Professor Helen Cox
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Dr Iain Tough / Postdoc
Ms Sian Fairbrother / PhD student
Ms Alison Ray / PhD student
Ms Sian Fairbrother / PhD student
Ms Alison Ray / PhD student
Enteric Peptides and their Receptors
The enteric nervous system (ENS) with entero-endocrine cells, detect and integrate information within the intestinal wall, in order to respond appropriately to luminal contents. These local signals control digestion, epithelial ion secretion, motility, vascular perfusion and tissue defence, and when signalling abnormalities occur the consequences can be fatal, e.g. cholera toxin infection or debilitating as in irritable bowel syndrome (IBS). The ENS is also plastic, changing with age and it can be hijacked by infectious agents e.g. prion protein, providing a potential route of infectivity to the brain (see Professor Morris’s summary). Elucidating how neurohumoral peptides alter intestinal function at tissue, cell and molecular levels will provide a rational basis for receptor-targeted drugs against specific bowel disorders.
Enteric peptides and their functional roles
The ENS comprises excitatory and inhibitory motor, secretomotor and sensory neurons each releasing transmitters that differentially stimulate receptors with distinct expression patterns and functions. Prominent amongst the G protein-coupled receptor- (GPCR) activating peptides are neuropeptide Y (NPY, Gi-coupled) and neurokinin (NK, Gq-coupled) peptides. NPY is involved in intestinal ion absorption, obesity and metabolism, all roles mediated by Y receptors. Three Y receptors stimulate the broad-spectrum inhibition of ion secretion in human colon;- Y1, Y2 and Y4, the latter providing a gut-specific therapeutic target. NK's and their receptors (NK1, NK2 and NK3) in contrast, are primarily involved in sensory neurotransmission and are up-regulated at sites of inflammation. Thus NK antagonists are predicted gut anti-inflammatories while Y agonists are potential anti-diarrhoeals.
The ENS comprises excitatory and inhibitory motor, secretomotor and sensory neurons each releasing transmitters that differentially stimulate receptors with distinct expression patterns and functions. Prominent amongst the G protein-coupled receptor- (GPCR) activating peptides are neuropeptide Y (NPY, Gi-coupled) and neurokinin (NK, Gq-coupled) peptides. NPY is involved in intestinal ion absorption, obesity and metabolism, all roles mediated by Y receptors. Three Y receptors stimulate the broad-spectrum inhibition of ion secretion in human colon;- Y1, Y2 and Y4, the latter providing a gut-specific therapeutic target. NK's and their receptors (NK1, NK2 and NK3) in contrast, are primarily involved in sensory neurotransmission and are up-regulated at sites of inflammation. Thus NK antagonists are predicted gut anti-inflammatories while Y agonists are potential anti-diarrhoeals.
Figure 1 The GPCR lifecycle
The receptor C terminus governs its organisation in the plasma membrane, endocytosis, recycling and degradation. Some of the protein modifications and interactions involved in the receptor's life cycle are highlighted. Drawing by Dr Nicholas Holliday
Receptor targeting, signalling and recycling
GPCRs provide a major molecular recognition mechanism for nutrients, neurotransmitters and hormones. We are interested in how specific GPCRs are targeted to the cell surface, and what controls their recycling and degradation (Figure 1). For example, the C-terminus of the Y1 receptor is crucial for correct signalling and trafficking after NPY stimulation (Figure 2). When removed, the resulting truncated receptor continuously internalises and recycles. In contrast, a GPCR sensitive to the stomach hormone ghrelin, is constitutively active (i.e. it signals without agonist) and endocytosed. Here internalisation can be blocked by a negative agonist that turns off the receptor (Figure 2). Identifying the protein motifs that act as switches to guide GPCRs along these pathways is of fundamental importance in predicting drug effectiveness in the long term (e.g. potential NPY and ghrelin receptor ligands for obesity).
Receptor targeting, signalling and recycling
GPCRs provide a major molecular recognition mechanism for nutrients, neurotransmitters and hormones. We are interested in how specific GPCRs are targeted to the cell surface, and what controls their recycling and degradation (Figure 1). For example, the C-terminus of the Y1 receptor is crucial for correct signalling and trafficking after NPY stimulation (Figure 2). When removed, the resulting truncated receptor continuously internalises and recycles. In contrast, a GPCR sensitive to the stomach hormone ghrelin, is constitutively active (i.e. it signals without agonist) and endocytosed. Here internalisation can be blocked by a negative agonist that turns off the receptor (Figure 2). Identifying the protein motifs that act as switches to guide GPCRs along these pathways is of fundamental importance in predicting drug effectiveness in the long term (e.g. potential NPY and ghrelin receptor ligands for obesity).
Figure 2. Studying the trafficking of GPCRs and their mutants.
Green-labelled Y1 receptors in HEK293 cells mainly on the cell surface are rapidly internalised in response to NPY (B), where they colocalise with a red marker (transferrin) for recycling endosomes. Truncation of the Y1 C terminus (S352*) generates a constitutively internalising receptor (C) whose trafficking is retarded at the plasma membrane after NPY treatment (D). In contrast, the spontaneously active ghrelin receptor undergoes constitutive internalisation (E) which is blocked by a negative agonist (SP-A; F). scale bar = 10 µm.
Epithelial luminal sensors
In polarised cells GPCRs are targeted to specific domains in the plasma membrane, for example dendrites or axons in neurones, and apical or basolateral membranes in epithelial cells. Many epithelial GPCRs are basolateral, but some act as apical sensors for the luminal nutrient environment. Preliminary studies with an intestinal receptor GPR40 sensitive to mid/long chain fatty acids, show that it is targeted towards the apical domain in epithelia and may thus provide a mechanism by which fatty acids stimulate epithelial/mucosal responses in the gut. Future studies will determine how this receptor is processed differently from other basolateral GPCRs, and how it signals in both epithelial and endocrine cells.
In polarised cells GPCRs are targeted to specific domains in the plasma membrane, for example dendrites or axons in neurones, and apical or basolateral membranes in epithelial cells. Many epithelial GPCRs are basolateral, but some act as apical sensors for the luminal nutrient environment. Preliminary studies with an intestinal receptor GPR40 sensitive to mid/long chain fatty acids, show that it is targeted towards the apical domain in epithelia and may thus provide a mechanism by which fatty acids stimulate epithelial/mucosal responses in the gut. Future studies will determine how this receptor is processed differently from other basolateral GPCRs, and how it signals in both epithelial and endocrine cells.