Papers by Sebastian Furness
British journal of pharmacology, 2015
Clinical use of cinacalcet in hyperparathyroidism is complicated by its tendency to induce hypoca... more Clinical use of cinacalcet in hyperparathyroidism is complicated by its tendency to induce hypocalcaemia, arising partly from activation of calcium-sensing receptors (CaS receptors) in the thyroid and stimulation of calcitonin release. CaS receptor allosteric modulators that selectively bias signalling towards pathways that mediate desired effects [e.g. parathyroid hormone (PTH) suppression] rather than those mediating undesirable effects (e.g. elevated serum calcitonin), may offer better therapies. We characterized the ligand-biased profile of novel calcimimetics in HEK293 cells stably expressing human CaS receptors, by monitoring intracellular calcium (Ca(2+) i ) mobilization, inositol phosphate (IP)1 accumulation, ERK1/2 phosphorylation (pERK1/2) and receptor expression. Phenylalkylamine calcimimetics were biased towards allosteric modulation of Ca(2+) i mobilization and IP1 accumulation. S,R-calcimimetic B was biased only towards IP1 accumulation. R,R-calcimimetic B and AC-26534...
ACS pharmacology & translational science, Jan 11, 2019
The calcitonin receptor (CTR) is a class B G protein-coupled receptor (GPCR) that responds to the... more The calcitonin receptor (CTR) is a class B G protein-coupled receptor (GPCR) that responds to the peptide hormone calcitonin (CT). CTs are clinically approved for the treatment of bone diseases. We previously reported a 4.1 Å structure of the activated CTR bound to salmon CT (sCT) and heterotrimeric Gs protein by cryo-electron microscopy (Liang, Y.-L., et al. Phase-plate cryo-EM structure of a class B GPCR-G protein complex. Nature 2017, 546, 118−123). In the current study, we have reprocessed the electron micrographs to yield a 3.3 Å map of the complex. This has allowed us to model extracellular loops (ECLs) 2 and 3, and the peptide N-terminus that previously could not be resolved. We have also performed alanine scanning mutagenesis of ECL1 and the upper segment of transmembrane helix 1 (TM1) and its extension into the receptor extracellular domain (TM1 stalk), with effects on peptide binding and function assessed by cAMP accumulation and ERK1/2 phosphorylation. These data were combined with previously published alanine scanning mutagenesis of ECL2 and ECL3 and the new structural information to provide a comprehensive 3D map of the molecular surface of the CTR that controls binding and signaling of distinct CT and related peptides. The work highlights distinctions in how different, related, class B receptors may be activated. The new mutational data on the TM1 stalk and ECL1 have also provided critical insights into the divergent control of cAMP versus pERK signaling and, collectively with previous mutagenesis data, offer evidence that the conformations linked to these different signaling pathways are, in many ways, mutually exclusive. This study furthers our understanding of the complex nature of signaling elicited by GPCRs and, in particular, that of the therapeutically important class B subfamily.
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, May 4, 2016
Glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) agonists have emerged as treatment options for ... more Glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) agonists have emerged as treatment options for type 2 diabetes mellitus (T2DM). GLP-1R signals through G-protein-dependent, and G-protein-independent pathways by engaging the scaffold protein b-arrestin; preferential signalling of ligands through one or the other of these branches is known as 'ligand bias'. Here we report the discovery of the potent and selective GLP-1R G-protein-biased agonist, P5. We identified P5 in a high-throughput autocrine-based screening of large combinatorial peptide libraries, and show that P5 promotes G-protein signalling comparable to GLP-1 and Exendin-4, but exhibited a significantly reduced b-arrestin response. Preclinical studies using different mouse models of T2DM demonstrate that P5 is a weak insulin secretagogue. Nevertheless, chronic treatment of diabetic mice with P5 increased adipogenesis, reduced adipose tissue inflammation as well as hepatic steatosis and was more effective at correcting hyperglycaemia and lowering haemoglobin A 1c levels than Exendin-4, suggesting that GLP-1R G-protein-biased agonists may provide a novel therapeutic approach to T2DM.
