Cellular Signalling 78 (2021) 109862 Contents lists available at ScienceDirect Cellular Signalling journal homepage: www.elsevier.com/locate/cellsig Biased agonism at chemokine receptors Dylan Scott Eiger a, Noelia Boldizsar b, Christopher Cole Honeycutt b, Julia Gardner b, Sudarshan Rajagopal a,c,* a Department of Biochemistry, Duke University, Durham, NC 27710, USA b Trinity College of Arts and Sciences, Duke University, Durham, NC 27710, USA c Department of Medicine, Duke University, Durham, NC 27710, USA ARTICLE INFO ABSTRACT Keywords: In the human chemokine system, interactions between the approximately 50 known endogenous chemokine Biased agonism ligands and 20 known chemokine receptors (CKRs) regulate a wide range of cellular functions and biological G protein-coupled receptors processes including immune cell activation and homeostasis, development, angiogenesis, and neuromodulation. Chemokine System CKRs are a family of G protein-coupled receptors (GPCR), which represent the most common and versatile class of receptors in the human genome and the targets of approximately one third of all Food and Drug Administration-approved drugs. Chemokines and CKRs bind with significant promiscuity, as most CKRs can be activated by multiple chemokines and most chemokines can activate multiple CKRs. While these ligand-receptor interactions were previously regarded as redundant, it is now appreciated that many chemokine:CKR interactions display biased agonism, the phenomenon in which different ligands binding to the same receptor signal through different pathways with different efficacies,leading to distinct biological effects. Notably, these biased responses can be modulated through changes in ligand, receptor, and or the specificcellular context (system). In this review, we explore the biochemical mechanisms, functional consequences, and therapeutic potential of biased agonism in the chemokine system. An enhanced understanding of biased agonism in the chemokine system may prove transformative in the understanding of the mechanisms and consequences of biased signaling across all GPCR subtypes and aid in the development of biased pharmaceuticals with increased therapeutic efficacyand safer side effect profiles. 1. Chemokine System normal and pathologic conditions. The role of CKR activation by che­ mokines was first recognized in the immune response, specifically as Chemokine receptors (CKRs) are a subfamily of G protein-coupled chemoattractants to direct leukocyte migration, a process known as receptors (GPCRs) that bind a group of small (8-12 kDa) and highly chemotaxis [5,6]. While the functions and roles of chemokines in leu­ conserved chemotactic cytokines known as chemokines [1]. The human kocytes are well known, it is now appreciated that chemokines and CKRs chemokine system is composed of approximately 20 known CKRs and 50 are also produced in a variety of non-leukocyte cell types, including known chemokines (Fig. 1). The chemokines are classified into four epithelial cells, fibroblasts,endothelial cells, and neurons [7], and play a subtypes (C, CC, CXC, CX3C) based on the number, positioning, and key role in a wide range of other cellular functions and biological pro­ spacing of conserved N-terminal cysteine residues [2]. Similarly, CKRs cesses including development, angiogenesis, neuromodulation, and are organized and classified according to the ligands they bind [3]. immune cell homeostasis [8–11]. For example, the expression of Chemokines are also categorized as homeostatic chemokines, which are neuronal chemokine ligands and receptors has recently been shown to constitutively expressed in a variety of specifictissues and cell types, and be involved in synaptic transmission and neuronal survival [12], as well inflammatorychemokines, which are induced during immune responses as in guidance of central nervous system (CNS) cellular interactions via primarily to recruit leukocytes to sites of inflammation[ 4]. Homeostatic neuron-astrocyte, neuron-microglia, and neuron-neuron interactions and inflammatory classifications of chemokines are not mutually [13]. exclusive, as some CKRs and chemokine ligands are involved in both Due to the chemokine system’s involvement in a wide variety of * Corresponding author at: Box 102147, Duke University Medical Center, Durham, NC 27710, USA. E-mail addresses: [email protected] (D.S. Eiger), [email protected] (N. Boldizsar), [email protected] (C.C. Honeycutt), julia. [email protected] (J. Gardner), [email protected] (S. Rajagopal). https://doi.org/10.1016/j.cellsig.2020.109862 Received 9 September 2020; Received in revised form 7 November 2020; Accepted 24 November 2020 Available online 27 November 2020 0898-6568/© 2020 Elsevier Inc. All rights reserved. D.S. Eiger