AGS3-based optogenetic GDI induces GPCR-independent Gβγ signalling and macrophage migration
G-protein-coupled receptors (GPCRs) are efficient guanine nucleotide exchange factors (GEFs) and exchange GDP to GTP on the Gα subunit of G-protein heterotrimers in response to various extracellular stimuli, including neurotransmitters and light. GPCRs primarily broadcast signals through activated G...
Saved in:
| Main Authors: | , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
The Royal Society
2025-02-01
|
| Series: | Open Biology |
| Subjects: | |
| Online Access: | https://royalsocietypublishing.org/doi/10.1098/rsob.240181 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1832540446756175872 |
|---|---|
| author | Waruna Thotamune Sithurandi Ubeysinghe Chathuri Rajarathna Dinesh Kankanamge Koshala Olupothage Aditya Chandu Bryan A. Copits Ajith Karunarathne |
| author_facet | Waruna Thotamune Sithurandi Ubeysinghe Chathuri Rajarathna Dinesh Kankanamge Koshala Olupothage Aditya Chandu Bryan A. Copits Ajith Karunarathne |
| author_sort | Waruna Thotamune |
| collection | DOAJ |
| description | G-protein-coupled receptors (GPCRs) are efficient guanine nucleotide exchange factors (GEFs) and exchange GDP to GTP on the Gα subunit of G-protein heterotrimers in response to various extracellular stimuli, including neurotransmitters and light. GPCRs primarily broadcast signals through activated G proteins, GαGTP and free Gβγ and are major disease drivers. Evidence shows that the ambient low threshold signalling required for cells is likely supplemented by signalling regulators such as non-GPCR GEFs and guanine nucleotide dissociation inhibitors (GDIs). Activators of G-protein signalling 3 (AGS3) are recognized as a GDI involved in multiple health and disease-related processes. Nevertheless, understanding of AGS3 is limited, and no significant information is available on its structure–function relationship or signalling regulation in living cells. Here, we employed in silico structure-guided engineering of a novel optogenetic GDI, based on the AGS3’s G-protein regulatory motif, to understand its GDI activity and induce standalone Gβγ signalling in living cells on optical command. Our results demonstrate that plasma membrane recruitment of OptoGDI efficiently releases Gβγ, and its subcellular targeting generated localized PIP3 and triggered macrophage migration. Therefore, we propose OptoGDI as a powerful tool for optically dissecting GDI-mediated signalling pathways and triggering GPCR-independent Gβγ signalling in cells and in vivo. |
| format | Article |
| id | doaj-art-354a10f30b574cb09df0dc40bb263277 |
| institution | Kabale University |
| issn | 2046-2441 |
| language | English |
| publishDate | 2025-02-01 |
| publisher | The Royal Society |
| record_format | Article |
| series | Open Biology |
| spelling | doaj-art-354a10f30b574cb09df0dc40bb2632772025-02-05T00:05:20ZengThe Royal SocietyOpen Biology2046-24412025-02-0115210.1098/rsob.240181AGS3-based optogenetic GDI induces GPCR-independent Gβγ signalling and macrophage migrationWaruna Thotamune0Sithurandi Ubeysinghe1Chathuri Rajarathna2Dinesh Kankanamge3Koshala Olupothage4Aditya Chandu5Bryan A. Copits6Ajith Karunarathne7Department of Chemistry, Saint Louis University, Saint Louis, MO 63103, USADepartment of Chemistry, Saint Louis University, Saint Louis, MO 63103, USADepartment of Chemistry, Saint Louis University, Saint Louis, MO 63103, USADepartment of Anesthesiology, Washington University Pain Center, Washington University School of Medicine, St. Louis, MO 63110, USADepartment of Chemistry and Biochemistry, The University of Toledo, Toledo, OH 43606, USADepartment of Chemistry, Saint Louis University, Saint Louis, MO 63103, USADepartment of Anesthesiology, Washington