify potential phosphorylation targets.


RESULTS
We characterized a 90 kDa protein that had undergone a pH shift and subsequently identified it as heat shock protein-90 (hsp-90) following in-gel trypsin digestion and mass spectroscopy. This was confirmed by Western blot using a monoclonal antibody against hsp-90. Western blot analysis of the 2D gel images using a polyclonal antibody directed at phosphoserine/threonine sites showed that TSH induced the phosphorylation of hsp-90. Western blotting of hsp-90 following stimulators of the signal transduction systems mediated by TSH indicated that TSH-mediated hsp-90 phosphorylation occurs through protein kinases A and C.


CONCLUSIONS
In summary, we have demonstrated that TSH action stimulates the phosphorylation of hsp-90 in FRTL-5 thyroid cells. Abnormalities of hsp-90 phosphorylation may be a mediator in the development of thyroid disease.
Ginsberg
Jody
J
Division of Endocrinology and Metabolism, Department of Medicine, University of Alberta, Edmonton, AB, Canada.
Labedz
Teresa
T
Brindley
David N
DN
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D015180:Electrophoresis, Gel, Two-Dimensional; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010766:Phosphorylation; D051381:Rats; D011916:Rats, Inbred F344; D015398:Signal Transduction; D016622:Silver Staining; D013961:Thyroid Gland / Q000378:metabolism*; D013972:Thyrotropin / Q000378:metabolism*
D018841:HSP90 Heat-Shock Proteins; D013972:Thyrotropin