The Experts below are selected from a list of 714 Experts worldwide ranked by ideXlab platform
Robert M Dores - One of the best experts on this subject based on the ideXlab platform.
-
ACTH, Melanocortin Receptors, MRAP Accessory Proteins
Encyclopedia of Endocrine Diseases, 2020Co-Authors: Robert M DoresAbstract:The evolution of the pituitary hormone, ACTH, the Melanocortin Receptors, a family of G protein-coupled Receptors, and the Melanocortin Receptor accessory proteins, MRAP1 and MRAP2, are intertwined, and parallel the evolution of the chordates. While all Melanocortin-related peptides (i.e., ACTH, αMSH, βMSH, γMSH, δMSH) have the canonical HFRW amino acid motif required for activation of Melanocortin Receptors (i.e., MC1R, MC2R, MC3R, MC4R, MC5R), during the radiation of the gnathostomes (jawed vertebrates) the ligand selectivity of Melanocortin-2 Receptor orthologs has diverged between cartilaginous fishes and bony vertebrates in a manner that has implications for the Hypothalamus/Pituitary/Adrenal-Interrenal axis.
-
pharmacological properties of whale shark rhincodon typus Melanocortin 2 Receptor and melancortin 5 Receptor interaction with mrap1 and mrap2
General and Comparative Endocrinology, 2020Co-Authors: Brianne Hoglin, Marin Miner, Robert M DoresAbstract:Abstract In the current study, the whale shark (ws; Rhincodon typus) Melanocortin-2 Receptor (MC2R) co-expressed with wsMRAP1 in Chinese Hamster Ovary (CHO) Cells could be stimulated in a dose dependent manner by ACTH(1–24) with an EC50 of 2.6 × 10−10 M ± 9.7 × 10−11. When the Receptor was expressed alone, stimulation was only observed at [10−6 M]. A comparable increase in sensitivity to stimulation by srDes-Ac-αMSH was also observed when the Receptor was co-expressed with wsMRAP1. In addition, co-expression with wsMRAP1 significantly increased the trafficking of wsMC2R to the plasma membrane of CHO cells. Surprisingly, co-expression with wsMRAP2 also increased sensitivity to stimulation by ACTH(1–24) and srDes-Ac-αMSH, and increased trafficking of the Receptor to the plasma membrane. These observations are in sharp contrast to the response of MC2R orthologs of bony vertebrates which have an obligate requirement for co-expression with MRAP1 for both trafficking to the plasma membrane and activation, and while co-expression with MRAP2 increases trafficking, it has minimal effects on activation. In addition, when comparing the activation features of wsMC2R with those of the elephant shark MC2R and red stingray MC2R orthologs, both similarities and differences are observed. The spectrum of features for cartilaginous fish MC2R orthologs will be discussed. A second objective of this study was to determine whether wsMC5R has features in common with wsMC2R in terms of ligand selectivity and interaction with wsMRAP paralogs. While wsMC5R can be activated by either srACTH(1–24) or srDes-Ac-αMSH, and co-expression with wsMRAP1 enhances this activation, wsMRAP1 had no effect on the trafficking of wsMC5R. Co-expression with wsMRAP2 had no positive or negative effect on either ligand sensitivity or trafficking of wsMC5R.
-
evidence for diversity in the activation of the Melanocortin 2 Receptor a study on gar elephant shark and stingray mc2rs
Peptides, 2020Co-Authors: Brianne Hoglin, Amanda Ferguson, Sheila Pahlavan, Robert M DoresAbstract:Abstract The Melanocortin-2 Receptor (MC2R) is a critical component of the HPI and HPA axes of cartilaginous fishes, teleosts and tetrapods. Studies on teleost and tetrapod orthologs suggest two contact sites between ACTH and the Receptor involving the following motifs on ACTH: H6F7R8W9 and K15K16R17R18P19. Using spotted gar (g) MC2R as a representative bony fish MC2R ortholog, we found that activation of gMC2R in Chinese Hamster Ovary (CHO) cells was diminished following stimulation of the transfected cells with hACTH(1–24) analogs substituted with alanine at either the H6F7R8W9 or K15K16R17R18P19 motifs compared to stimulation with hACTH(1–24). This observation suggests two ligand contact sites necessary for activation of the gMC2R. The same experiments were done with elephant shark (es) MC2R, however only the H6F7R8W9 analogs blocked activation, pointing to a single contact on esMC2R. Conversely, the red stingray (sr) MC2R activation was blocked by both the H6F7R8W9 and K15K16R17R18P19 alanine-substituted analogs. Together these results build a picture of the evolution of the ligand and Receptor interaction between ACTH and MC2R orthologs of different taxa. These results will be discussed in light of the parallel evolution of MC2R orthologs in cartilaginous fishes and bony vertebrates.
