Doxorubicin, a cardiotoxic antineoplastic, disrupts the cardiac-specific program of gene expression (Kurabayashi, M., Dutta, S., Jeyaseelan, R., and Kedes, L. (1995) Mol. Cell. Biol. 15, 6386-6397). We have now identified neonatal rat cardiomyocyte mRNAs rapidly sensitive to doxorubicin, or its congener daunomycin, including transcripts of nuclear genes encoding enzymes critical in production of energy in cardiomyocytes: ADP/ATP translocase, a heart- and muscle-specific isoform; Reiske iron-sulfur protein (RISP), a ubiquitously expressed electron transport chain component; and a muscle isozyme of phosphofructokinase. Loss of these mRNAs following doxorubicin or daunomycin is evident as early as 2 h and precedes significant reduction of intracellular ATP. ATP levels in control cardiomyocytes (17.9 +/- 2.9 nM/mg of protein) fall only after 14 h and reach residual levels of 10.4 +/- 0.9 nM (doxorubicin; p = <0.006) and 6.7 +/- 1.9 nM (daunomycin; p = <0. 001) by 24 h. Loss of mRNAs generating ATP was highly selective since mRNAs for other energy production enzymes, (cytochrome c, cytochrome b, and malate dehydrogenase), and genes important in glycolysis (pyruvate kinase and glyceraldehyde-3-phosphate dehydrogenase) were unaffected even at 24 and 48 h. The drugs had no effect on levels of ubiquitously expressed RISP mRNA in fibroblasts. These findings could link doxorubicin-induced damage to membranes and signaling pathways with 1) suppression of transcripts encoding myofibrillar proteins and proteins of energy production pathways and 2) depletion of intracellular ATP stores, myofibrillar degeneration, and related cardiotoxic effects.