We describe a novel cytokine receptor named GP130 Like receptor, or GPL, that displays similarities with the interleukin-6 and interleukin-12 family of signaling receptors. Four different isoforms diverging in their carboxyl terminus were isolated, corresponding to proteins encompassing 560, 610, 626, and 745 amino acids. Sequences included a signal peptide of 32 amino acids, followed by a cytokine binding domain containing four conserved cysteines, a WSDWS motif, and a region consisting of three fibronectin type III domain repeats. No immunoglobulin-like module was identified in the GPL sequences. The intracellular part of longer isoforms contained a proline-rich region defining a box1 motif for interaction with the Janus kinases. The Gpl gene is organized in 15 exons and is located on 5q11.2 in tandem with the gp130 gene. Both genes were only separated by 24 kilobases, with opposite transcriptional orientations. The GPL receptor displayed a 28% identity with gp130. Specific GPL transcripts were observed in tissues involved in reproduction. Transcripts were also found in blood cells and in bone marrow, revealing expression of GPL in all of the myelomonocytic lineage, from hematopoietic stem cells to activated dendritic cells. In monocytes and dendritic cells, expression of GPL was strongly up-regulated by interferon-␥, indicating a possible involvement of GPL in Th1-type immune responses. The molecular basis of cell signaling mediated by GPL was studied using chimeric receptors where external portions of ␣ or ␤ interleukin-5 receptor subunits were fused to the internal portion of GPL or of related receptors. Results indicated that association of GPL to the intracellular portions of gp130, or LIF receptor, allowed the signaling cascade.

Ciliary neurotrophic factor (CNTF) is involved in the survival of a number of different neural cell types, including motor neurons. CNTF functional responses are mediated through a tripartite membrane receptor composed of two signalling receptor chains, gp130 and the leukaemia inhibitory factor receptor (LIFR), associated with a non‐signalling CNTF binding receptor α component (CNTFR). CNTFR‐deficient mice show profound neuronal deficits at birth, leading to a lethal phenotype. In contrast, inactivation of the CNTF gene leads only to a slight muscle weakness, mainly during adulthood, suggesting that CNTFR binds to a second ligand that is important for development. Modelling studies of the interleukin‐6 family member cardiotrophin‐like cytokine (CLC) revealed structural similarities with CNTF, including the conservation of a site I domain involved in binding to CNTFR. Co‐expression of CLC and CNTFR in mammalian cells generates a secreted composite cytokine, displaying activities on cells expressing the gp130–LIFR complex on their surface. Correspondingly, CLC–CNTFR activates gp130, LIFR and STAT3 signalling components, and enhances motor neuron survival. Together, these observations demonstrate that CNTFR induces the secretion of CLC, as well as mediating the functional responses of CLC.

The selective toxicity of the polyene antibiotic amphotericin B between pathogenic eukaryotic organisms and animal cells has often been said to originate in the presence of ergosterol in fungal membranes instead of cholesterol, found in membranes of animal cells. We have tested this hypothesis by measuring the proton efflux induced by amphotericin B in egg yolk phosphatidylcholine small unilamellar vesicles. By measuring circular dichroism under the same conditions, we monitored the interaction of the antibiotic and its conformational changes. Sterol-free vesicles are sensitive to amphotericin B, but the sensitivity of sterol-containing vesicles is always greater and increasingly so with increasing sterol concentration. Ergosterol-containing vesicles are more sensitive than cholesterol-containing vesicles. On the other hand, numerous amphotericin B conformers can be detected in sterol-containing vesicles, depending upon both the concentration of sterol and the amphotericin B sterol ratio. It appears that one conformer, or maybe two at high amphotericin B concentration, is responsible for the induced permeability. From their circular dichroism spectra, these two conformers are the same in the presence of ergosterol or cholesterol. The concentration of amphotericin B necessary to obtain the two conformers is higher with cholesterol than with ergosterol, which agrees with the permeability results.

Class A G-protein-coupled receptors (GPCRs) constitute the largest family of transmembrane receptors in the human genome. Understanding the mechanisms which drove the evolution of such a large family would help understand the specificity of each GPCR sub-family with applications to drug design. To gain evolutionary information on class A GPCRs, we explored their sequence space by metric multidimensional scaling analysis (MDS). Three-dimensional mapping of human sequences shows a non-uniform distribution of GPCRs, organized in clusters that lay along four privileged directions. To interpret these directions, we projected supplementary sequences from different species onto the human space used as a reference. With this technique, we can easily monitor the evolutionary drift of several GPCR sub-families from cnidarians to humans. Results support a model of radiative evolution of class A GPCRs from a central node formed by peptide receptors. The privileged directions obtained from the MDS analysis are interpretable in terms of three main evolutionary pathways related to specific sequence determinants. The first pathway was initiated by a deletion in transmembrane helix 2 (TM2) and led to three sub-families by divergent evolution. The second pathway corresponds to the differentiation of the amine receptors. The third pathway corresponds to parallel evolution of several sub-families in relation with a covarion process involving proline residues in TM2 and TM5. As exemplified with GPCRs, the MDS projection technique is an important tool to compare orthologous sequence sets and to help decipher the mutational events that drove the evolution of protein families.