A splice variant of the human luteinizing hormone (LH) receptor modulates the expression of wild-type human LH receptor

We previously reported a splice variant form of human LH receptor [hLHR(exon 9)] that lacks exon 9, coding the N-terminal extracellular region close to the first transmembrane domain. Several recent studies suggest that G protein-coupled receptors are able to form dimerization or oligomerization of the receptor, suggesting an intermolecular interaction between hLHR(exon 9) and the wild-type LH receptor (hLHR). The aim of this study, using coimmunoprecipitation, is to examine whether hLHR forms an association with hLHR(exon 9). An interaction between hLHR(exon 9) with the immature band (68 kDa) of hLHR and not with the mature band (85 kDa) was seen. When hLHR and hLHR(exon 9) were coexpressed, the density of hLHR expression was significantly reduced, compared with hLHR expressed alone. The human chorionic gonadotropin-stimulated cAMP accumulation in the cells expressing hLHR(exon 9) was also impaired, compared with the cells expressing hLHR. In this study, we demonstrated that hLHR is capable of forming receptor complexes. Our findings may expand the possibility of a splice variant of hLHR specifically modulating the functional property of the wild-type hLHR.     

Carboxymethylation of methionine residues in bovine pituitary luteinizing hormone and its subunits. Effects on the binding activity with receptor sites and interactions between subunits.

The properties of the component 'X' identified as the primary electron acceptor of Photosystem I in spinach was investigated by electron-paramagnetic-resonance spectroscopy and the complete spectrum obtained for the first time. Component 'X' has gx = 1.78, gy = 1.88 and gz = 2.08; it can be observed only at very low temperatures (8--13K) and high microwave powers. Component X was identified in Photosystem I particles prepared with the French press or with Triton X-100. In samples reduced with ascorbate, illumination at low temperatures results in the photo-oxidation of P700 and reduction of a bound iron-sulphur protein; this is irreversible at low temperature. In samples in which the iron-sulphur proteins are reduced by sodium dithionite, illumination at low temperature results in the oxidation of P700 and the reduction of component 'X'; this is reversible at low temperature. The light-induced P700 signal is the same size with either ascorbate or dithionite as reducing agent, showing that all of the P700 involved in reduction of bound ferredoxin also functions in the reduction of component 'X'.     

Gonadotropin-releasing hormone (GnRH) inhibits ovulation induced with luteinizing hormone (LH) in proestrous hypophysectomized rats

In this paper we present evidence that a single low dose of the natural synthetic gonadotropin-releasing hormone (GnRH), inhibits ovulation induced by LH in proestrous-hypophysectomized rats. Rats hypophysectomized by the parapharyngeal route in the morning of proestrus received an intravenous injection of 100 or 300 ng GnRH at 1400 h immediately followed by 1.0 microgram LH per 100 g bw. In control groups, either one or both hormones were replaced with 0.9% NaCl. Ovulation was assessed the following morning by counting the ova present in oviductal flushings. All the rats treated with LH alone ovulated, and the addition of GnRH reduced significantly the number of ovulating rats and the number of ova per ovulating rat. In other groups of rats hypophysectomized in the morning of proestrus and treated in the same way, ovarian or adrenal secretory rates of estradiol and/or progesterone were measured after cannulation of the corresponding vein, in the afternoon of proestrus. In these animals, GnRH failed to inhibit either the ovarian progesterone surge observed 2 h after LH administration, or the adrenal progesterone secretion. All hypophysectomized rats showed lower ovarian secretory rate of estradiol than intact rats; this rate was not affected by treatment with LH or LH plus GnRH. The systemic estradiol levels in plasma of hypophysectomized rats were distributed within a range of 20 pg/ml to 50 pg/ml. The number of rats whose levels were above 21 pg/ml on estrus day was significantly higher in rats receiving 300 ng GnRH as compared to those receiving 100 ng GnRH, reaching values that surpassed the concentration found in intact, untreated animals at the same time of estrus. This effect did not depend on LH administration.     

Comparison of monoclonal luteinizing hormone (LH) receptor antibody and LH binding sites in vibratome sections of rat ovary by immunohistochemistry

To identify luteinizing hormone (LH) receptors, a monoclonal antibody (MAb) was produced by immunization of Balb/c mice with rat luteal cell membranes. Hybridomas, produced by a method for proteins of low antigenicity, were selected by competition with [125I]-hCG (LH) for luteal membrane binding. Conditions for analysis of LH receptor antibody (IgG2b isotype) binding by immunohistochemistry with an avidin-biotin-peroxidase complex were examined and results compared to localization of bound hCG, to detect receptors. By light microscopy, both bound hCG and the LH receptor antibody were located on luteal cell surfaces. In addition, the LH receptor antibody was associated with luteal cell cytoplasm. Cell surface membrane binding, but not cytoplasmic staining, was reduced in ovaries from rats injected with hCG. By electron microscopy, LH receptor antibody was observed in patches on luteal cell surface membranes and was associated with polysomes, small vesicles, and occasionally with discrete areas of endoplasmic reticulum. Therefore, detection of LH receptors with bound hCG may be limited to receptors found on cell surfaces, while additional LH receptors are revealed by use of a receptor antibody. The cytoplasmic LH receptor may represent stages in the processing of receptor protein. Furthermore, the methodology used in this study should be generally useful for immunohistochemistry with other MAb to receptors.