Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

Activators of G-Protein Signaling (AGS)

Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_455

Synonyms

Historical Background

Activators of G-protein signaling (AGS) proteins define a group of proteins identified in a yeast-based functional screen of mammalian cDNA libraries as receptor-independent activators of the G-protein signaling cascade. Activators of G-protein signaling are one subgroup of accessory proteins for G-protein signaling systems. Accessory proteins are generally defined as proteins other than the core triad of receptor (R), G-protein, and effector (E) that regulate the efficiency and/or specificity of signal transfer from G-protein coupled receptors (GPCRs) to G-proteins, segregate a signaling complex to microdomains of the cell, regulate the basal activity of the system, and/or provide alternative modes of signal input to G-protein signaling systems that operate independent of a typical GPCR. Such accessory proteins may influence G-protein signaling systems operating at the cell cortex or at intracellular locations (Fig. 1).
Activators of G-Protein Signaling (AGS), Fig. 1

Schematic diagram indicating points of influence of accessory proteins on G-protein signaling systems. AGS and related proteins may serve as alternative binding partners for G-protein subunits and also regulate signal transfer at multiple points within the broader G-protein signaling system (The figure is a modification of the concept figure presented by Sato et al. 2006)

The concept of accessory proteins resulted from a confluence of several independent lines of investigation (see Blumer et al. 2007, 2011; Gonczy 2008; Knoblich 2010; Sato et al. 2006; Willard et al. 2004). These lines of investigation included cell-specific differences in signal transfer from R to G, the partial purification of a putative non-receptor G-protein activator from extracts of NG108-15 cells, identification of G-protein subunits in intracellular organelles, identification of unexpected binding partners for G-protein subunits, and the identification of non-receptor proteins that could influence the activation state of G-proteins eventually leading to the development of a functional yeast-based screen for mammalian entities that activated G-protein signaling in the absence of a receptor (Sato et al. 2006; Cismowski and Lanier 2005; Cismowski et al. 1999; Takesono et al. 1999). Interspersed with these biochemical approaches was the realization that there were changes in signal processing through G-protein signaling systems that occurred independent of any obvious changes in receptor number or G-protein expression levels, suggesting additional undefined regulatory mechanisms. Another line of investigation evolved out of the study of asymmetric cell division in Drosophila melanogaster neuroblasts and sensory organ precursor cells in parallel with the Caenorhabditis elegans embryo. Thus, a confluence of biochemical, cell biology, and model organism data indicated unexpected modes of regulation for heterotrimeric G-proteins associated with previously unknown functional roles for this signaling system.

The discovery of AGS proteins involved the development of a functional readout that would allow rapid screening of mammalian cDNAs for their ability to activate G-protein signaling in the absence of a receptor. The initial reports of the discovery of AGS proteins using the yeast-based functional screen (Cismowski et al. 1999) included three AGS proteins with each defining a distinct group of AGS proteins (Fig. 2) (Cismowski et al. 1999; Takesono et al. 1999). AGS1 behaved as a guanine nucleotide exchange factor for Gαi/Gαo. AGS3 is the prototype for Group II AGS proteins. AGS2 defined Group III AGS proteins, which generally interact with Gβγ. In the yeast-based functional platform, Group I and II AGS proteins were functionally active in yeast strains expressing Gαi2, Gαi3, but not Gα16 or Gαs whereas the activity of Group III AGS proteins was independent of the type of Gα expressed. Subsequent screening of different mammalian cDNA libraries generated from different tissues exposed to different challenge paradigms or exhibiting specific pathologies yielded 13 distinct mammalian cDNAs that exhibited bioactivity in this functional platform (Fig. 2). AGS11-13, which are actually transcription factors, were identified in the functional screen using yeast expressing Gα16, but there is not currently enough information to determine how they should be subclassified within the larger group of AGS proteins (Sato et al. 2011).
Activators of G-Protein Signaling (AGS), Fig. 2

Functional roles of different Activators of G-protein signaling. This figure includes proteins that were not isolated in the yeast-based functional screen as AGS proteins, but were identified in other protein interaction or functional screens and shown to exhibit biological activity consistent with their inclusion as Group 1 (RIC8A, CCDC88A, GET3) or Group II (GPSM4, RAP1GAP, RGS14) AGS proteins (Modified from Blumer et al. 2007)

Group I AGS Proteins

To date, AGS1 is the only cDNA isolated in the yeast expression cloning system exhibiting functional activity consistent with its classification as a guanine nucleotide exchange factor (GEF). Initially isolated as a dexamethasone-inducible cDNA (Kemppainen and Behrend 1998), AGS1 (RASD1) appears to inhibit cell growth (Vaidyanathan et al. 2004) and is downregulated in various cancers (Dalgin et al. 2007; de Souza Rocha Simonini et al. 2010; Furuta et al. 2006; Nojima et al. 2009). AGS1 is also reported to interact with Gβγ (Hiskens et al. 2005). Additional non-receptor GEFs such as  Ric-8A and  Ric-8B (Miller et al. 2000; Tall et al. 2003), GIV/Girdin (Le-Niculescu et al. 2005), and Arr4 (Lee and Dohlman 2008) have been identified by other functional and protein interaction screens, and these proteins would also fit the definition of Group I AGS proteins.