-
identifying common features in the activation of Melanocortin 2 Receptors studies on the xenopus tropicalis Melanocortin 2 Receptor
International Journal of Molecular Sciences, 2019Co-Authors: Perry E Davis, Emily C Wilkinson, Robert M DoresAbstract:The interaction between the pituitary hormone, adrenocorticotropin (ACTH), and Melanocortin-2 Receptor (MC2R) orthologs involves the H6 F7 R8 W9 and R/K15 K16 R17 R18 motifs in ACTH making contact with corresponding contact sites on MC2R. Earlier studies have localized the common HFRW binding site of all Melanocortin Receptors to residues in TM2, TM3, and TM6 that are located close to the extracellular space. The current study has identified residues in Xenopus tropicalis (xt) MC2R in TM4 (I158, F161), in EC2 (M166), and in TM5 (V172) that also are involved in activation of xtMC2R, and may be in the R/KKRR contact site of xtMC2R. These results are compared to earlier studies on the corresponding domains of human MC2R and rainbow trout MC2R in an effort to identify common features in the activation of teleost and tetrapod MC2R orthologs following stimulation with ACTH.
-
cadmium disrupts Melanocortin 2 Receptor signaling in rainbow trout
Aquatic Toxicology, 2019Co-Authors: Navdeep Sandhu, Liang Liang, Robert M Dores, James C Mcgeer, Mathilakath M VijayanAbstract:Abstract Cadmium is an endocrine disruptor and inhibits corticosteroid production, but the mechanisms are far from clear. We tested the hypothesis that sublethal exposure to environmentally realistic levels of cadmium impairs cortisol production by disrupting the Melanocortin 2 Receptor (MC2R) signaling in rainbow trout (Oncorhynchus mykiss). Fish were exposed to sublethal concentrations of cadmium (0.75 or 2.0 μg/L) in a flow-through system for 7 d and subjected to an acute secondary stressor to evoke a cortisol response. Cadmium exposure for 7 d did not affect plasma cortisol concentrations, but head kidney mc2r mRNA levels were higher than in control fish. The cortisol stress performance to a secondary-stressor was attenuated in the cadmium groups, and this corresponded with transient reduction in transcript abundance of mc2r and the gene encoding its accessory protein MRAP1 but not MRAP2 in the head kidney. Furthermore, in vivo cadmium exposure attenuated the adrenocorticotropic hormone (ACTH)-, but not 8-br-cAMP-stimulated cortisol production in head kidney slices ex vivo. This corresponded with reduced transcript abundance of mc2r and mrap1, but not mrap2 in these tissue slices. Also, reporter assays with CHO cells transiently transfected with rainbow trout mc2r and zebrafish mrap1 revealed a dose-independent inhibition in ACTH-stimulated luciferase activity by cadmium. Collectively, waterborne exposure to environmentally realistic concentration of cadmium compromises the stressor-induced cortisol response, and a mode of action involves the disruption of MC2R signaling in rainbow trout.
Adrian J L Clark - One of the best experts on this subject based on the ideXlab platform.
-
The Melanocortin-2 Receptor in Normal Adrenocortical Function and Familial Adrenocorticotropic Hormone Resistance
The Melanocortin Receptors, 2020Co-Authors: Adrian J L ClarkAbstract:The physiologic role of adrenocorticotropic hormone (ACTH) and its part in the pituitary—adrenal axis is one of the most intensively studied systems in endocrinology. ACTH was one of the first hormones that was found to stimulate cAMP production by the adrenal gland (1), and the notion that this effect was mediated via a specific cell surface Receptor was confirmed by the elegant studies of Lefkowitz and colleagues (2) in work that set a standard for Receptor characterization. Nevertheless, progress on the understanding of the ACTH Receptor has been relatively slow. It is now clear that the MC2-R is synonymous with the ACTH Receptor, and both terms are used in this chapter. In general, the term ACTH Receptor is used to describe the functional entity for example, ligand binding to adrenal cells, whereas the term MC2-R is used to describe aspects that can clearly be related to this gene.
-
Stability and Turnover of the ACTH Receptor Complex.
