Assessing real-time signaling and agonist-induced CRHR1 internalization by optical methods
The development of live-cell sensors for real-time measurement of signaling responses, with improved spatial and temporal resolution with respect to classical biochemical methods, has changed our understanding of cellular signaling. Examination of cAMP generation downstream activated GPCRs has shown...
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| Formato: | SER |
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| Acceso en línea: | http://hdl.handle.net/20.500.12110/paper_0091679X_v149_n_p239_dosSantosClaro |
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todo:paper_0091679X_v149_n_p239_dosSantosClaro2023-10-03T14:55:03Z Assessing real-time signaling and agonist-induced CRHR1 internalization by optical methods dos Santos Claro, P.A. Inda, C. Armando, N.G. Piazza, V.G. Attorresi, A. Silberstein, S. Shukla A.K. CRH receptors signaling CRHR1 Cyclic AMP Fluorescent flow cytometry FRET-based biosensors GPCR endocytosis Live-cell imaging The development of live-cell sensors for real-time measurement of signaling responses, with improved spatial and temporal resolution with respect to classical biochemical methods, has changed our understanding of cellular signaling. Examination of cAMP generation downstream activated GPCRs has shown that signaling responses can be short-lived (generated from the cell surface) or prolonged after receptor internalization. Class B secretin-like Corticotropin-releasing hormone receptor 1 (CRHR1) is a key player in stress pathophysiology. By monitoring real-time signaling in living cells, we uncovered cell context-dependent temporal characteristics of CRHR1-elicited cAMP responses and disclosed a specific link between cAMP generation and receptor signaling from internal compartments. We describe technical aspects and elaborate the protocols for cell line expression of Förster resonance energy transfer (FRET)-based biosensors to study the dynamics of cAMP and calcium signaling responses downstream activated CRHR1, live-cell imaging and analysis, and fluorescence flow cytometry to determine receptor levels at the cell surface. © 2019 Elsevier Inc. SER info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_0091679X_v149_n_p239_dosSantosClaro |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
CRH receptors signaling CRHR1 Cyclic AMP Fluorescent flow cytometry FRET-based biosensors GPCR endocytosis Live-cell imaging |
| spellingShingle |
CRH receptors signaling CRHR1 Cyclic AMP Fluorescent flow cytometry FRET-based biosensors GPCR endocytosis Live-cell imaging dos Santos Claro, P.A. Inda, C. Armando, N.G. Piazza, V.G. Attorresi, A. Silberstein, S. Shukla A.K. Assessing real-time signaling and agonist-induced CRHR1 internalization by optical methods |
| topic_facet |
CRH receptors signaling CRHR1 Cyclic AMP Fluorescent flow cytometry FRET-based biosensors GPCR endocytosis Live-cell imaging |
| description |
The development of live-cell sensors for real-time measurement of signaling responses, with improved spatial and temporal resolution with respect to classical biochemical methods, has changed our understanding of cellular signaling. Examination of cAMP generation downstream activated GPCRs has shown that signaling responses can be short-lived (generated from the cell surface) or prolonged after receptor internalization. Class B secretin-like Corticotropin-releasing hormone receptor 1 (CRHR1) is a key player in stress pathophysiology. By monitoring real-time signaling in living cells, we uncovered cell context-dependent temporal characteristics of CRHR1-elicited cAMP responses and disclosed a specific link between cAMP generation and receptor signaling from internal compartments. We describe technical aspects and elaborate the protocols for cell line expression of Förster resonance energy transfer (FRET)-based biosensors to study the dynamics of cAMP and calcium signaling responses downstream activated CRHR1, live-cell imaging and analysis, and fluorescence flow cytometry to determine receptor levels at the cell surface. © 2019 Elsevier Inc. |
| format |
SER |
| author |
dos Santos Claro, P.A. Inda, C. Armando, N.G. Piazza, V.G. Attorresi, A. Silberstein, S. Shukla A.K. |
| author_facet |
dos Santos Claro, P.A. Inda, C. Armando, N.G. Piazza, V.G. Attorresi, A. Silberstein, S. Shukla A.K. |
| author_sort |
dos Santos Claro, P.A. |
| title |
Assessing real-time signaling and agonist-induced CRHR1 internalization by optical methods |
| title_short |
Assessing real-time signaling and agonist-induced CRHR1 internalization by optical methods |
| title_full |
Assessing real-time signaling and agonist-induced CRHR1 internalization by optical methods |
| title_fullStr |
Assessing real-time signaling and agonist-induced CRHR1 internalization by optical methods |
| title_full_unstemmed |
Assessing real-time signaling and agonist-induced CRHR1 internalization by optical methods |
| title_sort |
assessing real-time signaling and agonist-induced crhr1 internalization by optical methods |
| url |
http://hdl.handle.net/20.500.12110/paper_0091679X_v149_n_p239_dosSantosClaro |
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