Intermolecular cross-talk between NTR1 and NTR2 neurotensin receptor promotes intracellular sequestration and functional inhibition of NTR1 receptors.

G-protein-coupled receptors (GPCR) are now regarded as being able to acquire heterodimer conformations affecting their pharmacology, signaling and trafficking. In co-immunoprecipitation studies using differentially epitope-tagged receptors, we herein provide direct evidence for heterodimerization of human neurotensin type 1 receptor (hNTR1) and type 2 receptor (hNTR2). Using chimeric constructs, we also identified the hNTR2 transmembrane 2 (TM2) to TM4 region as crucial for the formation of the dimerization interface.

Phylogenetic history, pharmacological features, and signal transduction of neurotensin receptors in vertebrates.

Neurotensin (NTS) plays important roles in neurotransmission and neuromodulation in the nervous system. NTS exerts its effects mainly by binding to the neurotensin receptor 1 (NTSR1) and receptor 2 (NTSR2) that belong to the G protein-coupled receptor superfamily. While studies on NTS and NTSR have been conducted mainly in mammalian systems, little is known about this ligand-receptor pair in nonmammalian species.

Molecular cloning and expression of steroidogenic acute regulatory protein from bullfrog (Rana catesbeiana).

Steroidogenic acute regulatory protein (StAR) transfers cholesterol from the outer mitochondrial membrane to the inner membrane where the cytochrome P450 side chain cleavage enzyme (P450scc) resides. This process is the rate-limiting step in steroidogenesis. StAR cDNAs have been cloned and characterized from a range of different species.

A gonadotropin-releasing hormone-II antagonist induces autophagy of prostate cancer cells.

Gonadotropin-releasing hormone-I (GnRH-I) is known to directly regulate prostate cancer cell proliferation. However, the role of GnRH-II in prostate cancer is unclear. Here, we investigated the effect of the GnRH-II antagonist trptorelix-1 (Trp-1) on growth of PC3 prostate cancer cells.

Molecular evolution of multiple forms of kisspeptins and GPR54 receptors in vertebrates.

Kisspeptin and its receptor GPR54 play important roles in mammalian reproduction and cancer metastasis. Because the KiSS and GPR54 genes have been identified in a limited number of vertebrate species, mainly in mammals, the evolutionary history of these genes is poorly understood. In the present study, we have cloned multiple forms of kisspeptin and GPR54 cDNAs from a variety of vertebrate species.

Molecular cloning of the bullfrog kisspeptin receptor GPR54 with high sensitivity to Xenopus kisspeptin.

Kisspeptin and its receptor, GPR54, play important roles in mammalian reproduction and cancer development. However, little is known about their function in nonmammalian species. In the present study, we have isolated the cDNA encoding the kisspeptin receptor, GPR54, from the bullfrog, Rana catesbeiana.

Identification of farnesyl pyrophosphate and N-arachidonylglycine as endogenous ligands for GPR92.

A series of small compounds acting at the orphan G protein-coupled receptor GPR92 were screened using a signaling pathway-specific reporter assay system. Lipid-derived molecules including farnesyl pyrophosphate (FPP), N-arachidonylglycine (NAG), and lysophosphatidic acid were found to activate GPR92. FPP and lysophosphatidic acid were able to activate both G(q/11)- and G(s)-mediated signaling pathways, whereas NAG activated only the G(q/11)-mediated signaling pathway.

Involvement of amino acids flanking Glu7.32 of the gonadotropin-releasing hormone receptor in the selectivity of antagonists.

The Glu/Asp(7.32) residue in extracellular loop 3 of the mammalian type-I gonadotropin-releasing hormone receptor (GnRHR) interacts with Arg(8) of GnRH-I, conferring preferential ligand selectivity for GnRH-I over GnRH-II. Previously, we demonstrated that the residues (Ser and Pro) flanking Glu/Asp(7.

Extracellular loop 3 (ECL3) and ECL3-proximal transmembrane domains VI and VII of the mesotocin and vasotocin receptors confer differential ligand selectivity and signaling activity.

Mesotocin (MT) and vasotocin (VT) are the nonmammalian orthologs of mammalian oxytocin (OT) and arginine vasopressin (AVP), respectively. The OT/AVP family of peptides has arisen from gene duplication but has evolved to possess high selectivity toward their cognate receptors. The process of molecular evolution of receptors to confer high selectivity to their cognate ligands, however, is poorly understood.