Frontiers in Endocrinology, 2019Co-Authors: Adrian J L Clark, Li F ChanAbstract:Glucocorticoid production in mammals is principally regulated by the action of the pituitary hormone adrenocorticotropin (ACTH) acting on its cognate membrane Receptor on the zona fasciculata cells of the adrenal cortex. The Receptor for ACTH consists of two essential components, a small seven transmembrane domain G protein-coupled Receptor of the Melanocortin Receptor subgroup known as the Melanocortin 2 Receptor (MC2R) and a small single transmembrane domain protein that adopts a antiparallel homodimeric form and which is known as the Melanocortin 2 Receptor accessory protein (MRAP). MRAP is essential for the trafficking of the MC2R to the cell surface as well as being required for Receptor responsiveness to ACTH at physiological concentrations – probably by facilitating ACTH binding, but possibly also by supporting G protein interaction with the MC2R. A number of studies have shown that ACTH stimulates the expression of functional Receptor at the cell surface and the transcription of both MC2R and MRAP mRNA. However the time course of these transcriptional effects differs such that MRAP is expressed relatively rapidly whereas MC2R transcription responds much more slowly. Furthermore, recent data suggests that MRAP protein is turned over with a short half-life whereas MC2R has a significantly longer half-life. These findings imply that these two ACTH Receptor proteins have distinct trajectories and that it is likely that MRAP-independent MC2R is present at the cell surface. In such a situation newly transcribed and translated MRAP could enable the rapid recruitment of functional Receptor at the plasma membrane without the need for new MC2R translation. This may be advantageous in circumstances of significant stress in that the potentially complex and perhaps inefficient process of de novo MC2R translation, folding, post-translational modification and trafficking can be avoided.
-
MRAP Deficiency Impairs Adrenal Progenitor Cell Differentiation and Gland Zonation
The FASEB Journal, 2018Co-Authors: Tatiana V. Novoselova, Adrian J L Clark, Peter J King, Leonardo Guasti, Louise A Metherell, Mashal Hussain, Marika Charalambous, Li F ChanAbstract:Melanocortin 2 Receptor accessory protein (MRAP) is a single transmembrane domain accessory protein and a critical component of the hypothamo-pituitary-adrenal axis. MRAP is highly expressed in the...
-
Melanocortin 4 Receptor becomes an acth Receptor by coexpression of Melanocortin Receptor accessory protein 2
Molecular Endocrinology, 2013Co-Authors: Maria Josep Agulleiro, Raúl Cortés, Raúl Guillot, Adrian J L Clark, Begona Fernandezduran, S Navarro, Eirini Meimaridou, Jose Miguel CerdareverterAbstract:Melanocortin 2 Receptor (MC2R) is the only canonical ACTH Receptor. Its functional expression requires the presence of an accessory protein, known as Melanocortin Receptor 2 accessory protein 1 (MRAP1). The vertebrate genome exhibits a paralogue gene called MRAP2, which is duplicated in zebrafish (MRAP2a and MRAP2b), although its function remains unknown. In this paper, we demonstrate that MRAP2a enables MC4R, a canonical MSH Receptor, to be activated by ACTH with a similar sensitivity to that exhibited by MC2R. Both proteins physically interact and are coexpressed in the neurons of the preoptic area, a key region in the control of the energy balance and hypophyseal secretion in fish. ACTH injections inhibit food intake in wild-type zebrafish but not in fish lacking functional MC4R. Both MRAP1 and MRAP2a are hormonally regulated, suggesting that these proteins are substrates for feed-back regulatory pathways of Melanocortin signaling. Fasting has no effect on the central expression of MRAP2a but stimulate...
-
neonatal presentation of familial glucocorticoid deficiency resulting from a novel splice mutation in the Melanocortin 2 Receptor accessory protein
European Journal of Endocrinology, 2011Co-Authors: Vandana Jain, Adrian J L Clark, Louise A Metherell, Alessia David, Rajni Sharma, Pradeep Sharma, Li F ChanAbstract:Background: Familial glucocorticoid deficiency (FGD) is a rare autosomal recessive disorder characterised by isolated glucocorticoid deficiency. Mutations in the ACTH Receptor/Melanocortin 2 Receptor (MC2R), the MC2R accessory protein (MRAP) or the STAR protein (STAR) cause FGD types 1, 2 and 3, respectively, accounting for w50% of all cases. Patient and methods: We report a neonate of Indian origin, who was diagnosed with FGD in the first few days of life. He presented with hypoglycaemic seizures and was noted to have generalised intense hyperpigmentation and normal male genitalia. Biochemical investigations revealed hypocortisolaemia (cortisol 0.223 mg/dl; NR 1‐23 mg/dl) and elevated plasma ACTH (170 pg/ml). Serum electrolytes, aldosterone and plasma renin activity were normal. Peak cortisol following a standard synacthen test was 0.018 mg/dl. He responded to hydrocortisone treatment and continues on replacement. Patient DNA was analysed by direct sequencing. The effect of the novel mutation was assessed by an in vitro splicing assay using wild type and mutant heterologous minigenes. Results: A novel homozygous mutation c.106C2_3dupTAwas found in theMRAP gene. Both parents were heterozygous for the mutation. In an invitro splicing assay, the mutation resulted in the skipping of exon 3. Conclusion :W e have identif ied an ovelMRAP mutation where disruption of the intron 3 splice-site results in a prematurely terminated translation product. This protein (if produced) would lack the transmembrane domain that is essential for MC2R interaction. We predict that this would cause complete lack of ACTH response thus explaining the early presentation in this case.
