Improving the traffic operations and safety of full cloverleaf interchanges /

Mode of access: Internet.

GPCR modulation by RAMPs

Our conceptual understanding of the molecular architecture of G-protein coupled receptors (GPCRs) has transformed over the last decade. Once considered as largely independent functional units (aside from their interaction with the G-protein itself), it is now clear that a single GPCR is but part of a multifaceted signaling complex, each component providing an additional layer of sophistication. Receptor activity-modifying proteins (RAMPs) provide a notable example of proteins that interact with GPCRs to modify their function. They act as pharmacological switches, modifying GPCR pharmacology for a particular subset of receptors. However, there is accumulating evidence that these ubiquitous proteins have a broader role, regulating signaling and receptor trafficking. This article aims to provide the reader with a comprehensive appraisal of RAMP literature and perhaps some insight into the impact that their discovery has had on those who study GPCRs. © 2005 Elsevier Inc. All rights reserved.

Heterodimers and family-B GPCRs:RAMPs, CGRP and adrenomedullin

RAMPs (receptor activity-modifying proteins) are single-pass transmembrane proteins that associate with certain family-B GPCRs (G-protein-coupled receptors). Specifically for the CT (calcitonin) receptor-like receptor and the CT receptor, this results in profound changes in ligand binding and receptor pharmacology, allowing the generation of six distinct receptors with preferences for CGRP (CT gene-related peptide) adrenomedullin, amylin and CT. There are three RAMPs: RAMP1-RAMP3. The N-terminus appears to be the main determinant of receptor pharmacology whereas the transmembrane domain contributes to association of the RAMP with the GPCR. The N-terminus of all members of the RAMP family probably contains two disulphide bonds; a potential third disulphide is found in RAMP1 and RAMP3. The N-terminus appears to be in close proximity to the ligand and plays a key role in its binding, either directly or indirectly. BIBN4096BS, a CGRP antagonist, targets RAMP1 and this gives the compound very high selectivity for the human CGRP(1) receptor.

Comparison of the expression of calcitonin receptor-like receptor (CRLR) and receptor activity modifying proteins (RAMPs) with CGRP and adrenomedullin binding in cell lines

1. The calcitonin receptor-like receptor (CRLR) and specific receptor activity modifying proteins (RAMPs) together form receptors for calcitonin gene-related peptide (CGRP) and/or adrenomedullin in transfected cells. 2. There is less evidence that innate CGRP and adrenomedullin receptors are formed by CRLR/RAMP combinations. We therefore examined whether CGRP and/or adrenomedullin binding correlated with CRLR and RAMP mRNA expression in human and rat cell lines known to express these receptors. Specific human or rat CRLR antibodies were used to examine the presence of CRLR in these cells. 3. We confirmed CGRP subtype 1 receptor (CGRP(1)) pharmacology in SK-N-MC neuroblastoma cells. L6 myoblast cells expressed both CGRP(1) and adrenomedullin receptors whereas Rat-2 fibroblasts expressed only adrenomedullin receptors. In contrast we could not confirm CGRP(2) receptor pharmacology for Col-29 colonic epithelial cells, which, instead were CGRP(1)-like in this study. 4. L6, SK-N-MC and Col-29 cells expressed mRNA for RAMP1 and RAMP2 but Rat-2 fibroblasts had only RAMP2. No cell line had detectable RAMP3 mRNA. 5. SK-N-MC, Col-29 and Rat-2 fibroblast cells expressed CRLR mRNA. By contrast, CRLR mRNA was undetectable by Northern analysis in one source of L6 cells. Conversely, a different source of L6 cells had mRNA for CRLR. All of the cell lines expressed CRLR protein. Thus circumstances where CRLR mRNA is apparently absent by Northern analysis do not exclude the presence of this receptor. 6. These data strongly support CRLR, together with appropriate RAMPs as binding sites for CGRP and adrenomedullin in cultured cells.

Modulating receptor function through RAMPs:can they represent drug targets in themselves?

G protein-coupled receptors (GPCRs) are successfully exploited as drug targets. As our understanding of how distinct GPCR subtypes can be generated expands, so do possibilities for therapeutic intervention via these receptors. Receptor activity-modifying proteins (RAMPs) are excellent examples of proteins that enhance diversity in. GPCR function. They facilitate the creation of binding pockets, controlling the pharmacology of some GPCRs. Moreover, they have the ability to regulate cell-surface trafficking, internalisation and signalling of GPCRs, creating novel opportunities for drug discovery. RAMPs could be directly targeted by drugs, or advantage could be taken of unique RAMP/GPCR interfaces for generating highly selective ligands.

RAMPs and the modulation of G-protein coupled receptors

Receptor activity modifying proteins (RAMPs) are a family of three proteins that can interact with G-protein coupled receptors (GPCRs) to create new receptors and also alter the signalling and trafficing of existing receptors. There are frequently changes in RAMP expression in cardiovascular disease RAMPs represent important drug targets.

RAMPs and CGRP receptors

Receptor activity modifying protein 1 (RAMP1) forms a complex with calcitonin receptor-like receptor (CLR) to produce the receptor for calcitonin gene-related peptide (CGRP). RAMP1 has two main roles. It facilitates the cell-surface expression of CLR. It is also essential for the binding of CGRP to the receptor. It seems likely that Y66, F93, H97 and F101, amongst other residues, form a binding site for CLR. These cluster together on the same face of the extracellular portion of RAMP1, probably close to where it enters the plasma membrane. Residues at the other end of RAMP1 are most likely to be involved in CGRP recognition, although it is currently unclear how they do this. Within this area, W74 is important for the binding of the nonpeptide antagonist, BIBN4096BS, although it does not seem to be involved in the binding of CGRP itself. It has been shown that there is an epitope within residues 23-60 of CLR that are essential for RAMP recognition. Under some circumstances, changes in the expression of RAMP1 can alter the sensitivity of cells to CGRP, demonstrating that regulation of its levels may be of physiological or pathophysiological importance.