Cloning and activation of the bullfrog apelin receptor: Gi/o coupling and high affinity for [Pro1]apelin-13.

In mammals, apelin and its G protein-coupled receptor, APJ, regulate blood pressure, intake of food and water, and cardiac contractility. In this study, we report the cloning and functional characterization of APJ in the bullfrog, Rana catesbeiana. Bullfrog APJ (bfAPJ) cDNA contains an open reading frame of 1083 nucleotides encoding a protein of 360 amino acid residues.

Effects of butyltin compounds on follicular steroidogenesis in the bullfrog(Rana catesbeiana).

The effects of butyltin compounds on follicular steroidogenesis in amphibians were examined using ovarian follicles of Rana catesbeiana. Isolated follicles were cultured for 18h in the presence and absence of frog pituitary homogenate (FPH) or various steroid precursors, and the steroid levels in the follicles or culture media were measured by radioimmunoassay (RIA). Among the butyltin compounds, tributyltin (TBT) strongly inhibited the FPH-induced synthesis of pregnenolone (P(5)), progesterone (P(4)) and testosterone (T).

Salivary cortisol and DHEA levels in the Korean population: age-related differences, diurnal rhythm, and correlations with serum levels.

Purpose: The primary objective of this study was to examine the changes of basal cortisol and DHEA levels present in saliva and serum with age, and to determine the correlation coefficients of steroid concentrations between saliva and serum. The secondary objective was to obtain a standard diurnal rhythm of salivary cortisol and DHEA in the Korean population.

Materials And Methods: For the first objective, saliva and blood samples were collected between 10 and 11 AM from 359 volunteers ranging from 21 to 69 years old (167 men and 192 women).

Molecular evolution of neuropeptide receptors with regard to maintaining high affinity to their authentic ligands.

Recently, we cloned many of the bullfrog neuropeptide G protein-coupled receptors (GPCRs), including receptors for vasotocin (VT), mesotocin, gonadotropin-releasing hormone (GnRH), neurotensin, apelin, and metastin. Bullfrog GPCRs usually have high affinity for bullfrog ligands but relatively low affinity for mammalian ligands. Reciprocally, synthetic agonists and antagonists developed based upon mammalian ligands display lower affinity at bullfrog receptors.

Cellular and molecular biology of orphan G protein-coupled receptors.

The superfamily of G protein-coupled receptors (GPCRs) is the largest and most diverse group of membrane-spanning proteins. It plays a variety of roles in pathophysiological processes by transmitting extracellular signals to cells via heterotrimeric G proteins. Completion of the human genome project revealed the presence of approximately 168 genes encoding established nonsensory GPCRs, as well as 207 genes predicted to encode novel GPCRs for which the natural ligands remained to be identified, the so-called orphan GPCRs.

Vasotocin and mesotocin stimulate the biosynthesis of neurosteroids in the frog brain.

The neurohypophysial nonapeptides vasopressin (VP) and oxytocin (OT) modulate a broad range of cognitive and social activities. Notably, in amphibians, vasotocin (VT), the ortholog of mammalian VP, plays a crucial role in the control of sexual behaviors. Because several neurosteroids also regulate reproduction-related behaviors, we investigated the possible effect of VT and the OT ortholog mesotocin (MT) in the control of neurosteroid production.

The CCAAT enhancer-binding protein-alpha negatively regulates the transactivation of androgen receptor in prostate cancer cells.

The basic leucine zipper transcription factor, CCAAT enhancer-binding protein-alpha (C/EBPalpha), negatively regulates cell proliferation and induces terminal differentiation of various cell types. C/EBPalpha is expressed in the prostate, but its potential role in the tissue is unknown. Herein, we show that C/EBPalpha is highly expressed at the stage of growth arrest during prostate development.

Modulation of androgen receptor transactivation by the SWI3-related gene product (SRG3) in multiple ways.

The SWI3-related gene product (SRG3), a component of the mouse SWI/SNF complex, has been suggested to have an alternative function. Here, we demonstrate that in the prostate transactivation of the androgen receptor (AR) is modulated by SRG3 in multiple ways. The expression of SRG3, which is developmentally regulated in the prostate, is induced by androgen through AR.