Liang Liang - One of the best experts on this subject based on the ideXlab platform.
-
cadmium disrupts Melanocortin 2 Receptor signaling in rainbow trout
Aquatic Toxicology, 2019Co-Authors: Navdeep Sandhu, Liang Liang, Robert M Dores, James C Mcgeer, Mathilakath M VijayanAbstract:Abstract Cadmium is an endocrine disruptor and inhibits corticosteroid production, but the mechanisms are far from clear. We tested the hypothesis that sublethal exposure to environmentally realistic levels of cadmium impairs cortisol production by disrupting the Melanocortin 2 Receptor (MC2R) signaling in rainbow trout (Oncorhynchus mykiss). Fish were exposed to sublethal concentrations of cadmium (0.75 or 2.0 μg/L) in a flow-through system for 7 d and subjected to an acute secondary stressor to evoke a cortisol response. Cadmium exposure for 7 d did not affect plasma cortisol concentrations, but head kidney mc2r mRNA levels were higher than in control fish. The cortisol stress performance to a secondary-stressor was attenuated in the cadmium groups, and this corresponded with transient reduction in transcript abundance of mc2r and the gene encoding its accessory protein MRAP1 but not MRAP2 in the head kidney. Furthermore, in vivo cadmium exposure attenuated the adrenocorticotropic hormone (ACTH)-, but not 8-br-cAMP-stimulated cortisol production in head kidney slices ex vivo. This corresponded with reduced transcript abundance of mc2r and mrap1, but not mrap2 in these tissue slices. Also, reporter assays with CHO cells transiently transfected with rainbow trout mc2r and zebrafish mrap1 revealed a dose-independent inhibition in ACTH-stimulated luciferase activity by cadmium. Collectively, waterborne exposure to environmentally realistic concentration of cadmium compromises the stressor-induced cortisol response, and a mode of action involves the disruption of MC2R signaling in rainbow trout.
-
the Melanocortin 2 Receptor of the rainbow trout identifying a role for critical positions in transmembrane domain 4 extracellular loop 2 and transmembrane domain 5 in the activation of rainbow trout mc2r
General and Comparative Endocrinology, 2017Co-Authors: Liang Liang, Perry Davis, Michael R Dores, Robert M DoresAbstract:Abstract The activation of either teleost or tetrapod Melanocortin-2 Receptor (MC2R) orthologs requires interaction between the HFRW motif and R/KKRRP motif in the primary sequence of ACTH, and two corresponding sites on the Melanocortin 2 Receptor. While the HFRW contact site on MC2R appears to involve residues in TM2, TM3, and TM6, several studies on human MC2R point to the EC2/TM5 region of MC2R as a possible location for the R/KKRRP contact site. In this study nineteen single-alanine mutants of rainbow trout (rt) MC2R were made beginning at V 153 in TM4, at all positions in EC2 (extracellular loop 2), to F 175 in TM5. For twelve of these alanine mutants (i.e., V 153 , G 155 , C 162 , D 163 , T 165 , V 166 , I 167 , H 169 , F 170 , H 172 , V 173 , L 174 ), alanine substitution did not have a statistically significant effect on activation of the Receptor. For four of these alanine mutations (i.e., V 157 , M 158 , F 161 , K 168 ), while the negative shift in ligand sensitivity was statistically significant, the magnitude of the negative shift in activation was fivefold or less. However, for substitution at V 159 in TM4 (negative shift in activation: 110 fold), F 171 in TM5 (negative shift in activation: 48-fold), and F 175 in TM5 (negative shift in activation: 100 fold), the effect on activation was both statistically significant and may be physiologically relevant. To support this conclusion, a triple alanine mutant of rtMC2R (V 159 /A, F 171 /A, F 175 /A), and this mutant Receptor could not be activated by ACTH at concentrations as high as 10 −6 M. A Cell Surface ELISA analysis indicated that the trafficking of the triple alanine mutant rtMC2R to the plasma membrane was not impaired by the alanine substitutions. Collectively, these observations point to a critical role for TM4 and TM5 in the activation of the rainbow trout Melanocortin-2 Receptor.