Stem Cells, 2006
CD34 and its relatives, podocalyxin and endoglycan, comprise a family of surface sialomucins expr... more CD34 and its relatives, podocalyxin and endoglycan, comprise a family of surface sialomucins expressed by hematopoietic stem/progenitor cells and vascular endothelia. Recent data suggest that they serve as either pro-or antiadhesion molecules depending on their cellular context and their post-translational modifications. In addition, their ability to function as blockers of adhesion may be further regulated by their subcellular localization in membrane microdomains via activation-dependent linkage with the actin cytoskeleton. To gain further insights into the function and regulation of CD34-type molecules, we sought to identify the intracellular ligands that govern their localization. Using both genetic and biochemical approaches, we have identified the Na ؉ /H ؉ exchanger regulatory factor-1 (NHERF-1) as a selective ligand for podocalyxin and endoglycan but not for the closely related CD34. Furthermore, we show that NHERF-1 is expressed by all c-kit ؉ /lineage marker ؊ /Sca-1 ؉ cells, which are known to express podocalyxin and have long-term repopulating abilities. Finally, we show that these proteins relocalize and colocalize in response to cytokine signaling. The results suggest that this cytosolic adaptor protein may be important for mobilization of CD34-type proteins in the plasma membrane and may thereby regulate their ability to block or enhance hematopoietic cell adhesion. STEM
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, Jul 1, 2021
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, Apr 2, 2012
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, Aug 19, 2020
The microenvironment is critical for stem cell maintenance and can be of cellular and non-cellula... more The microenvironment is critical for stem cell maintenance and can be of cellular and non-cellular composition, including secreted growth factors and extracellular matrix (ECM)1–3. Although Notch and other signalling pathways have been reported to regulate quiescence4–9, the composition and source of niche molecules remain largely unknown. Here, we show that adult muscle satellite (stem) cells produce ECM collagens to maintain quiescence cell-autonomously. By ChIP-sequencing we identified NOTCH/RBPJ-bound regulatory elements adjacent to specific collagen genes, whose expression is deregulated in Notch mutant mice. Moreover, we show that satellite cell produced collagen V (COLV) is a critical component of the quiescent niche, as conditional deletion of Col5a1 leads to anomalous cell cycle entry and gradual diminution of the stem cell pool. Notably, the interaction of COLV with satellite cells is mediated by CALCR, for which COLV acts as a surrogate local ligand. Strikingly, systemic administration of a calcitonin derivative is sufficient to rescue the quiescence and self-renewal defects scored in COLV null satellite cells. This study unveils a Notch/COLV/CALCR signalling cascade that cell-autonomously maintains the satellite cell quiescent state and raises the possibility of a similar reciprocal mechanism acting in diverse stem cell populations.
Journal of Neuroendocrinology, Dec 20, 2021
The growth hormone secretagogue receptor 1a (GHSR1a) is intriguing because of its potential as a ... more The growth hormone secretagogue receptor 1a (GHSR1a) is intriguing because of its potential as a therapeutic target and its diverse molecular interactions. Initial studies of the receptor focused on the potential therapeutic ability for growth hormone (GH) release to reduce wasting in aging individuals, as well as food intake regulation for treatment of cachexia. Known roles of GHSR1a now extend to regulation of neurogenesis, learning and memory, gastrointestinal motility, glucose/lipid metabolism, the cardiovascular system, neuronal protection, motivational salience, and hedonic feeding. Ghrelin, the endogenous agonist of GHSR1a, is primarily located in the stomach and is absent from the central nervous system (CNS), including the spinal cord. However, ghrelin in the circulation does have access to a small number of CNS sites, including the arcuate nucleus, which is important in feeding control. At some sites, such as at somatotrophs, GHSR1a has high constitutive activity. Typically, ghrelin‐dependent and constitutive GHSR1a activation occurs via Gαq/11 pathways. In vitro and in vivo data suggest that GHSR1a heterodimerises with multiple G protein‐coupled receptors (GPCRs), including dopamine D1 and D2, serotonin 2C, orexin, oxytocin and melanocortin 3 receptors (MCR3), as well as the MCR3 accessory protein, MRAP2, providing possible mechanisms for its many physiological effects. In all cases, the receptor interaction changes downstream signalling and the responses to receptor agonists. This review discusses the signalling mechanisms of GHSR1a alone and in combination with other GPCRs, and explores the physiological consequences of GHSR1a coupling with other GPCRs.