Differential effects of gonadotropin-releasing hormone (GnRH)-I and GnRH-II on prostate cancer cell signaling and death.

Context: GnRH is known to directly regulate prostate cancer cell proliferation, but the precise mechanism of action of the peptide is still under investigation.

Objective: This study demonstrates differential effects of GnRH-I and GnRH-II on androgen-independent human prostate cancer cells.

Results: Both GnRH-I and GnRH-II increased the intracellular Ca(2+) concentration ([Ca(2+)](i)) either through Ca(2+) influx from external Ca(2+) source or via mobilization of Ca(2+) from internal Ca(2+) stores.

Extracellular loop 3 (EL3) and EL3-proximal transmembrane helix 7 of the mammalian type I and type II gonadotropin-releasing hormone (GnRH) receptors determine differential ligand selectivity to GnRH-I and GnRH-II.

Mammalian type I and II gonadotropin-releasing hormone (GnRH) receptors (GnRHRs) show differential ligand preference for GnRH-I and GnRH-II, respectively. Using a variety of chimeric receptors based on green monkey GnRHR-2 (gmGnRHR-2), a representative type II GnRHR, and rat GnRHR, a representative type I GnRHR, this study elucidated specific domains responsible for this ligand selectivity. A chimeric gmGnRHR-2 with the extracellular loop 3 (EL3) and EL3-proximal transmembrane helix 7 (TMH7) of rat GnRHR showed a great increase in ligand sensitivity to GnRH-I but not to GnRH-II.

Expression of the putative sterol binding protein Stard6 gene is male germ cell specific.

Mammalian spermatogenesis is orchestrated by many specific molecular and cellular events. To understand the detailed mechanism by which spermatogenesis is controlled, the specific genes involved in this process must be identified and studied. From the subtracted cDNA library of rat testis prepared using the representational difference analysis (RDA) method, we isolated the cDNA clone of steroidogenic acute regulatory (StAR) protein-related lipid transfer (START) protein 6 (Stard6).

Membrane-proximal region of the carboxyl terminus of the gonadotropin-releasing hormone receptor (GnRHR) confers differential signal transduction between mammalian and nonmammalian GnRHRs.

Recently, we demonstrated that the mammalian type-I GnRH receptor (GnRHR) has a high preference for the phospholipase C/protein kinase C (PLC/PKC)-linked signaling pathway, whereas non-mammalian bullfrog (bf) GnRHRs couple to both adenylate cyclase/protein kinase A (AC/PKA)- and PLC/PKC-linked signaling pathways. In the pre-sent study, using AC/PKA-specific reporter (cAMP-responsive element-luciferase) and PLC/PKC-specific reporter (serum-responsive element-luciferase) systems, we attempted to identify the motif responsible for this difference. A deletion of the intracellular carboxyl-terminal tail (C tail) of bfGnRHR-1 remarkably decreased its ability to induce the AC/PKA-linked signaling pathway.

Identification of amino acid residues that direct differential ligand selectivity of mammalian and nonmammalian V1a type receptors for arginine vasopressin and vasotocin. Insights into molecular coevolution of V1a type receptors and their ligands.

Arginine vasotocin (VT) is the ortholog in all nonmammalian vertebrates of arginine vasopressin (AVP) in mammals. We have previously cloned an amphibian V1atype vasotocin receptor (VT1R) that exhibited higher sensitivity for VT than AVP, while the mammalian V1a type receptor (V1aR) responded better to AVP than VT. In the present study, we identified the amino acid residues that confer differential ligand selectivity for AVP and VT between rat V1aR and bullfrog VT1R (bfVT1R).

GnRH-II analogs for selective activation and inhibition of non-mammalian and type-II mammalian GnRH receptors.

Recently, we identified three types of non-mammalian gonadotropin-releasing hormone receptors (GnRHR) in the bullfrog (designated bfGnRHR-1-3), and a mammalian type-II GnRHR in green monkey cell lines (denoted gmGnRHR-2). All these receptors responded better to GnRH-II than GnRH-I, while mammalian type-I GnRHR showed greater sensitivity to GnRH-I than GnRH-II. In the present study, we designed new GnRH-II analogs and examined whether they activated or inhibited non-mammalian and mammalian type-II GnRHRs.