-
identifying the activation motif in the n terminal of rainbow trout and zebrafish Melanocortin 2 Receptor accessory protein 1 mrap1 orthologs
General and Comparative Endocrinology, 2016Co-Authors: Robert M Dores, Liang Liang, Navdeep Sandhu, Rebecca E Hollmann, Mathilakath M VijayanAbstract:The activation of mammalian Melanocortin-2 Receptor (MC2R) orthologs is dependent on a four-amino acid activation motif (LDYL/I) located in the N-terminal of mammalian MRAP1 (Melanocortin-2 Receptor accessory protein). Previous alanine substitution analysis had shown that the Y residue in this motif appears to be the most important for mediating the activation of mammalian MC2R orthologs. Similar, but not identical amino acid motifs were detected in rainbow trout MRAP1 (YDYL) and zebrafish MRAP1 (YDYV). To determine the importance of these residues in the putative activation motifs, rainbow trout and zebrafish MRAP1 orthologs were individually co-expressed in CHO cells with rainbow trout MC2R, and the activation of this Receptor with either the wild-type MRAP1 ortholog or alanine-substituted analogs of the two teleost MRAP1s was analyzed. Alanine substitutions at all four amino acid positions in rainbow trout MRAP1 blocked activation of the rainbow trout MC2R. Single alanine substitutions of the D and Y residues in rainbow trout and zebrafish MRAP1 indicate that these two residues play a significant role in the activation of rainbow trout MC2R. These observations indicate that there are subtle differences in the way that teleost and mammalian MRAPs are involved in the activation of their corresponding MC2R orthologs.
-
structure function studies on the activation of the rainbow trout Melanocortin 2 Receptor
General and Comparative Endocrinology, 2015Co-Authors: Liang Liang, Joseph K Angleson, Kristin Schmid, Navdeep Sandhu, Mathilakath M Vijayan, Robert M DoresAbstract:Abstract Functional expression of the rainbow trout (rt) Melanocortin-2 Receptor (MC2R) in CHO cells requires co-expression with a teleost Melanocortin-2 Receptor accessory protein (MRAP) such as zebrafish (zf) MRAP. Transiently transfected rtMC2R/zfMRAP1 CHO cells were used to evaluate the efficacy of alanine substituted analogs of hACTH(1-24) in three motifs in the ligand: H6F7R8W9, G10K11P12V13G14, and K15K16R17R18P19. Alanine substitution at all positions in each motif either completely blocked activation of the Receptor (H6F7R8W9 and K15K16R17R18P19) or resulted in just over 400 fold increase in EC50 value (G10K11P12V13G14). Single alanine substitutions in the H6F7R8W9 motif indicated that substitution at either W9 or R8 resulted in a much larger increase in EC50 values as compared to substitutions at either F7 or W9. Alanine substitution at either K15K16 or R17R18P19 in the K15K16R17R18P19 motif resulted in a statistically equivalent increase in EC50 value of at least 600 fold. Finally, alanine substitutions in the G10K11P12V13G14 motif resulted in increases in EC50 values presumably as a result of altering the secondary structure of the ligand. However, truncated analogs of hACTH(1-24) in which either G10G14 (ACTH(1-22), or K11P12V13 (ACTH(1-21) were removed had no stimulatory activity. Finally, some of the hACTH(1-24) analogs were tested using rainbow trout head kidney pieces in vitro to confirm whether the response to analogs seen with the transient transfected rtMC2R CHO cells was similar to that of trout interrenal cells. The results of these alanine substitution analog studies are used to construct a multistep hypothetical model for the activation of teleost and tetrapod MC2Rs to account for the unique ligand selectivity of this Receptor.
-
analyzing the activation of the Melanocortin 2 Receptor of tetrapods
General and Comparative Endocrinology, 2014Co-Authors: Robert M Dores, Liang LiangAbstract:Abstract Following the biochemical characterization of the pituitary hormone, adrenocorticotropin (ACTH), in the 1950’s, a number of structure/function studies were done which identifies two amino acid motifs in ACTH, the HFRW motif and KKRR motif, as critical for the activation of the “ACTH” Receptor on adrenal cortex cells. In the 1990’s the “ACTH” Receptor was identified as a member of the Melanocortin Receptor gene family, and given the name Melanocortin-2 Receptor (MC2R). Since that time a number of studies on both tetrapod and teleost MC2R orthologs have established that these orthologs can only be activated by ACTH, but not by any of the MSH-sized Melanocortin ligands, and these orthologs require interaction with the Melanocortin-2 Receptor accessory protein (MRAP) for functional expression. This review summarizes recent structure/function studies on human ACTH, and points out the importance of the GKPVG motif in ACTH for the activation of the Receptor. In this regard, a multiple-step model for the activation of tetrapod and teleost MC2R orthologs is presented, and the evolution of gnathostome MC2R ligand selectivity and the requirement for MRAP interaction is discussed in light of a recent study on a cartilaginous fish MC2R ortholog. This review contains excerpts from the Gorbman/Bern Lecture presented at the Second Meeting of the North American Society for Comparative Endocrinology (NASCE).
