The five melanocortin receptors are members of the seven-transmembrane domain, G-protein coupled receptor (GPCR) superfamily. Melanocortins, the ligands of the receptors are a group of structurally similar peptides consisting of α-, β- and γ-melanocyte stimulating hormone (α, β, γ-MSH) and the adrenocorticotropic hormone (ACTH) all of which are derived from the post-translational processing of a common precursor peptide, proopiomelanocortin (POMC).
One of the most significant characteristics of the melanocortin signaling system is the presence of two endogenous antagonists, agouti (or agouti signaling protein, ASP) and agouti-related protein (AGRP). All five melanocortin receptors bind their agonists and their endogenous antagonists with different affinities. In this short article we report the use of Alomone Labs products in melanocortin-related research.
Melanocortin Receptors in the Kidney
Membranous Nephropathy (MN) is one of the most common kidney diseases in adults. Reports show that ACTH treatment reduces proteinurea, a symptom of the disease1. The mechanisms through which ACTH does so were investigated. MC1R was detected in podocytes of healthy and MN subjects in immunohistochemical staining and western blot analysis using Anti-MC1 Receptor Antibody (#AMR-020). In addition, MC1R was also detected in glomerular endothelial cells, mesangial cells, and tubular epithelial cells. In order to obtain more specific information about MC1R’s localization, immunohistochemistry studies showed that MC1R co-localizes with synaptopodin, but not with UAE1 (ulex), indicating that MC1R is localized to podocytes and not to endothelial cells (Figure 1). In summary, the data suggest that ACTH probably induces its effects through MC1R2.
Si et al.5 showed that MC1R, along with MC5R are expressed in the kidney using Alomone Labs Anti-MC1 Receptor Antibody (#AMR-020) and Anti-MC5 Receptor Antibody (#AMR-025). In this particular study, the reno-protective effects of ACTH in acute kidney injury (AKI) were explored. Immunostaining indicated that MC1R is expressed intensely and predominantly in renal tubules and very weakly in glomeruli, whereas MC5R is weakly and sporadically expressed in the interstitial cells. The authors go on to show that Tumor Necrosis Factor (TNF) injury enhanced the expression of MC1R in the tubules but had minor effect on MC5R. Cells treated with MC1R-specific siRNA showed a significant decrease in the MC1R signal in western blot analysis and immunocytochemistry using Anti-Melanocortin Receptor 1 antibody, thereby indicating the high specificity of MC1R antibody.
The findings show that ACTH protects kidney function and promotes general survival in animal models of AKI in part by activating MC1R5.
Figure 1. Expression of MC1R in Human Kidney.Immunohistochemical staining of human frozen kidney sections using Anti-MC1 Receptor Antibody (#AMR-020). MC1R (green) colocalizes with synaptopodin (red).Adapted from reference 2 with permission of the American Society of Nephrology.
Melanocortin Receptors in the Nervous System
MC4R is involved in processes of learning and memory function through its interaction with the neuropeptide α-melanocyte-stimulating hormone (α-MSH). Shen et al.4 showed that activation of MC4R enhances synaptic plasticity through the regulation of dendritic spine morphology and abundance of AMPA receptors, and that MC4R stimulates the AMPA receptor subunit GluA1 trafficking through phosphorylation of GluA1. The expression of MC4R in mouse hippocampal neurons was detected in immunocytochemical staining using Anti-MC4 Receptor (extracellular) Antibody (#AMR-024) (Figure 2A). In addition, the essential role MCR4 regarding synaptic functionality was demonstrated by treating hippocampal neurons with MCR4 targeted shRNA. The efficiency of shRNA treatment was observed in western blot analysis using Anti-MC4 Receptor (extracellular) Antibody (#AMR-024) in HEK293 cells transfected with MC4R expression construct (Figure 2B). Knocking down MCR4 significantly reduced dentritic spine density and the percentage of mature spines4.
Figure 2. Expression of MC4R in Mouse Hippocampal Neurons.A. Immunocytochemical staining of mouse hippocampal neurons. Extracellular staining of cells using Anti-MC4 Receptor (extracellular) Antibody (#AMR-024) (green). B. Western blot analysis of MC4R-transfected HEK 293 cells treated with and with shRNA targeting MC4R. shRNA treatment decreased the receptor level by 83%. MC4R protein levels were detected with Anti-Melanodortin Receptor 4 (extracellular) antibody.Adapted from reference 4 with permission of the Society for Neuroscience.
In a study examining hypothalamic neural circuits, the signaling of Bone Morphogenetic Protein (BMP) was found to regulate the development of feeding behavior3. As part of the study, a knockout mouse strain for the BMP receptor, BMPR1A was generated. Among BMPR1A-/- characteristics, mice exhibit significant weight loss and a decrease of agouti-related protein positive neurons. Hence, MC4R expression was verified by immunohistochemistry staining in mouse brain using Anti-MC4 Receptor (extracellular) Antibody. MC4R levels remained intact in knockout mice, and showed no significant change compared to the control3.
1. Berg, A.L. et al. (1999) Kidney Int. 56, 1534.
2. Lindskog, A. et al. (2010) J. Am. Soc. Nephrol. 21, 1290.
3. Peng, C.Y. et al. (2012) J. Neurosci. 32, 17211.
4. Shen, Y. et al. (2013) J. Neurosci. 33, 464.
5. Si, J. et al. (2013) Kidney Int. 83, 635.