et al. Cellular Signalling 78 (2021) 109862 biological processes, it is unsurprising that chemokines and CKRs are biased signaling for drug development in the chemokine system. implicated in various disease states including, but not limited to, auto­ immune disorders, infectious diseases, hypersensitivity reactions, 2. G Protein-Coupled Receptor Signaling and Biased Agonism atherosclerosis, and cancer [14–18]. The role of the chemokine system in chronic inflammatory diseases is particularly important and chemo­ CKRs are a subfamily of the rhodopsin class of GPCRs, the most kines play a central role in asthma, chronic obstructive pulmonary dis­ common and versatile superfamily of receptors in the human genome ease, inflammatory bowel disease, arthritis, multiple sclerosis, and [25] and the target of ~34% of all Food and Drug Administration (FDA) psoriasis [19]. Additionally, certain disorders are directly associated approved pharmaceutical drugs [26]. Canonical GPCR signaling starts with mutations in the genes that encode CKRs, such as the Warts, Hy­ with agonist binding, upon which a GPCR undergoes conformational pogammaglobulinemia, Immunodeficiency,and Myelokathexis (WHIM) changes that induce the recruitment of heterotrimeric G proteins con­ Syndrome which is driven by an autosomal dominant truncation mutant sisting of Gα, Gβ, and Gγ subunits. The guanosine diphosphate (GDP)- in the receptor CXCR4 [20]. bound Gα subunit undergoes nucleotide exchange for guanosine While the chemokine system is known to play a significant role in triphosphate (GTP), leading to Gα activation and dissociation of the many disease states, there are relatively few drugs that target it directly. heterotrimeric complex into its Gα and Gβγ constituents [27]. The Gα Chemokines and CKRs bind with significantpromiscuity, wherein most subunits are classified into four families based on sequence similarity: CKRs can be activated by multiple chemokine ligands and most che­ Gαs, Gαi/o, Gαq/11, and Gα12/13 [28]. The activated Gα proteins typically mokines can activate multiple CKRs [21]. This promiscuity was thought regulate the production and subsequent signaling of secondary mes­ to lead to “redundancy” between chemokines and their receptors, sengers, such as adenosine 3’,5’-cyclic monophosphate (cAMP), intra­ serving as a mechanism for a robust physiologic response [22]. As cellular calcium, and inositol triphosphate [29]. Most chemokine adequate chemokine levels are imperative for immune cell function, receptors signal through Gαi/o, which inhibits adenylyl cyclase and re­ redundant chemokine signaling would provide sufficient signals to duces intracellular concentrations of cAMP [30]. There are various direct leukocyte chemotaxis and function that is relatively insensitive to isoforms of the Gβ and Gγ subunits, and at chemokine receptors the Gβγ variations in the concentration of any individual chemokine [23]. dimer has been shown to activate phosphoinositide-specific phospholi­ However, we now appreciate that many of these ligands can have pase Cβ (PLC) and phosphoinositide 3-kinase (PI3K). PLC then produces distinct signaling profiles at the same receptor and many receptors can diacylglycerol (DAG), leading to the activation of protein kinase C have distinct signaling profiles when stimulated by the same ligand, a (PKC), and inositol-triphosphate (IP3), which triggers calcium mobili­ phenomenon referred to as “biased agonism” [22,24]. Biased signaling zation [31]. The signaling messengers of the Gβγ dimer in chemokine through differences in ligands, receptors and the cellular context (sys­ receptors have been demonstrated to play a role in the promotion of tem) can have important effects on chemokine signaling and implica­ leukocyte migration, among other functions [32]. tions for drug development. Here, we review the current literature on Following G protein activation, G protein-coupled receptor kinases biased signaling within the chemokine system to highlight the complex (GRKs) are recruited to the receptor and phosphorylate the receptor C- and multidimensional nature of biased agonism and the biochemical terminus and intracellular loops. This phosphorylation promotes the mechanisms that underlie it, the importance of biased agonism within interaction of the receptor with the β-arrestins, which were first the chemokine system and its physiologic effects, and the implications of described for their function in the desensitization of G protein-mediated Fig. 1. The complexity of the human chemokine system. Chemokine receptors fall into fivecategories: CCRs, CXCRs, ACKRs, XCRs and CX3CRs. Chemokine ligands fall into four categories: CCLs, CXCLs, XCLs, CX3CLs. Lines connecting chemokine receptors to chemokines are colored