The cellular protein degradation system, such as proteasomal or autophagy-lysosomal system plays an important role in the pathogenesis of a variety of human diseases including cancer. Transcription factor EB (TFEB) is a master transcriptional factor in the regulation of autophagy-lysosome pathway (ALP), and it has multiple biological functions including protein degradation, cell homeostasis and cell survival. In the present study we show that the tumor suppressor p53 can regulate TFEB nuclear translocation and activity in lung cancer cells. We found p53 deletion or chemical inhibition of p53 using pifithrin-α could promote the translocation of TFEB from cytoplasm to the nucleus, thus increased the TFEB-mediated lysosomal and autophagosomal biogenesis in lung cancer cells. Moreover, re-expression of p53 could decrease the expression levels of TFEB-targeting genes involved in ALP, and knockdown of TFEB could abolish the effect of p53 on the regulation of ALP gene expression. Taken together, our data indicate that p53 affects ALP through regulating TFEB nuclear translocation in lung cancer cells. Importantly, our study reveals a critical link between two keys factors in tumourigenesis and autophagy, and suggests a potential important role of p53-TFEB signaling axis in lung cancer.