RAMPs as drug targets

The receptor activity-modifying protein (RAMP) family of membrane proteins regulates G protein-coupled receptor (GPCR) function in several ways. RAMPs can alter their pharmacology and signalling as well as the trafficking of these receptors to and from the cell surface. Accordingly, RAMPs may be exploited as drug targets, offering new opportunities for regulating the function of therapeutically relevant RAMP-interacting GPCRs. For example, several small molecule antagonists of RAMP1/ calcitonin receptor-like receptor complexes, which block the actions of the neuropeptide calcitonin gene-related peptide are in development for the treatment of migraine headache.

Receptor Activity Modifying Proteins (RAMPs) interact with the VPAC2 Receptor and CRF1 receptors and modulate their function

Background and Purpose Although it is established that the receptor activity modifying proteins (RAMPs) can interact with a number of GPCRs, little is known about the consequences of these interactions. Here the interaction of RAMPs with the glucagon-like peptide 1 receptor (GLP-1 receptor), the human vasoactive intestinal polypeptide/pituitary AC-Activating peptide 2 receptor (VPAC) and the type 1 corticotrophin releasing factor receptor (CRF) has been examined. Experimental Approach GPCRs were co-transfected with RAMPs in HEK 293S and CHO-K1 cells. Cell surface expression of RAMPs and GPCRs was examined by elisa. Where there was evidence for interactions, agonist-stimulated cAMP production, Ca mobilization and GTPγS binding to G, G, G and G were examined. The ability of CRF to stimulate adrenal corticotrophic hormone release in Ramp2 mice was assessed. Key Results The GLP-1 receptor failed to enhance the cell surface expression of any RAMP. VPAC enhanced the cell surface expression of all three RAMPs. CRF enhanced the cell surface expression of RAMP2; the cell surface expression of CRF was also increased. There was no effect on agonist-stimulated cAMP production. However, there was enhanced G-protein coupling in a receptor and agonist-dependent manner. The CRF: RAMP2 complex resulted in enhanced elevation of intracellular calcium to CRF and urocortin 1 but not sauvagine. In Ramp2 mice, there was a loss of responsiveness to CRF. Conclusions and Implications The VPAC and CRF receptors interact with RAMPs. This modulates G-protein coupling in an agonist-specific manner. For CRF, coupling to RAMP2 may be of physiological significance. © 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.

The relationship of boat movements to environmental management at some Queensland marinas

Marinas currently exist primarily to service recreational boats, and these vessels are a potential cause of both problems and opportunities in environmental management. Thus, on the one hand, destructive fuel and other pollutants may be expelled, boat wakes can cause littoral soil erosion, physical damage results from collisions with marine life, and litter and noise pollution occur in otherwise pristine habitat. Boats also provide access to otherwise inaccessible natural environments for educational and other management reasons. In this study, boat traffic at three large marinas located along the Queensland coastline has been field surveyed for introductory information. No attempt was made at this juncture to survey the behaviour of the boat crews and passengers (concerning actual destinations, activities on board, etc. or to survey the recreational boat industry. Such studies rely on boat registration records and personal questionnaires. Some other surveys relating to fishing draw on boat ramp surveys and direct submissions by recreational fishers; these provide some data on daily usage of boat ramps, but without particular attention to boats. We believe field observations of overall boat activities in the water are necessary for environmental management purposes. The aim of the survey was to provide information to help prioritize the potential impacts that boats’ activities have on the surrounding natural environment. Any impact by boats will be a product of their numbers, size, frequency of movement, carrying capacity and routes/destinations. The severity of impacts will dictate the appropriate management action.

Differences in the performance of safety performance functions estimated for total crash count and for crash count by crash type

In recent years the development and use of crash prediction models for roadway safety analyses have received substantial attention. These models, also known as safety performance functions (SPFs), relate the expected crash frequency of roadway elements (intersections, road segments, on-ramps) to traffic volumes and other geometric and operational characteristics. A commonly practiced approach for applying intersection SPFs is to assume that crash types occur in fixed proportions (e.g., rear-end crashes make up 20% of crashes, angle crashes 35%, and so forth) and then apply these fixed proportions to crash totals to estimate crash frequencies by type. As demonstrated in this paper, such a practice makes questionable assumptions and results in considerable error in estimating crash proportions. Through the use of rudimentary SPFs based solely on the annual average daily traffic (AADT) of major and minor roads, the homogeneity-in-proportions assumption is shown not to hold across AADT, because crash proportions vary as a function of both major and minor road AADT. For example, with minor road AADT of 400 vehicles per day, the proportion of intersecting-direction crashes decreases from about 50% with 2,000 major road AADT to about 15% with 82,000 AADT. Same-direction crashes increase from about 15% to 55% for the same comparison. The homogeneity-in-proportions assumption should be abandoned, and crash type models should be used to predict crash frequency by crash type. SPFs that use additional geometric variables would only exacerbate the problem quantified here. Comparison of models for different crash types using additional geometric variables remains the subject of future research.