Peter J King - One of the best experts on this subject based on the ideXlab platform.
-
MRAP Deficiency Impairs Adrenal Progenitor Cell Differentiation and Gland Zonation
The FASEB Journal, 2018Co-Authors: Tatiana V. Novoselova, Adrian J L Clark, Peter J King, Leonardo Guasti, Louise A Metherell, Mashal Hussain, Marika Charalambous, Li F ChanAbstract:Melanocortin 2 Receptor accessory protein (MRAP) is a single transmembrane domain accessory protein and a critical component of the hypothamo-pituitary-adrenal axis. MRAP is highly expressed in the...
-
localisation of the Melanocortin 2 Receptor and its accessory proteins in the developing and adult adrenal gland
Journal of Molecular Endocrinology, 2011Co-Authors: Rebecca J Gorrigan, Peter J King, Adrian J L Clark, Leonardo Guasti, Li F ChanAbstract:The Melanocortin-2-Receptor (MC2R)/MC2R accessory protein (MRAP) complex is critical to the production of glucocorticoids from the adrenal cortex. Inactivating mutations in either MC2R or MRAP result in the clinical condition familial glucocorticoid deficiency. The localisation of MC2R together with MRAP within the adrenal gland has not previously been reported. Furthermore, MRAP2, a paralogue of MRAP, has been shown in vitro to have a similar function to MRAP, facilitating MC2R trafficking and responsiveness to ACTH. Despite similar MC2R accessory functions, in vivo, patients with inactivating mutations of MRAP fail to be rescued by a functioning MRAP2 gene, suggesting differences in adrenal expression, localisation and/or function between the two MRAPs. In this study on the rat adrenal gland, we demonstrate that while MRAP and MC2R are highly expressed in the zona fasciculata, MRAP2 is expressed throughout the adrenal cortex in low quantities. In the developing adrenal gland, both MRAP and MRAP2 are equally well expressed. The MC2R/MRAP2 complex requires much higher concentrations of ACTH to activate compared with the MC2R/MRAP complex. Interestingly, expression of MC2R and MRAP in the undifferentiated zone would support the notion that ACTH may play an important role in adrenal cell differentiation and maintenance.
-
Agonist activated adrenocorticotropin Receptor internalizes via a clathrin-mediated G protein Receptor kinase dependent mechanism.
Endocrine Research, 2009Co-Authors: Asma H. Baig, Peter J King, Francesca Swords, László Hunyady, Márta Szaszák, Adrian J L ClarkAbstract:The physiological effects of the pituitary hormone, adrenocorticotropic hormone (ACTH) on the adrenal are mediated by the Melanocortin 2 Receptor (MC2R), a G protein coupled Receptor (GPCR) that signals via adenylate cyclase to elevate intracellular cyclic AMP (cAMP) levels. The function and expression of the Receptor is likely to be a major determinant of the response to ACTH. Following repeated stimulation, the cAMP signal is diminished or desensitized. Prolonged desensitization may involve internalization of the Receptor. Internalization may occur by at least two mechanisms—Receptor mediated endocytosis via clathrin-coated pits and by caveolae mediated internalization. The mode of internalization for the endogenous MC2R in Y1 cells was determined using radiolabelled ACTH. Treatment of Y1 cells with hypertonic sucrose or with concanavalin A, which inhibit clathrin-mediated endocytosis, blocked internalization. Filipin and nystatin, which inhibit caveolae formation, did not influence internalization. A d...
-
the majority of adrenocorticotropin Receptor Melanocortin 2 Receptor mutations found in familial glucocorticoid deficiency type 1 lead to defective trafficking of the Receptor to the cell surface
The Journal of Clinical Endocrinology and Metabolism, 2008Co-Authors: Tengteng Chung, Tom R Webb, Li F Chan, Sadani N Cooray, J P Chapple, Peter J King, Louise A Metherell, Adrian J L ClarkAbstract:Context: There are at least 24 missense, nonconservative mutations found in the ACTH Receptor [Melanocortin 2 Receptor (MC2R)] that have been associated with the autosomal recessive disease familial glucocorticoid deficiency (FGD) type 1. The characterization of these mutations has been hindered by difficulties in establishing a functional heterologous cell transfection system for MC2R. Recently, the Melanocortin 2 Receptor accessory protein (MRAP) was identified as essential for the trafficking of MC2R to the cell surface; therefore, a functional characterization of MC2R mutations is now possible. Objective: Our objective was to elucidate the molecular mechanisms responsible for defective MC2R function in FGD. Methods: Stable cell lines expressing human MRAPα were established and transiently transfected with wild-type or mutant MC2R. Functional characterization of mutant MC2R was performed using a cell surface expression assay, a cAMP reporter assay, confocal microscopy, and coimmunoprecipitation of MRAP...
