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SHR Neuro Cancer Cardio Lipid Metab Microb

Erdoğan, YC; Pilic, J; Gottschalk, B; Yiğit, EN; Zaki, AG; Öztürk, G; Eroğlu, E; Okutan, B; Sommer, NG; Weinberg, AM; Schindl, R; Graier, WF; Malli, R.
Development of a Dual Reporter System to Simultaneously Visualize Ca²⁺ Signals and AMPK Activity.
ACS Sens. 2024; 9(9):4680-4689 Doi: 10.1021/acssensors.4c01058 [OPEN ACCESS]
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Leading authors Med Uni Graz: Erdogan Yusuf Ceyhun; Malli Roland
Co-authors Med Uni Graz: Gottschalk Benjamin; Graier Wolfgang; Okutan Begüm; Pilic Johannes; Schindl Rainer; Sommer Nicole; Weinberg Annelie-Martina
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Abstract:: In this study, we introduce a new separation of phases-based activity reporter of kinase (SPARK) for AMP-activated kinase (AMPK), named AMPK-SPARK, which reports the AMPK activation by forming bright fluorescent clusters. Furthermore, we introduce a dual reporter system, named GCaMP-AMPK-SPARK, by incorporating a single-fluorescent protein (FP)-based Ca²⁺ biosensor, GCaMP6f, into our initial design, enabling simultaneous monitoring of Ca²⁺ levels and AMPK activity. This system offers the essential quality of information by single-channel fluorescence microscopy without the need for coexpression of different biosensors and elaborate filter layouts to overcome spectral limitations. We used AMPK-SPARK to map endogenous AMPK activity in different cell types and visualized the dynamics of AMPK activation in response to various stimuli. Using GCaMP-AMPK-SPARK, we revealed cell-to-cell heterogeneities in AMPK activation by Ca²⁺ mobilization. We anticipate that this dual reporter strategy can be employed to study the intricate interplays between different signaling networks and kinase activities.
Find related publications in this database (using NLM MeSH Indexing): AMP-Activated Protein Kinases - metabolism; Humans - administration & dosage; Calcium - metabolism; Calcium Signaling - administration & dosage; Biosensing Techniques - methods; HEK293 Cells - administration & dosage; Microscopy, Fluorescence - administration & dosage; Animals - administration & dosage; Enzyme Activation - administration & dosage
Find related publications in this database (Keywords): AMPK reporter; Ca2+ biosensor; fluorescencemicroscopy; phase separation-based activity reporter of kinase; SPARK; GCaMP; multiplexing