Internalization of Ovine Luteinizing Hormone/Human Chorionic Gonadotropin Recombinants: Differential Effects of theα- andβ-Subunits*

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Abstract
The rate of internalization and degradation of radioiodinated hCG [human chorionic gonadotropin] and ovine LH [luteinizing hormone] (oLH) as well as homologous and heterologous recombinants of their subunits was studied in suspensions of ovine luteal cells. Hormone bound to the plasma membrane was quantified by treating the cells with acidic buffer (pH 3.0) and quantification of the radioactivity that was removed from receptor. The radioactivity remaining in the cell pellet was considered to be intracellular. The extent of degradation of the radioactive hormones was determined by subjecting the radioactivity released into the medium during incubation to precipitation with 20% trichloroacetic acid. The nonprecipitable radioactivty was considered to have been degraded. Radioactivity was lost from the cell membrane with a t1/2 of 16.8 .+-. 2.5 h for radioiodinated hCG, 22.8 .+-. 3.8 h for the .alpha.-subunit of hCG recombined with radioiodinated .beta.-subunit of hCG, 8.9 .+-. 4.5 h for the .alpha.-subunit of oLH recombined with the radioiodinated .beta.-subunit of hCG, 0.5 .+-. 0.1 h for the .alpha.-subunit of hCG recombined with the radioiodinated .beta.-subunit of oLH, 0.5 .+-. 0.1 h for radioiodinated oLH and 0.7 .+-. 0.2 for the .alpha.-subunit of oLH recombined with the radioiodinated .beta.-subunit of oLH. In general, the levels of radioactivity that were intracellular and the rate of degradation of the hormone were inversely related to the quantities of hormone that remained on the plasma membrane. The preparations that contained the .beta.-subunit of hCG were all internalized at a slower rate (t1/2 = 9-22 h) and consequently were degraded more slowly than any of the preparations that contained the .beta.-subunit of oLH (t1/2 = 0.5-0.7 h). When the radioiodine in oLH was present in the .beta.-subunit, intracellular levels of radioactivity were higher and degradation occurred more rapidly than when the radioiodine was in the .alpha.-subunit. Although the differences were less dramatic, hCG with radioiodine in the .beta.-subunit also reached higher levels intracellularly, but was degraded to a lesser extent at 16 and 24 h than was hcG with radioiodine in the .alpha.-subunit. These data demonstrated a dramatic difference (.apprx. 30-fold) in the rate of loss of oLH vs. that of hCG from the membrane of ovine luteal cells. Further, it appears that the .beta.-subunit of each of the 2 hormones is the major factor in determining the rate of internalization of the intact hormone. The differences is how the 2 hormones are processed at the receptor level may help explain the known differences in their steroiodogenic potencies.