HIGH-AFFINITY BINDING OF THYROTROPIN TO THE EXTRACELLULAR DOMAIN OF ITS RECEPTOR TRANSFECTED IN CHINESE-HAMSTER OVARY CELLS

Thyrotropin (TSH) receptor is a cell surface receptor that shares a hi gh degree of homology with other glycoprotein hormone receptors includ ing lutropin-choriogonadotropin (LH/CG) and follicle-stimulating hormo ne (FSH) receptors. Although the extracellular domain of TSH receptor is important for ligand binding, no direct information is available on whether extracellular domain alone is sufficient for high-affinity bi nding. Moreover, mutations made in the second cytoplasmic loop or the cytoplasmic tail of TSH receptor were reported to reduce significantly the affinity of TSH binding. In an attempt to determine whether TSH r eceptor extracellular domain is sufficient for high-affinity TSH bindi ng or whether it requires transmembrane regions, we made a construct ( TSHR-EX/CMV) that encodes for only the extracellular domain plus a for eign hydrophobic tail. The TSHR-EX/CMV was transfected and stably expr essed in Chinese hamster ovary (CHO) cells. The truncated receptor was anchored to the cell surface through the hydrophobic tail at the carb oxyl terminus. High-affinity TSH binding was observed comparable to th at of the cells transfected with full-length TSH receptor. The CHO cel ls transfected with TSHR-EX/CMV did not respond to TSH stimulation of adenylate cyclase, whereas the cells transfected with the full-length TSH receptor cDNA did. The data presented here show that the extracell ular domain of TSH receptor is sufficient to confer high-affinity TSH binding.