Heterotrimeric Gq proteins as therapeutic targets? Kostenis, Evi; Pfeil, Eva Marie; Annala, Suvi
Journal of biological chemistry/The Journal of biological chemistry,
04/2020, Volume:
295, Issue:
16
Journal Article
Peer reviewed
Open access
Heterotrimeric G proteins are the core upstream elements that transduce and amplify the cellular signals from G protein–coupled receptors (GPCRs) to intracellular effectors. GPCRs are the largest ...family of membrane proteins encoded in the human genome and are the targets of about one-third of prescription medicines. However, to date, no single therapeutic agent exerts its effects via perturbing heterotrimeric G protein function, despite a plethora of evidence linking G protein malfunction to human disease. Several recent studies have brought to light that the Gq family–specific inhibitor FR900359 (FR) is unexpectedly efficacious in silencing the signaling of Gq oncoproteins, mutant Gq variants that mostly exist in the active state. These data not only raise the hope that researchers working in drug discovery may be able to potentially strike Gq oncoproteins from the list of undruggable targets, but also raise questions as to how FR achieves its therapeutic effect. Here, we place emphasis on these recent studies and explain why they expand our pharmacological armamentarium for targeting Gq protein oncogenes as well as broaden our mechanistic understanding of Gq protein oncogene function. We also highlight how this novel insight impacts the significance and utility of using G(q) proteins as targets in drug discovery efforts.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Endogenous opioid peptides and prescription opioid drugs modulate pain, anxiety and stress by activating opioid receptors, currently classified into four subtypes. Here we demonstrate that ...ACKR3/CXCR7, hitherto known as an atypical scavenger receptor for chemokines, is a broad-spectrum scavenger of opioid peptides. Phylogenetically, ACKR3 is intermediate between chemokine and opioid receptors and is present in various brain regions together with classical opioid receptors. Functionally, ACKR3 is a scavenger receptor for a wide variety of opioid peptides, especially enkephalins and dynorphins, reducing their availability for the classical opioid receptors. ACKR3 is not modulated by prescription opioids, but we show that an ACKR3-selective subnanomolar competitor peptide, LIH383, can restrain ACKR3's negative regulatory function on opioid peptides in rat brain and potentiate their activity towards classical receptors, which may open alternative therapeutic avenues for opioid-related disorders. Altogether, our results reveal that ACKR3 is an atypical opioid receptor with cross-family ligand selectivity.
Immune cells congregate at specific loci to fight infections during inflammatory responses, a process that must be transient and self-resolving. Cell dispersal promotes resolution, but it remains ...unclear how transition from clustering to dispersal is regulated. Here we show, using quantitative live imaging in zebrafish, that differential ligand-induced trafficking of chemokine receptors such as Cxcr1 and Cxcr2 orchestrates the state of neutrophil congregation at sites of tissue damage. Through receptor mutagenesis and biosensors, we show that Cxcr1 promotes clustering at wound sites, but is promptly desensitized and internalized, which prevents excess congregation. By contrast, Cxcr2 promotes bidirectional motility and is sustained at the plasma membrane. Persistent plasma membrane residence of Cxcr2 prolongs downstream signaling and is required for sustained exploratory motion conducive to dispersal. Thus, differential trafficking of two chemokine receptors allows coordination of antagonistic cell behaviors, promoting a self-resolving migratory response.
Some 865 genes in man encode G‐protein‐coupled receptors (GPCRs). The heterotrimeric guanine nucleotide‐binding proteins (G‐proteins) function to transduce signals from this vast panoply of receptors ...to effector systems including ion channels and enzymes that alter the rate of production, release or degradation of intracellular second messengers. However, it was not until the 1970s that the existence of such transducing proteins was even seriously suggested. Combinations of bacterial toxins that mediate their effects via covalent modification of the α‐subunit of certain G‐proteins and mutant cell lines that fail to generate cyclic AMP in response to agonists because they either fail to express or express a malfunctional G‐protein allowed their identification and purification. Subsequent to initial cloning efforts, cloning by homology has defined the human G‐proteins to derive from 35 genes, 16 encoding α‐subunits, five β and 14 γ. All function as guanine nucleotide exchange on–off switches and are mechanistically similar to other proteins that are enzymic GTPases. Although not readily accepted initially, it is now well established that β/γ complexes mediate as least as many functions as the α‐subunits. The generation of chimeras between different α‐subunits defined the role of different sections of the primary/secondary sequence and crystal structures and cocrystals with interacting proteins have given detailed understanding of their molecular structure and basis of function. Finally, further modifications of such chimeras have generated a range of G‐protein α‐subunits with greater promiscuity to interact across GPCR classes and initiated the use of such modified G‐proteins in drug discovery programmes.
