polyP detection and manipulation in rabbit heart mitochondria and cardiomyocytes. (A) Average amount of polyP in rabbit heart mitochondria (left) and gel images of polyP standard and polyP sample from isolated rabbit mitochondria (right). (B) Images of control GFP (left) and GFP-PPX–expressing cells (right). The top panel shows global GFP fluorescence at 500–530 nm that reveals the mitochondrial fluorescence pattern in PPX-expressing cells and a homogeneously distributed fluorescence in control cells. The middle panel shows the decrease in DAPI fluorescence in polyP-depleted cells. The bottom panel shows colocalization of GFP-PPX signal with mitochondria. TMRM was used as a mitochondrial signal, and the degree of overlay is presented in shades of yellow in the merged image. (C) Fluorescence spectrum of 5 µM of DAPI-loaded myocytes expressing control GFP (black), PPX (red), and control GFP cells not loaded with DAPI (gray). (D) Mean values from the DAPI spectrum obtained in C for GFP (black)- and PPX (red)-expressing myocytes at λem = 584 nm. a.u., arbitrary fluorescence units. The numbers in parenthesis indicate the number of hearts (A) or number of cells (D).
