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Rob Leurs

Publications and source records attributed to Rob Leurs.

62 records · Page 4Linked to original sources
PubMed2002

Chimaeric gonadotropin-releasing hormone (GnRH) peptides with improved affinity for the catfish (Clarias gariepinus) GnRH receptor.

The gonadotropin-releasing hormone (GnRH) receptor in catfish differs from its mammalian counterparts in showing a very low affinity for the hypothalamic GnRH form [i.e. catfish GnRH (cfGnRH)] and a very high affinity for the highly conserved mesencephalic GnRH, chicken GnRH-II (cGnRH-II). In the present study we investigated the molecular interactions between ligand and receptor involved in determining the ligand selectivity of the catfish GnRH receptor. Studies on the binding characteristics of the catfish GnRH receptor for cfGnRH and cGnRH-II as well as for mammalian GnRH (mGnRH) and synthetic chimaeric GnRHs, differing at positions 5, 7 and 8, revealed that the low affinity of the catfish receptor for cfGnRH can be improved by replacing Leu(7) by a tryptophan residue and/or Asn(8) by either a tyrosine or an arginine residue. Testing cfGnRH and cGnRH-II as well as mGnRH and the chimaeric GnRHs on Asp(304)-->Ala, Asp(304)-->Glu and Asp(304)-->Asn mutant catfish GnRH receptors revealed that Asp(304) of the catfish receptor mediates the recognition of Arg(8) in mGnRH, as well as in the chimaeric peptides [Arg(8)]cfGnRH and [Arg(8)]cGnRH-II, but seems to be less important for the recognition of Tyr(8) in cGnRH-II. On the basis of these results, a three-dimensional model for the binding of [Arg(8)]cGnRH-II to the catfish GnRH receptor is proposed.

Alanine↗
PubMed2002

A novel phenylaminotetralin radioligand reveals a subpopulation of histamine H(1) receptors.

Previously, (-)-trans-1-phenyl-3-N,N-dimethylamino-1,2,3,4-tetrahydronaphthalene ([-]-trans-H(2)-PAT) was shown to activate stereospecifically histamine H(1) receptors coupled to modulation of tyrosine hydroxylase activity in guinea pig and rat forebrain in vitro and in vivo. Furthermore, the novel radioligand [(3)H](-)-trans-H(2)-PAT was shown to label selectively H(1) receptors in guinea pig and rat brain with high affinity (K(D), ~0.1 and 0.5 nM, respectively) and a B(max) about 50 and 15%, respectively, of that observed for the H(1) antagonist radioligand [(3)H]mepyramine. In the current study, [(3)H](-)-trans-H(2)-PAT-labeled cloned guinea pig and human H(1) receptors in Chinese hamster ovary (CHO) cell membranes with high affinity (K(D), ~0.08 and 0.23 nM, respectively) and a B(max) about 15% of that observed for [(3)H]mepyramine. The binding of H(2)-PAT to H(1) receptors in both CHO-H(1) cell lines was stereoselective with the (-)-trans-isomer having affinity (K(i), ~1.5 nM) about 4-, 20-, and 50-times higher than the (-)-cis-, (+)-trans-, and (+)-cis-isomers, respectively; the affinity of (-)-trans-H(2)-PAT was unaffected by excess GTP. In functional assays, (-)-trans-H(2)-PAT was a full antagonist of histamine H(1)-mediated stimulation of phospholipase C (PLC) and [(3)H]inositol phosphates (IP) formation in CHO-H(1) cells, a full inverse agonist of constitutively active H(1) receptors in COS-7-H(1) cells, and a full competitive antagonist (pA(2) = 9.2) of histamine H(1)-mediated contraction of guinea pig ileum. It is concluded that (-)-trans-H(2)-PAT is an antagonist at H(1) receptors coupled to PLC/IP formation and smooth muscle contraction. Meanwhile, the observation that [(3)H](-)-trans-H(2)-PAT labels only a subpopulation of H(1) receptors and that (-)-trans-H(2)-PAT activates H(1) receptors coupled to modulation of tyrosine hydroxylase suggests that there may be post-translational H(1) receptor heterogeneity.

Animals↗
PubMed2002

The rat cytomegalovirus R33-encoded G protein-coupled receptor signals in a constitutive fashion.

