Autophagy is a lysosomal degradation pathway for bulk cytosolic proteins and damaged organelles, and is well known to act as a cell survival mechanism. Acetaminophen (APAP) overdose can cause liver injury in animals and humans by inducing necrosis due to mitochondrial damage. We recently found that pharmacological induction of autophagy by rapamycin protects against, whereas pharmacological suppression of autophagy by chloroquine exacerbates, APAP-induced liver injury in mice. Autophagy is induced to remove APAP-induced damaged mitochondria and thus attenuates APAP-induced hepatocyte necrosis. To our surprise, we found that liver-specific Atg5 knockout mice are not more susceptible, but are resistant to APAP-induced liver injury due to compensatory effects. Our work suggests that pharmacological modulation of autophagy is a novel therapeutic approach to ameliorate APAP-induced liver injury. Moreover, our work also suggests that caution needs to be exercised when using genetic autophagy gene knockout mice for pathophysiological studies.