In vitro thyroid hormone rapidly modulates protein phosphorylation in cerebrocortical synaptosomes from adult rat brain.
Thyroid hormones induced rapid changes in phosphorylation in a membrane-containing lysate of synaptosomes purified from adult rat cerebral cortex. The in vitro addition of 3,5,3'-L-triiodothyronine or L-thyroxine strongly influenced incorporation of label from [gamma-32P]-ATP into proteins in a cerebrocortical synaptosomal lysate. Incubation with 3,5,3'-L-triiodothyronine or L-thyroxine had strong biphasic dose-dependent effects on the phosphorylation of 38+/-1, 53+/-1, 62+/-1, and 113+/-1 kDa proteins (which we termed alpha, beta, gamma, and delta, respectively) and several others. Although we observed differing levels of phosphorylation among the four proteins, doses of 3,5,3'-L-triiodothyronine or L-thyroxine ranging from 1 to 30 nM caused significant dose-dependent stimulation of the phosphorylation of all of them, an effect which occurred within three minutes. In each case, the enhancement of phosphorylation diminished with higher concentrations (100 nM-1 microM) of 3,5,3'-L-triiodothyronine. In contrast, incubations with similar doses of 3,3',5'-L-triiodothyronine (reverse L-triiodothyronine) were without significant effect, indicating a specificity for 3,5,3'-L-triiodothyronine and L-thyroxine. Western blots of synaptosomal lysates incubated with 3,5,3'-L-triiodothyronine (1 nM-1 microM) demonstrated phosphorylation at the serine residues of a 112 kDa protein (matching delta) and phosphorylation at tyrosyl residues of a distinct 95 kDa protein. These data support the contention that thyroid hormones have a variety of rapid nongenomic pathways for regulation of protein phosphorylation in mature mammalian brain.
Sarkar
P K
PK
Department of Biology, Rutgers University, 315 Penn Street, Camden, NJ 08102-1411, USA. psarkar@nycc.edu
Durga
N D
ND
Morris
J J
JJ
Martin
J V
JV
D000818:Animals; D015153:Blotting, Western; D002540:Cerebral Cortex / Q000187:drug effects* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D008297:Male; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology*; D013574:Synaptosomes / Q000187:drug effects* / Q000378:metabolism; D013963:Thyroid Hormones / Q000378:metabolism / Q000494:pharmacology*
D011506:Proteins; D013963:Thyroid Hormones