Nitric oxide (NO) is a free radical that mediates signal transduction and influences a wide variety of biological processes. It is generated by three different nitric oxide synthase (NOS) isoforms (I-III). Although all NOS enzymes have been identified in the thyroid cell, NOS III is the highest expressed. We reported that NO acts as a regulator of the TSH-induced gene expression and that TSH is able to increases NO production and NOS III mRNA expression.
The aim of this work was to analyze the signal pathway involved in the TSH-induced expression of NOS III in the thyroid cells FRTL-5.
TSH stimulated NOS III protein expression in a time and concentration dependent manner. We evaluated a possible post-transcriptional effect of TSH on NOS III protein stabilization. A significant increase in NOS III protein half-life was induced by TSH treatment. In order to examine the signal pathway involved, we measured the effect of TSH on NOS III protein expression in the presence of different kinase inhibitors. We observed a significant inhibition of the TSH-induced protein up-regulation in presence of PKA and Akt inhibitors. As Akt is canonically activated by PI3K, we evaluated the effect of PI3K inhibitors on the TSH-increased NOS III expression but no evident blockage was observed. When we evaluated Akt phosphorylation in response to TSH, a TSH-dependent Akt phosphorylation that involves PKA activity was found.
In conclusion, our results provide evidence of a novel regulation of NOS III expression by TSH involving a PKA-mediated activation of the Akt pathway.
