Visualization of the thyrotropin-releasing hormone receptor and its ligand during endocytosis and recycling.
AUTOR(ES)
Ashworth, R
RESUMO
Endocytosis and recycling of both thyrotropin-releasing hormone (TRH) and its G-protein-coupled receptor were visualized by conventional and confocal fluorescence microscopy in pituitary cells using a rhodamine-labeled TRH analog (Rhod-TRH) and indirect immunofluorescent staining of cells stably transfected with an epitope-tagged TRH receptor (TRHR). The epitope-tagged TRHR was confined to the cell surface prior to agonist treatment. Both Rhod-TRH and TRHR were also localized on the plasma membrane after agonist binding at 0 degrees C. Ligand binding at 37 degrees C resulted in rapid endocytosis, and both Rhod-TRH and the epitope-tagged TRHR appeared in cytoplasmic vesicles within 5 min. Fluorescently labeled TRH and transferrin colocalized in the same endocytotic vesicles, and internalization of Rhod-TRH and TRHR was inhibited by hypertonic medium, suggesting that endocytosis occurred by a clathrin-dependent mechanism. Internalized TRHRs returned to the membrane within 20 min after removal of TRH, and cycloheximide did not block receptor recycling. A mutant TRHR truncated at Cys335 signaled but did not internalize Rhod-TRH, confirming the importance of the carboxyl terminus of the TRHR in receptor-mediated endocytosis. Thus, the TRH-TRHR complex is endocytosed via clathrin-coated vesicles and the receptor is recycled to the plasma membrane.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=42771Documentos Relacionados
- The Ups and Downs of Thyrotropin-Releasing Hormone
- Assessment of Thyrotropin-Releasing Hormone and Thyrotropin Reserve in Man
- Direct visualization of receptors for thyrotropin-releasing hormone with a fluorescein-labeled analog.
- Thyroid Hormone Inhibition of the Prolactin Response to Thyrotropin-Releasing Hormone
- Metabolism and Excretion of Exogenous Thyrotropin-Releasing Hormone in Humans