Ion Channels: Solving a specificity mystery
Being able to quickly respond to danger is an essential survival skill. In humans, the sympathetic branch of the autonomic nervous system is responsible for the body’s ‘fight-or-flight’ response. It activates the physiological changes we perceive as an adrenaline rush, including a rapid increase in heart rate mediated by beta adrenergic receptors in the pacemaker cells of the heart. When the danger has passed, the parasympathetic branch of the nervous system activates an alternative ‘rest-and-digest’ program, which slows down the heart by activating other receptors called muscarinic acetylcholine receptors.
Despite their opposite effects, these receptors are both G protein-coupled receptors (GPCRs). When activated, the receptors in this family signal by causing a G protein to split into a Gα subunit and a Gβγ subunit (Figure 1). A G protein can have a Gαs or a Gαi/o subunit, among other possibilities. Beta adrenergic receptors prefer to interact with G proteins that contain the former, whereas muscarinic acetylcholine receptors (M2Rs) favor G proteins that contain the latter. A longstanding mystery in cell biology is why the Gβγ subunits released by M2Rs are able to activate potassium channels called GIRKs, which causes the heart rate to drop, whereas the same Gβγ subunits released by beta adrenergic receptors cannot (Dascal, 1997).
One possibility is that M2Rs and GIRKs can form large complexes, which means that the M2Rs could release the Gβγ subunits right where they are needed (Clancy et al., 2005; Riven et al., 2006), whereas beta adrenergic receptors and GIRKs may not form such complexes and therefore would not benefit from a proximity effect. Now, in eLife, Kouki Touhara and Roderick MacKinnon of the Rockefeller University report that the mystery has a different solution (Touhara and MacKinnon, 2018).
Touhara and MacKinnon assessed whether the specificity of GIRKs for M2Rs compared to beta adrenergic receptors is universal, confirming that only M2Rs could activate the potassium channels regardless of the cell line tested. They also showed that the formation of a M2R-GIRK complex was neither necessary nor sufficient to explain why Gβγ subunits released by M2Rs can activate GIRKs and those released by beta adrenergic receptors cannot. So, what, then, is the explanation?
The next clue came from investigating the effect of G protein levels on GIRK signaling. Touhara and MacKinnon confirmed that when native levels of G proteins were present, only M2Rs could activate GIRK channels. However, when Gαsβγ levels were increased, beta adrenergic receptors were also able to activate the channels. These results even extended to non-GIRK channels, in which the researchers find the same pattern, further suggesting that under normal conditions the limited availability of G proteins allows M2Rs to selectively modulate ion channels.
To better understand the role of G protein levels, the rate at which each receptor causes Gβγ to split from Gα was measured. This revealed that M2Rs break up G proteins more quickly than adrenergic receptors do. With this information in hand, a mathematical model was constructed, incorporating previously measured values for reaction rates. The results suggest that the specific GIRK-GPCR signaling could be because G proteins containing Gαi/o subunits associate more quickly with M2Rs than the G proteins that contain Gαs subunits do with beta adrenergic receptors. This results in M2Rs liberating Gβγ subunits more quickly so that they accumulate to the high levels required for GIRK activation, and could explain how differences in a single association-rate constant can allow the parasympathetic and sympathetic branches of the nervous system to control heart rate without interfering with each other (Figure 1).
While the kinetic model offers a nearly complete picture, Touhara and MacKinnon point out that their model required a higher receptor concentration than expected. This is consistent with previous work showing that GPCRs may be preferentially concentrated in local ‘hotspot’ regions of the cell membrane (Sungkaworn et al., 2017). This could offer yet another level of regulation, providing an exciting avenue for future research. Other areas to explore include the molecular basis for the fast association of Gαiβγ with M2Rs, and how other GPCR-GIRK signaling systems have been tuned for the diverse biological roles they play.
Signalling via the G protein-activated K+ channelsCellular Signalling 9:551–573.https://doi.org/10.1016/S0898-6568(97)00095-8
Article and author information
- Version of Record published: January 16, 2019 (version 1)
© 2019, Zheng and Kruse
This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.
- Page views
Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
- Structural Biology and Molecular Biophysics
Class 1 cytokine receptors transmit signals through the membrane by a single transmembrane helix to an intrinsically disordered cytoplasmic domain that lacks kinase activity. While specific binding to phosphoinositides has been reported for the prolactin receptor (PRLR), the role of lipids in PRLR signalling is unclear. Using an integrative approach combining NMR spectroscopy, cellular signalling experiments, computational modelling and simulation, we demonstrate co-structure formation of the disordered intracellular domain of the human PRLR, the membrane constituent phosphoinositide-4,5-bisphosphate (PI(4,5)P2) and the FERM-SH2 domain of the Janus kinase 2 (JAK2). We find that the complex leads to accumulation of PI(4,5)P2 at the transmembrane helix interface and that mutation of residues identified to interact specifically with PI(4,5)P2 negatively affects PRLR-mediated activation of signal transducer and activator of transcription 5 (STAT5). Facilitated by co-structure formation, the membrane-proximal disordered region arranges into an extended structure. We suggest that the co-structure formed between PRLR, JAK2 and PI(4,5)P2 locks the juxtamembrane disordered domain of the PRLR in an extended structure, enabling signal relay from the extracellular to the intracellular domain upon ligand binding. We find that the co-structure exists in different states which we speculate could be relevant for turning signalling on and off. Similar co-structures may be relevant for other non-receptor tyrosine kinases and their receptors.
- Biochemistry and Chemical Biology
- Structural Biology and Molecular Biophysics
Aurora B, together with IN-box, the C-terminal part of INCENP, forms an enzymatic complex that ensures faithful cell division. The [Aurora B/IN-box] complex is activated by autophosphorylation in the Aurora B activation loop and in IN-box, but it is not clear how these phosphorylations activate the enzyme. We used a combination of experimental and computational studies to investigate the effects of phosphorylation on the molecular dynamics and structure of [Aurora B/IN-box]. In addition, we generated partially phosphorylated intermediates to analyze the contribution of each phosphorylation independently. We found that the dynamics of Aurora and IN-box are interconnected, and IN-box plays both positive and negative regulatory roles depending on the phosphorylation status of the enzyme complex. Phosphorylation in the activation loop of Aurora B occurs intramolecularly and prepares the enzyme complex for activation, but two phosphorylated sites are synergistically responsible for full enzyme activity.