Group II AGS Proteins

Group II AGS proteins are characterized by the presence of one to four, evolutionarily conserved, G-protein regulatory (GPR) or GoLoco motifs (Takesono et al. 1999; Siderovski et al. 1999), which are 20–25-amino acid cassettes that interact with Gαi/o-GDP and Gαt-GDP. Although the core residues within GPR motifs are conserved, individual GPR motifs may differ in their relative affinities for different Gαi/o and  Gαt family members. The GPR motif defines a novel, totally unexpected mechanism for regulation of the activation-deactivation cycle of heterotrimeric G-proteins with potentially broad conceptual implications. In contrast to Group I and III AGS proteins, each member of the Group II AGS proteins has a shared structural motif (GPR/GoLoco) (Fig. 3).
Activators of G-Protein Signaling (AGS), Fig. 3

Schematic representation of Group II AGS proteins. Where indicated, the lines underneath the schematic representation of individual entities correspond to the coding region actually isolated in the initial yeast-based functional screen (The figure is reproduced from Blumer et al. 2011)

The genes encoding AGS3, AGS4, and AGS5 were named G-protein signaling modulator (GPSM) 1, 3, and 2, respectively, by the HUGO Gene Nomenclature Committee. There are at least three additional GPR proteins in mammals, Pcp2/L7 (GPSM4), RGS14, and Rap1Gap (Transcript Variant 1, isoform a, Rap1GapII), which to date have not been identified in any of the yeast-based functional screens described above, likely reflecting variations in cDNA representation in the libraries used for screening or other factors which affect their activity in the yeast screen. In silico screens based on conserved GPR consensus sequences also identified a putative GPR motif in the protein WAVE1/Scar, although it is not yet known if the motif actively engages Gα-GDP in the context of WAVE1 function in the cell (Song et al. 2006).

Proteins with GPR motifs can be further subgrouped based on the number of GPR motifs they contain. One subset (AGS3, AGS4, AGS5, and Pcp2/L7) has 2–4 GPR motifs, whereas a second subset (AGS6/RGS12, RGS14, Rap1Gap) contains a single GPR motif. Within the first subset, AGS3 and AGS5 share a similar domain architecture and are ∼60% identical at the amino acid level with an amino-terminal tetratricopeptide repeat (TPR) domain and four carboxyl-terminal GPR motifs. AGS4 and Pcp2/L7, as well as the AGS3 variant AGS3-short, have 2–3 GPR motifs and lack any other clearly defined protein interaction domains. Within the second subset (AGS6/RGS12, RGS14, and Rap1Gap), in addition to their single GPR motif, each of the three proteins contain a GTPase-activating (GAP) domain in addition to other protein interaction or regulatory motifs. Thus, AGS6/RGS12 and RGS14 have an interesting arrangement of a GPR domain, which can dock GαGDP independently of Gβγ, and a GAP domain which accelerates GTP hydrolysis on the transition state of activated Gα, thus acting to terminate GαGTP-based signaling. Although conceptually this is a very interesting combination of domains with completely different, but perhaps complementary, effects on the nucleotide-bound state of Gα, the functional roles of these intramolecular domains within AGS6/RGS12 and RGS14 on the integration of G-protein signaling are not yet fully understood. The GAP domain in Rap1Gap does not regulate heterotrimeric Gα subunits but rather targets the ras-related small GTPase Rap1.

GPR motifs serve as docking sites for Gαi-GDP free of Gβγ and this GPR-Gα signaling module is regulated by both cell surface G-protein coupled receptors and non-receptor GEFs ( RIC-8, GIV/Girdin), which promote exchange of GDP for GTP and dissociation or rearrangement of Gα and the GPR motif in a manner analogous to that observed for G-protein-coupled receptor-mediated regulation of Gαβγ (Garcia-Marcos et al. 2011; Tall and Gilman 2005; Thomas et al. 2008; Vellano et al. 2011). The GPR-Gα signaling module plays a central role in asymmetric cell division in multiple organisms and system adaptation (Fig. 1) (Blumer et al. 2007; Gonczy 2008; Knoblich 2010). Determination of the X-ray crystal structure of the GPR/GoLoco peptide-Gαi1 complex provides a structural basis for understanding interaction of the motif with GαGDP (Bosch et al. 2011; Kimple et al. 2002). Structure-activity studies have revealed specific amino acids in the GPR motif that influence interaction with Gα (Peterson et al. 2002) and a key residue in Gα that is required for interaction with GPR motifs (Bosch et al. 2011).

