Molecular and Functional Analysis of Human β-Defensin 3 Action at Melanocortin Receptors

Abstract

Summary The β-defensins are a class of small, cationic proteins first recognized as antimicrobial components of the innate and adaptive immune system. More recently, one of the major β-defensins produced in skin, β-defensin 3, has been discovered to function as a melanocortin receptor ligand in vivo and in vitro, but its biophysical and pharmacological basis of action has been enigmatic. Here we report functional and biochemical studies focused on human β-defensin 3 (HBD3) and melanocortin receptors 1 and 4. Genetic and pharmacologic studies indicate that HBD3 acts as a neutral melanocortin receptor antagonist, capable of blocking the action of either stimulatory agonists, such as α-melanocyte stimulating hormone, or inhibitory inverse agonists such as Agouti signaling protein (Asip) and Agouti-related protein (Agrp). A comprehensive structure-function analysis demonstrates that two patches of positively charged residues, located on opposite poles of HBD3 and spatially organized by the compact β-defensin fold, are primarily responsible for high affinity binding to melanocortin receptors. These findings identify a distinct mode of melanocortin receptor-ligand interactions based primarily on electrostatic complementarity, with implications for designing ligands that target melanocortin and potentially other seven transmembrane receptors.

Membrane Protein 4F2/CD98 Is a Cell Surface Receptor Involved in the Internalization and Trafficking of Human β-Defensin 3 in Epithelial Cells

“…In the CD98 region that interacts with the highly charged hBD3, the loops contain a large number of negatively charged residues (see Figure 7C). We propose that this complementary charge interaction is the structural determinant for CD98-hBD3 recognition, as observed for the electrostatic interaction of hBD3 with the melanocortin and chemokine receptors (Nix et al, 2013;Esseghir et al, 2006).…”

“…In particular, we addressed the question of whether hBD3 internalization is mediated by specific receptors on epithelial cells, as described for the receptors expressed on immune cells, the binding of which also triggers chemotaxis (Soruri et al, 2007;Rö hrl et al, 2010). In this context, it is noteworthy that hBD3 has been implicated in the modulation of melanocortin receptors (Beaumont et al, 2012;Nix et al, 2013). Our analysis identifies 4F2/CD98 as an hBD3-specific membrane receptor and shows that it is internalized by means of recycling endosomes.…”

60 YEARS OF POMC: Regulation of feeding and energy homeostasis by α-MSH

“…Cachexia is invariably linked to immunosuppression. As strongly charged molecules, defensins might interact with MCR solely through electrostatic interactions explaining their neutral antagonist effect (Nix et al 2013, 2015). There is little doubt that these interactions play a role in determining melanocortin response on MC1R, as defensins seem to block agouti in live animals (Candille et al 2007, Kerns et al 2007).…”

Morphological Evolution Repeatedly Caused by Mutations in Signaling Ligand Genes

“…In addition to Agouti, the β-defensin 3/CBD103 peptide is secreted by skin epithelia, strongly binds to MC1R, and was shown to be responsible for melanism in dogs (Candille et al 2007). In certain melanic dog breeds, one amino acid deletion in β-defensin 3/CBD103 results in dominant melanism, possibly by blocking the inhibitory activity of Agouti or by losing its blocking of α-MSH stimulatory binding (Nix et al 2013). Of note, the CBD103 ΔG23 melanic allele is revealing a complex history that blurs the boundary between wild and domesticated.…”