-
interaction of the Melanocortin 2 Receptor with nucleoporin 50 evidence for a novel pathway between a g protein coupled Receptor and the nucleus
The FASEB Journal, 2007Co-Authors: Marina Doufexis, Helen L Storr, Peter J King, Adrian J L ClarkAbstract:The adrenocorticotropin (ACTH) Receptor (Melanocortin 2 Receptor, or MC2R) is the smallest G-protein-coupled Receptor that, when activated by the peptide hormone ACTH, stimulates cAMP production an...
Mathilakath M Vijayan - One of the best experts on this subject based on the ideXlab platform.
-
cadmium disrupts Melanocortin 2 Receptor signaling in rainbow trout
Aquatic Toxicology, 2019Co-Authors: Navdeep Sandhu, Liang Liang, Robert M Dores, James C Mcgeer, Mathilakath M VijayanAbstract:Abstract Cadmium is an endocrine disruptor and inhibits corticosteroid production, but the mechanisms are far from clear. We tested the hypothesis that sublethal exposure to environmentally realistic levels of cadmium impairs cortisol production by disrupting the Melanocortin 2 Receptor (MC2R) signaling in rainbow trout (Oncorhynchus mykiss). Fish were exposed to sublethal concentrations of cadmium (0.75 or 2.0 μg/L) in a flow-through system for 7 d and subjected to an acute secondary stressor to evoke a cortisol response. Cadmium exposure for 7 d did not affect plasma cortisol concentrations, but head kidney mc2r mRNA levels were higher than in control fish. The cortisol stress performance to a secondary-stressor was attenuated in the cadmium groups, and this corresponded with transient reduction in transcript abundance of mc2r and the gene encoding its accessory protein MRAP1 but not MRAP2 in the head kidney. Furthermore, in vivo cadmium exposure attenuated the adrenocorticotropic hormone (ACTH)-, but not 8-br-cAMP-stimulated cortisol production in head kidney slices ex vivo. This corresponded with reduced transcript abundance of mc2r and mrap1, but not mrap2 in these tissue slices. Also, reporter assays with CHO cells transiently transfected with rainbow trout mc2r and zebrafish mrap1 revealed a dose-independent inhibition in ACTH-stimulated luciferase activity by cadmium. Collectively, waterborne exposure to environmentally realistic concentration of cadmium compromises the stressor-induced cortisol response, and a mode of action involves the disruption of MC2R signaling in rainbow trout.
-
identifying the activation motif in the n terminal of rainbow trout and zebrafish Melanocortin 2 Receptor accessory protein 1 mrap1 orthologs
General and Comparative Endocrinology, 2016Co-Authors: Robert M Dores, Liang Liang, Navdeep Sandhu, Rebecca E Hollmann, Mathilakath M VijayanAbstract:The activation of mammalian Melanocortin-2 Receptor (MC2R) orthologs is dependent on a four-amino acid activation motif (LDYL/I) located in the N-terminal of mammalian MRAP1 (Melanocortin-2 Receptor accessory protein). Previous alanine substitution analysis had shown that the Y residue in this motif appears to be the most important for mediating the activation of mammalian MC2R orthologs. Similar, but not identical amino acid motifs were detected in rainbow trout MRAP1 (YDYL) and zebrafish MRAP1 (YDYV). To determine the importance of these residues in the putative activation motifs, rainbow trout and zebrafish MRAP1 orthologs were individually co-expressed in CHO cells with rainbow trout MC2R, and the activation of this Receptor with either the wild-type MRAP1 ortholog or alanine-substituted analogs of the two teleost MRAP1s was analyzed. Alanine substitutions at all four amino acid positions in rainbow trout MRAP1 blocked activation of the rainbow trout MC2R. Single alanine substitutions of the D and Y residues in rainbow trout and zebrafish MRAP1 indicate that these two residues play a significant role in the activation of rainbow trout MC2R. These observations indicate that there are subtle differences in the way that teleost and mammalian MRAPs are involved in the activation of their corresponding MC2R orthologs.
