Prolonged fasting alters skeletal muscle gene expression in a manner that promotes myofiber atrophy, but the underlying mechanisms are not fully understood. Here, we examined the potential role of activating transcription factor 4 (ATF4), a transcription factor with an evolutionarily ancient role in the cellular response to starvation. In mouse skeletal muscle, fasting increases the level of ATF4 mRNA. To determine whether increased ATF4 expression was required for myofiber atrophy, we reduced ATF4 expression with an inhibitory RNA targeting ATF4 and found that it reduced myofiber atrophy during fasting. Likewise, reducing the fasting level of ATF4 mRNA with a phosphorylation-resistant form of eukaryotic initiation factor 2alpha decreased myofiber atrophy. To determine whether ATF4 was sufficient to reduce myofiber size, we overexpressed ATF4 and found that it reduced myofiber size in the absence of fasting. In contrast, a transcriptionally inactive ATF4 construct did not reduce myofiber size, suggesting a requirement for ATF4-mediated transcriptional regulation. To begin to determine the mechanism of ATF4-mediated myofiber atrophy, we compared the effects of fasting and ATF4 overexpression on global skeletal muscle mRNA expression. Interestingly, expression of ATF4 increased a small subset of five fasting-responsive mRNAs, including four of the 15 mRNAs most highly induced by fasting. These five mRNAs encode proteins previously implicated in growth suppression (p21(Cip1/Waf1), GADD45alpha, and PW1/Peg3) or titin-based stress signaling [muscle LIM protein (MLP) and cardiac ankyrin repeat protein (CARP)]. Taken together, these data identify ATF4 as a novel mediator of skeletal myofiber atrophy during starvation.