British Journal of Pharmacology (2006) 147, S46–S55. doi:10.1038/sj.bjp.0706405
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BFBNIB, DOBA, FZAB, GIS, IJS, IZUM, KILJ, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBMB, SIK, UILJ, UKNU, UL, UM, UPUK
G protein-independent, arrestin-dependent signaling is a paradigm that broadens the signaling scope of G protein-coupled receptors (GPCRs) beyond G proteins for numerous biological processes. ...However, arrestin signaling in the collective absence of functional G proteins has never been demonstrated. Here we achieve a state of "zero functional G" at the cellular level using HEK293 cells depleted by CRISPR/Cas9 technology of the Gs/q/12 families of Gα proteins, along with pertussis toxin-mediated inactivation of Gi/o. Together with HEK293 cells lacking β-arrestins ("zero arrestin"), we systematically dissect G protein- from arrestin-driven signaling outcomes for a broad set of GPCRs. We use biochemical, biophysical, label-free whole-cell biosensing and ERK phosphorylation to identify four salient features for all receptors at "zero functional G": arrestin recruitment and internalization, but-unexpectedly-complete failure to activate ERK and whole-cell responses. These findings change our understanding of how GPCRs function and in particular of how they activate ERK1/2.
Abstract
Obesity is the major driver of the global epidemic in type 2 diabetes (T2D). In individuals with obesity, impaired insulin action leads to increased lipolysis in adipocytes, resulting in ...elevated plasma free fatty acid (FFA) levels that promote peripheral insulin resistance, a hallmark of T2D. Here we show, by using a combined genetic/biochemical/pharmacologic approach, that increased adipocyte lipolysis can be prevented by selective activation of adipocyte G
q
signaling in vitro and in vivo (in mice). Activation of this pathway by a G
q
-coupled designer receptor or by an agonist acting on an endogenous adipocyte G
q
-coupled receptor (CysLT
2
receptor) greatly improved glucose and lipid homeostasis in obese mice or in mice with adipocyte insulin receptor deficiency. Our findings identify adipocyte G
q
signaling as an essential regulator of whole-body glucose and lipid homeostasis and should inform the development of novel classes of GPCR-based antidiabetic drugs.
GPCRs modulate a plethora of physiological processes and mediate the effects of one‐third of FDA‐approved drugs. Depending on which ligand activates a receptor, it can engage different intracellular ...transducers. This ‘biased signalling’ paradigm requires that we now characterize physiological signalling not just by receptors but by ligand–receptor pairs. Ligands eliciting biased signalling may constitute better drugs with higher efficacy and fewer adverse effects. However, ligand bias is very complex, making reproducibility and description challenging. Here, we provide guidelines and terminology for any scientists to design and report ligand bias experiments. The guidelines will aid consistency and clarity, as the basic receptor research and drug discovery communities continue to advance our understanding and exploitation of ligand bias. Scientific insight, biosensors, and analytical methods are still evolving and should benefit from and contribute to the implementation of the guidelines, together improving translation from in vitro to disease‐relevant in vivo models.
Full text
Available for:
BFBNIB, DOBA, FZAB, GIS, IJS, IZUM, KILJ, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBMB, SIK, UILJ, UKNU, UL, UM, UPUK
The potent and selective Gq protein inhibitor depsipeptide FR900359 (FR), originally discovered as the product of an uncultivable plant endosymbiont, is synthesized by a complex biosynthetic system ...comprising two nonribosomal peptide synthetase (NRPS) assembly lines. Here we characterize a cultivable bacterial FR producer, enabling detailed investigations into biosynthesis and attachment of the functionally important FR side chain. We reconstitute side chain assembly by the monomodular NRPS FrsA and the non-heme monooxygenase FrsH, and characterize intermolecular side chain transesterification to the final macrocyclic intermediate FR-Core, mediated by the FrsA thioesterase domain. We harness FrsA substrate promiscuity to generate FR analogs with altered side chains and demonstrate indispensability of the FR side chain for efficient Gq inhibition by comparative bioactivity, toxicity and docking studies. Finally, evolution of FR and side chain biosynthesis is discussed based on bioinformatics analyses. Side chain transesterification boosts potency and target affinity of selective Gq inhibitor natural products.
G protein-coupled receptors (GPCRs) transmit extracellular signals to the inside by activation of intracellular effector proteins. Different agonists can promote differential receptor-induced ...signaling responses – termed bias – potentially by eliciting different levels of recruitment of effector proteins. As activation and recruitment of effector proteins might influence each other, thorough analysis of bias is difficult. Here, we compared the efficacy of seven agonists to induce G protein, G protein-coupled receptor kinase 2 (GRK2), as well as arrestin3 binding to the muscarinic acetylcholine receptor M
3
by utilizing FRET-based assays. In order to avoid interference between these interactions, we studied GRK2 binding in the presence of inhibitors of G
i
and G
q
proteins and analyzed arrestin3 binding to prestimulated M
3
receptors to avoid differences in receptor phosphorylation influencing arrestin recruitment. We measured substantial differences in the agonist efficacies to induce M
3
R-arrestin3 versus M
3
R-GRK2 interaction. However, the rank order of the agonists for G protein- and GRK2-M
3
R interaction was the same, suggesting that G protein and GRK2 binding to M
3
R requires similar receptor conformations, whereas requirements for arrestin3 binding to M
3
R are distinct.
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