Tissue distribution, turnover, and glycosylation of the long and short growth hormone receptor isoforms in rat tissues

Two isoforms of the GH receptor, the full-length receptor (GHR(L)) and a short isoform (GHR(S)) that lacks the transmembrane and intracellular domains of GHR(L), have been analyzed in rat tissue extracts by Western blotting and immunoprecipitation. Although quantitative estimates of GHR(S) and GHR(L) based on coprecipitation of [I-125]GH indicated similar amounts of both isoforms in tissue extracts, the 110 kDa band corresponding to GHR(L) was generally not detected on Western blots without enrichment by immunoprecipitation. Two bands with electrophoretic mobilities corresponding to 38 and 42 kDa were present in extracts prepared from liver, muscle, and adipocytes. Western blots of the GH binding protein in rat serum also revealed two bands, but these had electrophoretic mobilities corresponding to 44 and 52 kDa. After digestion by endoglycosidase F, a single band with an electrophoretic mobility corresponding to 31 kDa was detected in samples from adipocytes, liver or serum, indicating that GHR(S) retained in tissues is glycosylated less extensively than that in rat serum. Digestion with neuraminidase indicated that the smaller glycoproteins in tissue extracts lack sialic acid residues that are present in serum samples. Furthermore, endoglycosidase H degraded GHR(S) in liver extracts to a 31 kDa band but did not degrade serum samples, suggesting that tissues retain a high mannose form of GHR(S). The abundance of GHR(S) or GHR(L) in tissues from male, virgin female, and pregnant rats was estimated from the amount of I-125-GH that was bound to each isoform after immunoprecipitation. Liver contained more than 10 times as much GHR(S) per gram of tissue as fat or muscle. In liver, muscle, and fat, the amount of GHR(S) exceeded that of GHR(L), sometimes by as much as 6-fold. GHBP levels in serum of females exceeded those in males, and rose even higher in pregnant females. The abundance of GHR(S) in all tissue extracts paralleled serum levels. In muscle and fat, the levels of GHR, did not differ in male, female and pregnant rats, whereas in liver, the pattern was similar to the GHR(S) pattern. In all tissues, pools of GHR(S) exceeded those of GHR(L) by a factor that grew larger as tissue and serum levels increased. The half life of GHBP in serum was estimated to be 2.4 h in rats treated with cycloheximide, whereas that of GHR(S) was 20 min in liver and 8.5 h in fat. These results suggest that GHR(S) is synthesized in liver 8 times faster than it is released into serum, whereas synthesis in fat is less than 30% of the rate at which it is released into serum by all tissues. Therefore, liver appears to be the major source of GHBP in serum. Although secretion into the circulatory system accounts for little or perhaps none of its turnover in some tissues, GHR(S) peels in tissues do appear to be regulated, suggesting that GHR(S) may function primarily in the cells in which it is synthesized.