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The Kiss1 gene codes for kisspeptin, which binds to GPR54, a G-protein-coupled receptor. Kisspeptin and GPR54 are expressed in discrete regions of the forebrain, and they have been implicated in the neuroendocrine regulation of reproduction. Kiss1-expressing neurons are thought to regulate the secretion of gonadotropin-releasing hormone (GnRH) and thus coordinate the estrous cycle in rodents; however, the precise role of kisspeptin-GPR54 signaling in the regulation of gonadotropin secretion is unknown. In this study, we used female mice with deletions in the GPR54 gene [GPR54 knock-outs (KOs)] to test the hypothesis that kisspeptin-GPR54 signaling provides the drive necessary for tonic GnRH/luteinizing hormone (LH) release. We predicted that tonic GnRH/LH secretion would be disrupted in GPR54 KOs and that such animals would be incapable of showing a compensatory rise in LH secretion after ovariectomy. As predicted, we found that GPR54 KO mice do not exhibit a postovariectomy rise in LH, suggesting that tonic GnRH secretion is disrupted in the absence of kisspeptin-GPR54 signaling. We also postulated that kisspeptin-GPR54 signaling is critical for the generation of the estradiol (E)-induced GnRH/LH surge and thus E should be incapable of inducing an LH surge in the absence of GPR54. However, we found that E induced Fos expression in GnRH neurons and produced a GnRH-dependent LH surge in GPR54 KOs. Thus, in mice, kisspeptin-GPR54 signaling is required for the tonic stimulation of GnRH/LH secretion but is not required for generating the E-induced GnRH/LH surge.

We studied the effects of changes in microtubule assembly status upon the intracellular transport of an integral membrane protein from the rough endoplasmic reticulum to the plasma membrane. The protein was the G glycoprotein of vesicular stomatitis virus in cells infected with the Orsay-45 temperature-sensitive mutant of the virus; the synchronous intracellular transport of the G protein could be initiated by a temperature shift-down protocol. The intracellular and surface-expressed G protein were separately detected and localized in the same cells at different times after the temperature shift, by double-immunofluorescence microscopic measurements, and the extent of sialylation of the G protein at different times was quantitated by immunoprecipitation and SDS PAGE of [3SS]methionine-labeled cell extracts. Neither complete disassembly of the cytoplasmic microtubules by nocodazole treatment, nor the radical reorganization of microtubules upon taxol treatment, led to any perceptible changes in the rate or extent of G protein sialylation, nor to any marked changes in the rate or extent of surface appearance of the G protein. However, whereas in control cells the surface expression of G was polarized, at membrane regions in juxtaposition to the perinuclear compact Golgi apparatus, in cells with disassembled microtubules the surface expression of the G protein was uniform, corresponding to the intracellular dispersal of the elements of the Golgi apparatus. The mechanisms of transfer of integral proteins from the rough endoplasmic reticulum to the Golgi apparatus, and from the Golgi apparatus to the plasma membrane, are discussed in the light of these observations, and compared with earlier studies of the intracellular transport of secretory proteins.There is good evidence that, during the intracellular transport of secretory and membrane components, the transfer of these components from the Golgi apparatus (GA) ~ to the cell surface occurs by way of specific vesicles that bud off the trans face of the GA and migrate to the plasma membrane (for a recent review, see reference 13). It is thought that a similar process occurs to transfer these secretory and membrane components from their sites of synthesis and insertion in the rough endoplasmic reticulum (RER) to the GA, with so-called "transition Abbreviations used in this paper: GA, Golgi apparatus; NRK, normal rat kidney; 0-45, temperature-sensitive mutant of vesicular stomatitis virus Orsay-45; RER, rough and endoplasmic reticulum; VSV, vesicular stomatitis virus.