ATG5 and ATG7 are considered as essential molecules for induction of macroautophagy. However, we found that cells lacking ATG5 or ATG7 can still form autophagosomes/autolysosomes and perform autophagy-mediated protein degradation when subjected to certain stresses. Although lipidation of LC3 is accepted to be a good indicator of macroautophagy, it did not occur during the ATG5/ATG7-independent alternative macroautophagy. Unlike conventional macroautophagy, autophagosomes seemed to be generated in a Rab9-dependent manner by the fusion of the phagophore with vesicles derived from the trans-Golgi and late endosomes. Mammalian macroautophagy can occur via at least two different pathways, which are an ATG5/ATG7-dependent conventional pathway and an ATG5/ATG7-independent alternative pathway.