Communication between and within cells represents one of the most important traits of living matter. Information is usually transported in form of chemical substances (hormones, neurotransmitters, proteins, etc.) through the extracellular space, as well as between intracellular compartments. Reception and transmission of extracellular information into the cell’s interior is often realized by membrane receptors. Of these, G- protein-coupled receptors (GPCRs) constitute the largest group in the
Already some decades ago, changes of intracellular calcium concentrations (Ca2+i) following single extracellular stimuli were found to not always occur as transient, monophasic responses but showed a repetitive or oscillating behavior [25]. Ca2+i oscillations have since then been investigated as possible time codes for biological information [26]. Gq-coupled GPCRs are prominent regulators of Ca2+i waves, a discovery underlined by early findings of repetitive calcium spikes in hepatocytes
Guanine nucleotide-binding proteins (G proteins) are essential transducers in GPCR signaling pathways. G proteins can be classified into monomeric (or small) and heterotrimeric G proteins (consisting of three subunits: Gα, Gβ, and Gγ) [2]. They are commonly described as molecular switches, which are in an ‘on’ [guanosine triphosphate (GTP)-bound] or an ‘off’ [guanosine diphosphate (GDP)-bound] state. G-protein activation is initiated by the dissociation of GDP from and subsequent binding of GTP
An initial receptor impulse may be generated at the plasma membrane by what is generally thought of as ‘canonical’, G-protein-mediated signaling: surface GPCRs activate G proteins at the plasma membrane that then dissociate from the receptor to interact with effector proteins at the membrane or at intracellular locations. Delayed signal impulses emerge either from (or near) the cell membrane or from internalized structures incorporating additional transducers (Figure 3A). The detailed
Recently, the concept of drug residence time for compound selection and optimization in drug discovery came to the fore and has been discussed elsewhere in detail 71, 72. Instead of focusing on the Kd values of the ligands as a measure of affinity, it was proposed to give more attention to their binding kinetics, particularly the koff values, because slowly dissociating ligands have been proven successful as antagonists [73]. The molecular mechanisms governing how such kinetically biased
Ligands targeting GPCRs can activate distinct pathways from a multitude of signaling routes, which is also denoted as ligand bias. Apart from subcellular environment (location) and physical composition (quality), GPCR ligands also employ dynamic aspects (time) to convey information from cell to cell. We therefore propose to extend the current view of signaling bias by a kinetic component: temporal bias. Temporal bias is distinct from ‘conformational bias’, where ligands induce different
The authors declare that there are no conflicts of interest. What are the manifestations of signaling dynamics? How are temporal and spatial aspects interlinked? Can we find a logic behind where and when a signaling event happens? What are the determinants in the dynamics of proteins that define a temporal code? What are the rhythmic patterns of conformational ensembles that dictate the signaling outcome? How are the different time scales connected, from microsecond molecular switches within proteins
First, we targeted Gαq. The Gαq protein is an activator of phospholipase Cβ (PLCβ), and activation of Gαq triggers Ca2+ release from the endoplasmic reticulum (ER) to the cytoplasm, and subsequent Ca2+ oscillations (Grundmann and Kostenis, 2017; Waldo et al., 2010). To control Gαq localization and activity using mDcTMP, a palmitoylation-deficient constitutively active form of Gαq was fused at the C terminus to mNeonGreen-tagged iK6DHFR (mNG-iK6DHFR-Gαq) and expressed in HeLa cells (Figure 3A).
In contrast, hM3Dq (a Gαq DREADD) can directly regulate [Ca2+]i signaling through the ubiquitous Gαq-PLCβ-IP3 pathway and IP3-mediated release of Ca2+ from the endoplasmic reticulum (ER) [59]. This pathway is one of the most commonly utilized calcium release mechanisms in cells [60], and is intimately associated with chondrocyte mechanotransduction [61] and oscillatory [Ca2+]i signaling behaviors [62]. Thus, hM3Dq represents an innovative tool for specifically and synthetically regulating GPCR-dependent [Ca2+]i signaling, and Gαq-dependent signaling more generally, having been leveraged with transformative success in the neurological sciences [63,64] and to control the metabolic activity of a number of non-neuronal cell types (e.g., liver, kidney, pancreas) [65–67].