Cargando…

Ratiometric measurement of MAM Ca(2+) dynamics using a modified CalfluxVTN

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+) conce...

Descripción completa

Detalles Bibliográficos
Autores principales: 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
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10276021/
https://www.ncbi.nlm.nih.gov/pubmed/37328454
http://dx.doi.org/10.1038/s41467-023-39343-2
Descripción
Sumario: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.