University Pain Center, Washington University School of Medicine, St. Louis, MO 63110, USADepartment of Chemistry, Saint Louis University, Saint Louis, MO 63103, USAG-protein-coupled receptors (GPCRs) are efficient guanine nucleotide exchange factors (GEFs) and exchange GDP to GTP on the Gα subunit of G-protein heterotrimers in response to various extracellular stimuli, including neurotransmitters and light. GPCRs primarily broadcast signals through activated G proteins, GαGTP and free Gβγ and are major disease drivers. Evidence shows that the ambient low threshold signalling required for cells is likely supplemented by signalling regulators such as non-GPCR GEFs and guanine nucleotide dissociation inhibitors (GDIs). Activators of G-protein signalling 3 (AGS3) are recognized as a GDI involved in multiple health and disease-related processes. Nevertheless, understanding of AGS3 is limited, and no significant information is available on its structure–function relationship or signalling regulation in living cells. Here, we employed in silico structure-guided engineering of a novel optogenetic GDI, based on the AGS3’s G-protein regulatory motif, to understand its GDI activity and induce standalone Gβγ signalling in living cells on optical command. Our results demonstrate that plasma membrane recruitment of OptoGDI efficiently releases Gβγ, and its subcellular targeting generated localized PIP3 and triggered macrophage migration. Therefore, we propose OptoGDI as a powerful tool for optically dissecting GDI-mediated signalling pathways and triggering GPCR-independent Gβγ signalling in cells and in vivo.https://royalsocietypublishing.org/doi/10.1098/rsob.240181GPCRG proteinsGDIoptogeneticsmacrophage migrationactivators of G-protein signalling 3 |
| spellingShingle | Waruna Thotamune Sithurandi Ubeysinghe Chathuri Rajarathna Dinesh Kankanamge Koshala Olupothage Aditya Chandu Bryan A. Copits Ajith Karunarathne AGS3-based optogenetic GDI induces GPCR-independent Gβγ signalling and macrophage migration Open Biology GPCR G proteins GDI optogenetics macrophage migration activators of G-protein signalling 3 |
| title | AGS3-based optogenetic GDI induces GPCR-independent Gβγ signalling and macrophage migration |
| title_full | AGS3-based optogenetic GDI induces GPCR-independent Gβγ signalling and macrophage migration |
| title_fullStr | AGS3-based optogenetic GDI induces GPCR-independent Gβγ signalling and macrophage migration |
| title_full_unstemmed | AGS3-based optogenetic GDI induces GPCR-independent Gβγ signalling and macrophage migration |
| title_short | AGS3-based optogenetic GDI induces GPCR-independent Gβγ signalling and macrophage migration |
| title_sort | ags3 based optogenetic gdi induces gpcr independent gβγ signalling and macrophage migration |
| topic | GPCR G proteins GDI optogenetics macrophage migration activators of G-protein signalling 3 |
| url | https://royalsocietypublishing.org/doi/10.1098/rsob.240181 |
| work_keys_str_mv | AT warunathotamune ags3basedoptogeneticgdiinducesgpcrindependentgbgsignallingandmacrophagemigration AT sithurandiubeysinghe ags3basedoptogeneticgdiinducesgpcrindependentgbgsignallingandmacrophagemigration AT chathurirajarathna ags3basedoptogeneticgdiinducesgpcrindependentgbgsignallingandmacrophagemigration AT dineshkankanamge ags3basedoptogeneticgdiinducesgpcrindependentgbgsignallingandmacrophagemigration AT koshalaolupothage ags3basedoptogeneticgdiinducesgpcrindependentgbgsignallingandmacrophagemigration AT adityachandu ags3basedoptogeneticgdiinducesgpcrindependentgbgsignallingandmacrophagemigration AT bryanacopits ags3basedoptogeneticgdiinducesgpcrindependentgbgsignallingandmacrophagemigration AT ajithkarunarathne ags3basedoptogeneticgdiinducesgpcrindependentgbgsignallingandmacrophagemigration |