Cardiac differentiation is associated with enhancement and restructuring of the adenylate kinase isoform network.

Abstract

<p><b>A</b> - Microarray analyses of total mRNA in embryonic stem (ES) cells and derived cardiomyocytes (CM). Adenylate kinase isoform genes are hierarchically clustered as mRNA copy numbers of CM <i>versus</i> ES transcripts with extent of fold change color-coded (n = 3 in each group). <b>B</b> – ES cells have lower total adenylate kinase activity than derived CM. <b>C</b> – CM have more AK1 transcripts and protein than ES cells. <b>D and E</b> - CM transcribe AK2 less and AK5 more frequently than ES cells but protein levels of both isoforms are approximately equal. <b>F</b> and <b>G</b> - Confocal immunocytochemistry indicate cytosolic and limited nuclear localization of AK1 (green) in ES cells (nuclear staining with DAPI, blue) and AK1 association with the cell membrane (G). <b>H</b> – CM have greater AK1 levels concentrated in the nucleus, perinuclear space and along myofibrils. <b>I</b> – In ES cell-derived CM, AK1 is present in intercellular nanotubular connections and nucleus (inset). <b>J</b> and <b>K</b> - AK2 is found in the cytoplasm both in ES cells and CM; higher levels of AK2 in CM formed a punctate pattern within the cytoplasm consistent with mitochondrial localization. <b>L</b> and <b>M</b> - The cytoplasmic localization of the AK5 isoform did vary between ES cells and ES-derived CM; AK5 protein was low in CM. <b>F</b> to <b>M</b> – representative images of n = 5–10. All scale bars are 10 µm. * <i>P</i><0.05.</p