The rat cytomegalovirus (RCMV) R33 gene is conserved among all betaherpesviruses and encodes a protein (pR33) that shows sequence similarity with chemokine-binding G protein-coupled receptors (GPCRs). Previously, the physiological significance of the R33 gene was demonstrated by the finding that an RCMV strain with R33 deleted is severely attenuated in vivo and is unable to either enter or replicate in the salivary glands of infected rats. Here, we report that RCMV pR33 is expressed as a functional GPCR that signals in an agonist-independent manner in both COS-7 and Rat2 cells. Transient expression of pR33 in COS-7 cells results in constitutive activation of phospholipase C (PLC) due to coupling to G proteins of the G(q) class. Interestingly, PLC activation is partially inhibited by cotransfection with G(alpha)-transducin subunits, which indicates the involvement of G(betagamma) as well as Galpha subunits in pR33-mediated signaling. Surprisingly, PLC activation is also partially inhibited by addition of pertussis toxin (PTX), suggesting that pR33 activates not only G(q) but also G(i/0) proteins. The constitutive activation of G(i/0) proteins by pR33 is further demonstrated by the PTX-sensitive decrease of CRE-mediated transcription and the PTX-sensitive increase of both NF-kappaB- and SRE-mediated transcription. In contrast to its homolog of human herpesvirus 6B (pU12), pR33 does not bind RANTES.

Animals↗
PubMed2002

Kaposi's sarcoma-associated herpesvirus-encoded G protein-coupled receptor ORF74 constitutively activates p44/p42 MAPK and Akt via G(i) and phospholipase C-dependent signaling pathways.

The G protein-coupled receptor encoded by Kaposi's sarcoma-associated herpesvirus, also referred to as ORF74, has been shown to stimulate oncogenic and angiogenic signaling pathways in a constitutively active manner. The biochemical routes linking ORF74 to these signaling pathways are poorly defined. In this study, we show that ORF74 constitutively activates p44/p42 mitogen-activated protein kinase (MAPK) and Akt via G(i)- and phospholipase C (PLC)-mediated signaling pathways. Activation of Akt by ORF74 appears to be phosphatidylinositol 3-kinase (PI3-K) dependent but, interestingly, is also mediated by activation of protein kinase C (PKC) and p44/p42 MAPK. ORF74 may signal to Akt via p44/p42 MAPK, which can be activated by G(i), through activation of PI3-K or through PKC via the PLC pathway. Signaling of ORF74 to these proliferative and antiapoptotic signaling pathways can be further modulated positively by growth-related oncogene (GROalpha/CXCL1) and negatively by human gamma interferon-inducible protein 10 (IP-10/CXCL10), thus acting as an agonist and an inverse agonist, respectively. Despite the ability of the cytomegalovirus-encoded chemokine receptor US28 to constitutively activate PLC, this receptor does not increase phosphorylation of p44/p42 MAPK or Akt in COS-7 cells. Hence, ORF74 appears to signal through a larger diversity of G proteins than US28, allowing it to couple to proliferative and antiapoptotic signaling pathways. ORF74 can therefore be envisioned as an attractive target for novel treatment of Kaposi's sarcoma.

Animals↗
PubMed2002

Two gonadotropin-releasing hormone receptors in the African catfish: no differences in ligand selectivity, but differences in tissue distribution.

Ligand-binding studies revealed the presence of GnRH-binding sites in African catfish ovary. However, our expression profiling studies failed to detect the previously identified catfish GnRH receptor (cfGnRH-R1) mRNA in this tissue. This negative result instigated us to clone an additional catfish GnRH receptor (cfGnRH-R2) cDNA and study its expression in different tissues in conjunction with the expression of the two catfish GnRH (i.e. cfGnRH and cGnRH-II) genes. The highest cfGnRH-R1 and cfGnRH-R2 mRNA levels were detected in pituitary for cfGnRH-R1 and in brain and ovary for cfGnRH-R2. cfGnRH mRNA was coexpressed with cfGnRH-R1 mRNA in pituitary and brain and with cfGnRH-R2 mRNA in brain and ovary. Ubiquitous expression of cGnRH-II mRNA was observed in all tissues tested, with the highest expression in brain, heart, pituitary, ovary, and head-kidney. Binding studies revealed that cfGnRH-R1 had a higher affinity than cfGnRH-R2 for cGnRH-II, cfGnRH, and various other GnRH agonists. However, this was not reflected in the inositol phosphate or cAMP signal transduction properties of both types of cfGnRH-R. We therefore conclude that in catfish, functional ligand/receptor units evolved by restricted coexpression of a particular receptor in combination with a particular GnRH in particular (nearby) tissue(s).

Amino Acid Sequence↗
PubMed2002

Histamine receptors: specific ligands, receptor biochemistry, and signal transduction.

During the past few years, there has been a tremendous increase in our understanding of the histamine receptors. Important progress has been made in the development of H1-receptor agonists and the rationalization of H1-receptor-ligand interaction. The recent observation of constitutive H1- and H2-receptor activity has led to a reclassification of H1- and H2-antagonists. For the H3-receptor, a wide variety of selective and potent ligands are currently available and await clinical application. The recent cloning of the H3-receptor gene and the anticipated generation of transgenic mice will facilitate this development. Within the field of signal transduction, a previously unanticipated complexity has been unravelled. With the cloning of the H3-receptor gene, a similar complexity is to be expected.

Animals↗