Mechanistically, proteins with GPR motifs may impart biological activity by influencing subunit interactions to promote or sustain dissociation of Gαβγ heterotrimers in the absence of nucleotide exchange. In the context of the G-protein activation-deactivation cycle, the GPR protein may also influence subunit interactions by binding GαGDP prior to reassociation with Gβγ. In either situation, alterations in Gβγ-regulated effector pathways would be expected. The GαGDP-GPR complex itself is actually suggested to be a putative “bioactive” entity (Gonczy 2008). The Gα-GPR signaling module can also function as a discreet signaling system that is regulated by non-receptor guanine nucleotide exchange factors and perhaps G-protein-coupled receptors. Interplay between the Gα-GPR and the classical heterotrimer Gαβγ signaling modules offers additional interesting mechanisms for signal integration (Blumer et al. 2011).

A nonsense mutation in GPSM2, which truncates the reading frame of the protein, is associated with nonsyndromic hearing loss likely as a result of altered planar cell polarity in the auditory system (Walsh et al. 2010; Yariz et al. 2011). Renal AGS3 (GPSM1) is markedly elevated in polycystic kidney disease and in response to renal injury (Nadella et al. 2010; Regner et al. 2011). In cell culture, AGS3 traffics into the aggresome pathway and it is also a central player in regulating autophagy (Garcia-Marcos et al. 2011).

Group III AGS Proteins

The mechanisms by which the Group III AGS proteins activate G-protein signaling in the yeast functional screen and function in mammalian signaling systems are not yet fully understood. As more information becomes available, it is likely that members of this loosely defined Group III will exhibit different mechanisms of action in terms of their ability to lead to the end readout of Gβγ-dependent growth in the yeast functional screen. One mechanism by which the Group III proteins may act is by an interaction with Gβγ and/or heterotrimer to influence subunit interactions in a way that there is productive effector engagement. AGS2 (tctex1/DYNLT1) is actually a light chain for the cytoplasmic motor protein dynein and regulates neurite outgrowth through its interaction with Gβγ and dynein (Sachdev et al. 2007). AGS7 (TRIP13) was identified as a thyroid receptor–interacting protein and AGS8 (FNDC1) promotes apoptosis of cardiac myocytes. AGS9 (Rpn10) is a component of the 26S proteasome.

Summary

The discovery of AGS and related proteins altered our basic concepts of G-protein signaling. First, Gα and Gβγ are processing signals within the cell distinct from their role as transducers for cell surface receptors and such signals involve previously unrecognized functional roles for heterotrimeric G-protein subunits. Secondly, Gα and Gβγ may complex with alternative binding partners independent of the classical Gαβγ heterotrimer, providing a distinct signaling pathway with its own set of activators and effectors.

AGS and related accessory proteins or signal regulators are intimately involved in generating signaling diversity in ways that are not yet fully recognized. Such accessory proteins have evolved to provide a mechanism for cells to adapt acutely and for a longer-term to physiological and pathological challenges without altering the major components of the signaling core itself. AGS proteins and related entities play unexpected and important functional roles in a number of systems and impact a number of signaling pathways that influence cell growth and survival. Rapidly accumulating data from disease tissue profiling and genomic-based technologies indicate that selected AGS proteins may serve as biomarkers for specific diseases and their altered expression or function in disease states suggests candidate signaling modules for therapeutic targeting.

Central questions in the field are as follows: What regulates the subcellular location of AGS proteins? What regulates the interaction of AGS proteins with Gα or Gβγ? What is downstream of the GPR-Gα signaling module? Are there pathologies associated with polymorphisms in AGS proteins? Are AGS proteins candidate therapeutic targets? How did the different “G-switch” modules (e.g., Gα-GPR, Gαβγ) and their regulation by non-receptor and cell surface receptors evolve in response to different evolutionary pressures in terms of signal processing?

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© Springer International Publishing AG 2018

Authors and Affiliations

  1. 1.Department of Cell and Molecular Pharmacology and Experimental TherapeuticsMedical University of South CarolinaCharlestonUSA
  2. 2.Department of NeurosciencesMedical University of South CarolinaCharlestonUSA