We have demonstrated previously that the STAT‐1 transcription factor plays a key role in ischemia/reperfusion (I/R)‐induced apoptosis in cardiac myocytes. In the present study we assessed which region of the STAT‐1 molecule mediates apoptosis in cardiac myocytes. A STAT‐1 construct (amino acid 350–750) lacking the N‐terminus could enhance I/R‐induced apoptosis in cardiac myocytes. However, a STAT‐1 construct, which lacks 60 amino acids at the C‐terminus (amino acid 691–750), was ineffective in promoting I/R‐induced apoptosis in cardiac myocytes. Furthermore, overexpression of a C‐terminal STAT‐1 construct (amino acid 691–750) containing the transcriptional activation domain, but not the DNA binding domain, strongly enhanced I/R‐induced apoptotic cell death. Cardiac myocytes isolated from mice expressing a truncated C‐terminal STAT‐1 were more sensitive to I/R‐induced cell death. Finally, isolated hearts from these animals exposed to I/R injury had larger infarct size and greater number of TUNEL‐positive myocytes than control hearts. These studies demonstrate that the C‐terminal transactivation domain of STAT‐1 is necessary and sufficient for I/R injury‐induced apoptosis in cardiac myocytes.