9 - Role of PDZ Proteins in Regulating Trafficking, Signaling, and Function of GPCRs: Means, Motif, and Opportunity

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Abstract

PDZ proteins, named for the common structural domain shared by the postsynaptic density protein (PSD95), Drosophila disc large tumor suppressor (DlgA), and zonula occludens-1 protein (ZO-1), constitute a family of 200–300 recognized members. These cytoplasmic adapter proteins are capable of assembling a variety of membrane-associated proteins and signaling molecules in short-lived functional units. Here, we review PDZ proteins that participate in the regulation of signaling, trafficking, and function of G protein-coupled receptors. Salient structural features of PDZ proteins that allow them to recognize targeted GPCRs are considered. Scaffolding proteins harboring PDZ domains may contain single or multiple PDZ modules and may also include other protein–protein interaction modules. PDZ proteins may impact receptor signaling by diverse mechanisms that include retaining the receptor at the cell membrane, thereby increasing the duration of ligand binding, as well as importantly influencing GPCR internalization, trafficking, recycling, and intracellular sorting. PDZ proteins are also capable of modifying the assembled complex of accessory proteins such as β-arrestins that themselves regulate GPCR signaling. Additionally, PDZ proteins may modulate GPCR signaling by altering the G protein to which the receptor binds, or affect other regulatory proteins that impact GTPase activity, protein kinase A, phospholipase C, or modify downstream signaling events. Small molecules targeting the PDZ protein–GPCR interaction are being developed and may become important and selective drug candidates.

Introduction

G protein-coupled receptors (GPCRs) form the largest family of signaling receptors that are expressed in vertebrate cells. They are responsible for transducing a strikingly vast array of extracellular signals to biological actions. GPCRs represent 2% of the human genome and are important drug targets. Effectively, these receptors are guanine nucleotide exchange factors, which when occupied by their cognate ligand, exchange guanosine diphosphate (GDP) for guanosine triphosphate (GTP) on the alpha subunit of the associated heterotrimeric nucleotide-binding protein. The activated alpha subunit then dissociates from the beta–gamma subunit. Signal transduction is mostly mediated by the alpha subunit but sometimes by the beta–gamma subunit pair. The two principal signaling pathways involved are by Gαs or inhibition by Gi of the adenylyl cyclase–cAMP–protein kinase A (PKA)/EPAC signaling pathway, and stimulation by Gαq of the phospholipase C (PLC)—Ca2+ + phosphatidylinositol pathway. GPCR desensitization provides a mechanism to protect cells against excessive stimulation, while GPCR resensitization guards cells against prolonged desensitization and hormone insensitivity. Desensitization and receptor internalization are the two primary mechanisms controlling GPCR signaling.

Although most receptors activate a single pathway, some receptors employ multiple signaling pathways. The type 1 parathyroid hormone receptor (PTH1R), for instance, in vascular smooth muscle cells, parathyroid hormone (PTH), stimulates adenylyl cyclase but not PLC (Maeda et al., 1996, Wu et al., 1993), whereas in keratinocytes (Orloff et al., 1995, Whitfield et al., 1992), cardiac myocytes (Rampe et al., 1991, Schlüter et al., 1995), and lymphocytes (Atkinson et al., 1987, Klinger et al., 1990, Whitfield et al., 1971), the PTH1R activates PLC but not adenylyl cyclase. In osteoblasts and kidney tubule cells, PTH activates both adenylyl cyclase and PLC (Abou-Samra et al., 1992, Friedman et al., 1996, Hruska et al., 1987). The origin of the cell-specific signaling remained obscure until the discovery that a PDZ adapter protein, present in some but not in all cells expressing the PTH1R switches signaling between adenylyl cyclase and PLC (Mahon et al., 2002). Increasing evidence now supports the view that cytoplasmic adapter proteins affect the signaling and trafficking of many GPCRs, and thereby their biological behavior. In this review, we describe emerging findings regarding the means by which modular PDZ proteins confer ligand- and cell-specific signaling and trafficking on select GPCRs, the corresponding recognition motifs engaged by the cognate proteins, and the physiological opportunities regulated by these interactions.

Section snippets

PDZ Proteins

PDZ proteins are soluble cytoplasmic adapter proteins that function as transient scaffolding structures to assemble multiprotein signaling complexes by virtue of highly conserved modules. The general arrangement for PDZ domains is based on the structure of PSD95, DLG, and ZO1, for which they are named. The human genome includes some 200–300 PDZ proteins. PDZ modules consist of an 80–90 amino acid sequence forming a three-dimensional globular structure that is composed of six β-sheets (βA–βF)

Family A GPCRs

Depending on how stringently one defines the consensus motif for PDZ-mediated protein interaction, a handful to a potentially large number of mammalian family A GPCRs have the ability to engage PDZ domain-containing proteins. Table III lists those family A GPCRs for which such interactions have been established most convincingly, and specifically linked to function.

G Proteins

Although we focus our discussion primarily on GPCRs interacting with PDZ proteins, it should be noted that several G proteins themselves have canonical or internal PDZ-recognition sequences that bind PDZ proteins. Both NHERF1 and NHERF2, for example, bind Gαq (Rochdi et al., 2002, Wang et al., 2010), and NHERF2, but not NHERF1, interacts with Gαi. Neither NHERF1 nor NHERF2 associates with Gαs. The ability of PDZ proteins to engage G proteins underscores their ability to act as molecular routers

Conclusion

Our understanding of multiprotein interactions and how they impart many of the characteristic features of GPCRs is a subject of intense investigation and consequently a rapidly changing arena. It is now clear that many of the heretofore irreconcilable reported findings between GPCR signaling, trafficking, and function in different cells or in response to distinct ligands now can be attributed to the participation of PDZ proteins and their ability to confer ligand- and cell-specific actions on

Acknowledgments

Original work conducted in the authors’ laboratories was supported by grants DK054171, DK069998 (PAF), and DA10711, DA12864 (MVZ) from the National Institutes of Health.

Conflict of Interest: The authors have no conflicts of interest to declare.

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