Targeting mitochondrial calcium transfer to treat amyotrophic lateral sclerosis using the novel photoconvertible ratiometric Ca2+ sensor CaMPARI

IIntracellular membrane contact sites facilitate the transfer of ions, metabolites, and other molecules between subcellular compartments. Mitochondrial-associated membranes (MAMs) of the ER are particularly important for regulating the metabolic and energetic equilibria of the cell, and dysfunctions in MAMs are implicated in many pathologies. One of the most important roles of the MAMs and an indicator of their functionality is the transfer of Ca2+ from the ER to the mitochondrion. However, measuring Ca2+ levels in subcellular compartments, despite the existence of many high fidelity, targetable dyes and indicators, is still limited to single-cell observations and not conducive to studying populations of cells. To address this limitation, a genetically encoded calcium indicator (GECI) called CaMPARI (Calmodulin modulatable photoactivatable ratiometric indicator) was characterized as an effective means of measuring intracellular Ca2+ in cell populations. CaMPARI was then used to measure intracellular both evoked and basal Ca2+ transfer dynamics in HEK293T, HeLa, and NSC34 cells differentially expressing the MAMs resident protein IP3R1 to model the Ca2+ dynamics of MAMs disturbance.

Intracellular membrane contact sites facilitate the transfer of ions, metabolites, and other molecules between subcellular compartments. Mitochondrial-associated membranes (MAMs) of the ER are particularly important for regulating the metabolic and energetic equilibria of the cell, and dysfunctions in MAMs are implicated in many pathologies. One of the most important roles of the MAMs and an indicator of their functionality is the transfer of Ca2+ from the ER to the mitochondrion. However, measuring Ca2+ levels in subcellular compartments, despite the existence of many high fidelity, targetable dyes and indicators, is still limited to single-cell observations and not conducive to studying populations of cells. To address this limitation, a genetically encoded calcium indicator (GECI) called CaMPARI (Calmodulin modulatable photoactivatable ratiometric indicator) was characterized as an effective means of measuring intracellular Ca2+ in cell populations. CaMPARI was then used to measure intracellular both evoked and basal Ca2+ transfer dynamics in HEK293T, HeLa, and NSC34 cells differentially expressing the MAMs resident protein IP3R1 to model the Ca2+ dynamics of MAMs disturbance.