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G protein-coupled receptors mobilize intracellular calcium via the Gs-βγ-PLCβ module

dc.contributor.advisorKostenis, Evi
dc.contributor.authorBrands, Julian Maximilian Heinrich
dc.date.accessioned2024-05-22T07:33:17Z
dc.date.available2025-06-02T22:00:35Z
dc.date.issued22.05.2024
dc.identifier.urihttps://hdl.handle.net/20.500.11811/11555
dc.description.abstractAccurate regulation of calcium is essential for many physiological processes. To fine-tune these processes, the cell can draw on a large portfolio of diverse mechanisms to govern calcium mobilization, in which GPCRs present an integral component. A key element of how the cell mobilizes calcium using GPCRs involves the large group of phospholipases Cß. These enzymes are canonically activated by Gq-coupled receptors, but it has been found that Gi coupled receptors can also activate PLCß2 and PLCß3 via their Gßγ subunit. Notably, this Gßγ-specific effect is fully dependent on active Gq. Upon activation, PLCß enzymes hydrolyze the membrane component PIP2 to DAG and IP3. While DAG triggers various cellular effectors, IP3 mediates a cytosolic increase in calcium via activation of endoplasmically located InsP3R.
By studying the Gi-ßγ-PLCß-Ca2+ pathway and identifying active Gq as the driver of Gi-ßγ-mediated Ca2+release, we came up with a new hypothesis. Could Gq be the missing link that would allow even other members of the G protein family to release calcium via their Gßγ subunit? Therefore, we hypothesized that activation of Gq may permit Gs-ßγ to activate PLCß enzymes, resulting in the release of intracellularly stored calcium. To test this hypothesis, we used a variety of CRISPR/Cas9-edited cells, engineered molecules, and inhibitors alongside illuminating techniques such as fluorescence-based calcium measurement, HTRF-based cAMP accumulation, label-free whole-cell biosensing, real-time BRET and Nanobit measurements.
However, examination of Gq-dependent Gs-calcium revealed that Gs-GPCRs trigger more than one Gq-dependent calcium release pathway in non-excitable HEK293 cells. This exploration led us to discriminate two distinct pathways utilized by Gs-GPCRs: i) a cAMP-dependent and ii) a cAMP-independent, previously unrecognized pathway. Both pathways are tightly governed by active Gaq, yet they diverge in terms of the underlying action. Because Gas mainly orchestrates cAMP-mediated effects, we focused on the unknown cAMP-independent Gs-mediated calcium release and were able to identify Gs-ßγ as the driving force. It appears that the Gs-ßγ module activates Gßγ-sensitive PLCß isoforms that mediate a calcium release from intracellular stores. Given the cell- and tissue-specific nature of calcium regulation, we extended our investigation to cells of greater physiological relevance. Thereby, we revealed a Gq-dependent Gs-mediated calcium response even in primary cell lines, including mouse brown adipose tissue and mouse embryonic fibroblasts, expanding the physiological significance of Gs-ßγ-mediated calcium release. Therefore, our results illustrate the remarkable adaptability of GPCR calcium signaling and highlight its capacity to rapidly adapt to ever-changing conditions (externally & internally). The Gs-GPCR system appears to be a competent regulator of calcium dynamics, whether through Gas-mediated, cAMP-dependent mechanisms or through a Gs-ßγ-mediated and cAMP-independent mechanism.
en
dc.language.isoeng
dc.rightsIn Copyright
dc.rights.urihttp://rightsstatements.org/vocab/InC/1.0/
dc.subject.ddc500 Naturwissenschaften
dc.subject.ddc570 Biowissenschaften, Biologie
dc.titleG protein-coupled receptors mobilize intracellular calcium via the Gs-βγ-PLCβ module
dc.typeDissertation oder Habilitation
dc.publisher.nameUniversitäts- und Landesbibliothek Bonn
dc.publisher.locationBonn
dc.rights.accessRightsopenAccess
dc.identifier.urnhttps://nbn-resolving.org/urn:nbn:de:hbz:5-75862
ulbbn.pubtypeErstveröffentlichung
ulbbnediss.affiliation.nameRheinische Friedrich-Wilhelms-Universität Bonn
ulbbnediss.affiliation.locationBonn
ulbbnediss.thesis.levelDissertation
ulbbnediss.dissID7586
ulbbnediss.date.accepted11.04.2024
ulbbnediss.instituteMathematisch-Naturwissenschaftliche Fakultät : Fachgruppe Pharmazie / Pharmazeutische Biologie
ulbbnediss.fakultaetMathematisch-Naturwissenschaftliche Fakultät
dc.contributor.coRefereeFleischmann, Bernd
ulbbnediss.date.embargoEndDate02.06.2025
ulbbnediss.contributor.gnd136786447X


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