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, Nov 22, 2022
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, May 23, 2019
The β 2 adrenergic receptor (β 2 AR) signals through both G s and G i in cardiac myocytes, and th... more The β 2 adrenergic receptor (β 2 AR) signals through both G s and G i in cardiac myocytes, and the G i pathway counteracts the G s pathway. However, G i coupling is much less efficient than G s coupling in most cell-based and biochemical assays, making it difficult to study β 2 AR−G i interactions. Here we investigate the role of phospholipid composition on G s and G i coupling. While negatively charged phospholipids are known to enhance agonist affinity and stabilize an active state of the β 2 AR, we find that they impair coupling to G i3 and facilitate coupling to G s. Positively charged Ca 2+ and Mg 2+ , known to interact with the negative charge on phospholipids, facilitates G i3 coupling. Mutational analysis suggests that Ca 2+ coordinates an interaction between phospholipid and the negatively charged EDGE motif on the amino terminal helix of G i3. Taken together, our observations suggest that local membrane charge modulates the interaction between β 2 AR and competing G protein subtypes.
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, Apr 2, 2012
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, May 28, 2020
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, Mar 9, 2015
Therapeutic Advances in Medical Oncology, 2020
Researchers are actively seeking novel targeted therapies for the brain tumour glioblastoma (GBM)... more Researchers are actively seeking novel targeted therapies for the brain tumour glioblastoma (GBM) as the mean survival is less than 15 months. Here we discuss the proposal that the calcitonin receptor (CT Receptor), expressed in 76-86% of patient biopsies, is expressed by both malignant glioma cells and putative glioma stem cells (GSCs), and therefore represents a potential therapeutic target. Forty-two per cent (42%) of high-grade glioma (HGG; representative of GSCs) cell lines express CT Receptor protein. CT Receptors are widely expressed throughout the life cycle of organisms and in some instances promote apoptosis. Which of the common isoforms of the CT Receptor are predominantly expressed is currently unknown, but a functional response to cell stress of the insert-positive isoform is hypothesised. A model for resistant malignancies is one in which chemotherapy plays a direct role in activating quiescent stem cells for replacement of the tumour tissue hierarchy. The putative role that the CT Receptor plays in maintenance of quiescent cancer stem cells is discussed in view of the activation of the Notch-CT Receptor-collagen V axis in quiescent muscle (satellite) stem cells. The pharmacological CT response profiles of four of the HGG cell lines were reported. Both CT responders and non-responders were sensitive to an immunotoxin based on an anti-CT Receptor antibody. The CALCR mRNA exhibits alternative splicing commonly associated with cancer cells, which could result in the atypical pharmacology exhibited by CT non-responders and an explanation of tumour suppression. Due to the inherent instability of CALCR mRNA, analysis of CT Receptor protein in patient samples will lead to improved data for the expression of CT Receptor in GBM and other cancers, and an understanding of the role and activity of the splice variants. This knowledge will aid the effective targeting of this receptor for treatment of GBM.
Neurogastroenterology and Motility, Dec 2, 2020
This is the author manuscript accepted for publication and has undergone full peer review but has... more This is the author manuscript accepted for publication and has undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record. Please cite this article as
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, Dec 18, 2013
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, Jun 2, 2021
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Papers by Sebastian Furness