Ratiometric measurement of MAM Ca2+ dynamics using a modified CalfluxVTN

• Published in: Nature communications Vol. 14; no. 1; p. 3586
• Main Authors: Cho, Eunbyul, Woo, Youngsik, Suh, Yeongjun, Suh, Bo Kyoung, Kim, Soo Jeong, Nhung, Truong Thi My, Yoo, Jin Yeong, Nghi, Tran Diem, Lee, Su Been, Mun, Dong Jin, Park, Sang Ki
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 16.06.2023; Nature Publishing Group; Nature Portfolio
• Subjects: 14/10; 14/5; 631/1647/1888/2249; 631/1647/245/2225; 631/45/321/1154; 631/61/350/59; 631/80/642; Alzheimer's disease; Biological activity; Bioluminescence; Calcium (intracellular); Calcium imaging; Calcium ions; Calcium signalling; Crosstalk; Humanities and Social Sciences; Kinases; Localization; Mitochondria; Movement disorders; multidisciplinary; Neurodegenerative diseases; Organelles; Parkinson's disease; Physiology; Proteins; Science; Science (multidisciplinary); Scientific imaging; Sensors; Steady state
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-023-39343-2

Mitochondria-associated ER membrane (MAM) is a structure where these calcium-regulating organelles form close physical contact sites for efficient Ca 2+ crosstalk. Despite the central importance of MAM Ca 2+ dynamics in diverse biological processes, directly and specifically measuring Ca 2+ concentrations inside MAM is technically challenging. Here, we develop MAM-Calflux, a MAM-specific BRET-based Ca 2+ indicator. The successful application of the bimolecular fluorescence complementation (BiFC) concept highlights Ca 2+ -responsive BRET signals in MAM. The BiFC strategy imparts dual functionality as a Ca 2+ indicator and quantitative structural marker specific for MAM. As a ratiometric Ca 2+ indicator, MAM-Calflux estimates steady-state MAM Ca 2+ levels. Finally, it enables the visualization of uneven intracellular distribution of MAM Ca 2+ and the elucidation of abnormally accumulated MAM Ca 2+ from the neurons of Parkinson’s disease mouse model in both steady-state and stimulated conditions. Therefore, we propose that MAM-Calflux can be a versatile tool for ratiometrically measuring dynamic inter-organellar Ca 2+ communication. MAM Ca 2+ dynamics play an important role in diverse biological processes, but directly and specifically measuring Ca 2+ concentrations in this region is technically challenging. Here the authors report a MAM-specific BRET-based Ca 2+ indicator called MAM-Calflux, which works as both a Ca 2+ indicator and a structural marker due to its ratiometric nature.