-
structure function studies on the activation of the rainbow trout Melanocortin 2 Receptor
General and Comparative Endocrinology, 2015Co-Authors: Liang Liang, Joseph K Angleson, Kristin Schmid, Navdeep Sandhu, Mathilakath M Vijayan, Robert M DoresAbstract:Abstract Functional expression of the rainbow trout (rt) Melanocortin-2 Receptor (MC2R) in CHO cells requires co-expression with a teleost Melanocortin-2 Receptor accessory protein (MRAP) such as zebrafish (zf) MRAP. Transiently transfected rtMC2R/zfMRAP1 CHO cells were used to evaluate the efficacy of alanine substituted analogs of hACTH(1-24) in three motifs in the ligand: H6F7R8W9, G10K11P12V13G14, and K15K16R17R18P19. Alanine substitution at all positions in each motif either completely blocked activation of the Receptor (H6F7R8W9 and K15K16R17R18P19) or resulted in just over 400 fold increase in EC50 value (G10K11P12V13G14). Single alanine substitutions in the H6F7R8W9 motif indicated that substitution at either W9 or R8 resulted in a much larger increase in EC50 values as compared to substitutions at either F7 or W9. Alanine substitution at either K15K16 or R17R18P19 in the K15K16R17R18P19 motif resulted in a statistically equivalent increase in EC50 value of at least 600 fold. Finally, alanine substitutions in the G10K11P12V13G14 motif resulted in increases in EC50 values presumably as a result of altering the secondary structure of the ligand. However, truncated analogs of hACTH(1-24) in which either G10G14 (ACTH(1-22), or K11P12V13 (ACTH(1-21) were removed had no stimulatory activity. Finally, some of the hACTH(1-24) analogs were tested using rainbow trout head kidney pieces in vitro to confirm whether the response to analogs seen with the transient transfected rtMC2R CHO cells was similar to that of trout interrenal cells. The results of these alanine substitution analog studies are used to construct a multistep hypothetical model for the activation of teleost and tetrapod MC2Rs to account for the unique ligand selectivity of this Receptor.
-
Structure/function studies on the activation of the rainbow trout Melanocortin-2 Receptor.
General and Comparative Endocrinology, 2014Co-Authors: Liang Liang, Joseph K Angleson, Kristin Schmid, Navdeep Sandhu, Mathilakath M Vijayan, Robert M DoresAbstract:Abstract Functional expression of the rainbow trout (rt) Melanocortin-2 Receptor (MC2R) in CHO cells requires co-expression with a teleost Melanocortin-2 Receptor accessory protein (MRAP) such as zebrafish (zf) MRAP. Transiently transfected rtMC2R/zfMRAP1 CHO cells were used to evaluate the efficacy of alanine substituted analogs of hACTH(1-24) in three motifs in the ligand: H6F7R8W9, G10K11P12V13G14, and K15K16R17R18P19. Alanine substitution at all positions in each motif either completely blocked activation of the Receptor (H6F7R8W9 and K15K16R17R18P19) or resulted in just over 400 fold increase in EC50 value (G10K11P12V13G14). Single alanine substitutions in the H6F7R8W9 motif indicated that substitution at either W9 or R8 resulted in a much larger increase in EC50 values as compared to substitutions at either F7 or W9. Alanine substitution at either K15K16 or R17R18P19 in the K15K16R17R18P19 motif resulted in a statistically equivalent increase in EC50 value of at least 600 fold. Finally, alanine substitutions in the G10K11P12V13G14 motif resulted in increases in EC50 values presumably as a result of altering the secondary structure of the ligand. However, truncated analogs of hACTH(1-24) in which either G10G14 (ACTH(1-22), or K11P12V13 (ACTH(1-21) were removed had no stimulatory activity. Finally, some of the hACTH(1-24) analogs were tested using rainbow trout head kidney pieces in vitro to confirm whether the response to analogs seen with the transient transfected rtMC2R CHO cells was similar to that of trout interrenal cells. The results of these alanine substitution analog studies are used to construct a multistep hypothetical model for the activation of teleost and tetrapod MC2Rs to account for the unique ligand selectivity of this Receptor.
-
molecular characterization tissue specific expression and regulation of Melanocortin 2 Receptor in rainbow trout
Endocrinology, 2008Co-Authors: Neelakanteswar Aluru, Mathilakath M VijayanAbstract:ACTH, the primary secretagogue for corticosteroid biosynthesis, binds to Melanocortin 2 Receptor (MC2R) and activates the signaling cascade leading to steroid biosynthesis in the adrenal cortex. Whereas MC2R regulation has been studied using mammalian models, little is known about the molecular mechanisms involved in ACTH signaling in nonmammalian vertebrates. A full-length cDNA encoding MC2R was sequenced from rainbow trout (Oncorhynchus mykiss) interrenal tissue (analogous to the adrenal cortex in mammals) and showed about 60 and about 44% amino acid sequence similarity to teleosts and humans, respectively. Phylogenetic analysis confirmed that MC2R from all species clustered together and was distant from other MCRs. Quantitative real-time PCR revealed a marked tissue-specific difference in MC2R mRNA abundance, with the highest levels observed in the interrenal tissue, ovary, and testis. Acute ACTH, but not α-MSH or [Nle4, d-Phe7]-MSH, stimulation resulted in a time- and dose-related elevation in MC2R mR...