Studies with a growth hormone antagonist and dual-fluorescent confocal microscopy demonstrate that the full-length human growth hormone receptor, butnot the truncated isoform, is very rapidly internalized independent of Jak2-Stat5 signaling

We have investigated trafficking of two negative regulators of growth hormo ne receptor (GHR) signaling: a human, truncated receptor, GHR1-279, and a G H antagonist, B2036, Fluorescent-labeled growth hormone (GH) was rapidly in ternalized by the full-length GHR, with >80% of the hormone internalized wi thin 5 min of exposure to GH, In contrast, <5% of labeled GH was internaliz ed by cells expressing truncated GHR1-279, Using another truncated receptor , GHR1-317 fused to enhanced green fluorescent protein (EGFP), we have expl oited fluorescence energy transfer to monitor the trafficking of ligand-rec eptor complexes. The data confirmed that internalization of this truncated receptor is very inefficient, It was possible to visualize the truncated GH R1-317-EGFP packaged in the endoplasmic reticulum, its rapid movement in me mbrane bound vesicles to the Golgi apparatus, and subsequent transport to t he cell membrane, The GH antagonist, B2036, blocked Jak2-Stat5-mediated GHR signaling but was internalized with a similar time course to native GH, Th e results: 1) demonstrate the rapid internalization of GH when studied unde r physiological conditions; 2) confirm the hypothesis that internalization of cytoplasmic domain truncated human GHRs is very inefficient, which expla ins their dominant negative action; and 3) show that the antagonist action of B2036 is independent of receptor internalization.