Modulation of α2C adrenergic receptor temperature-sensitive trafficking by HSP90

Decreasing the temperature to 30 °C is accompanied by significant enhancement of α2C-AR plasma membrane levels in several cell lines with fibroblast phenotype, as demonstrated by radioligand binding in intact cells. No changes were observed on the effects of low-temperature after blocking receptor internalization in α2C-AR transfected HEK293T cells. In contrast, two pharmacological chaperones, dimethyl sulfoxide and glycerol, increased the cell surface receptor levels at 37 °C, but not at 30 °C. Further, at 37 °C α2C-AR is co-localized with endoplasmic reticulum markers, but not with the lysosomal markers. Treatment with three distinct HSP90 inhibitors, radicicol, macbecin and 17-DMAG significantly enhanced α2C-AR cell surface levels at 37 °C, but these inhibitors had no effect at 30 °C. Similar results were obtained after decreasing the HSP90 cellular levels using specific siRNA. Co-immunoprecipitation experiments demonstrated that α2C-AR interacts with HSP90 and this interaction is decreased at 30 °C. The contractile response to endogenous α2C-AR stimulation in rat tail artery was also enhanced at reduced temperature. Similar to HEK293T cells, HSP90 inhibition increased the α2C-AR contractile effects only at 37 °C. Moreover, exposure to low-temperature of vascular smooth muscle cells from rat tail artery decreased the cellular levels of HSP90, but did not change HSP70 levels. These data demonstrate that exposure to low-temperature augments the α2C-AR transport to the plasma membrane by releasing the inhibitory activity of HSP90 on the receptor traffic, findings which may have clinical relevance for the diagnostic and treatment of Raynaud Phenomenon.

Research Highlights
► At 37 °C α2C-AR is localized mostly in the endoplasmic reticulum in HEK293T cells.
► Exposure to low-temperature stimulated the receptor transport to the cell surface.
► Inhibition of HSP90 has similar effects on the α2C-AR traffic as low-temperature.
► In rat tail artery, HSP90 inhibitors increase the contraction to α2C-AR stimulation.
► These findings may have clinical relevance in Raynaud Phenomenon.