Abstract
Autophagy and lysosomal activities play a key role in the cell by initiating and carrying out the degradation of misfolded proteins. Transcription factor EB (TFEB) functions as a master controller of lysosomal biogenesis and function during lysosomal stress, controlling most but, importantly, not all lysosomal genes. Here, we sought to better understand the regulation of lysosomal genes whose expression does not appear to be controlled by TFEB. Sixteen of these genes were screened for transactivation in response to diverse cellular insults. mRNA levels for lysosomal-associated membrane protein 3 (LAMP3), a gene that is highly up-regulated in many forms of cancer, including breast and cervical cancers, were significantly increased during the integrated stress response, which occurs in eukaryotic cells in response to accumulation of unfolded and misfolded proteins. Of note, results from siRNA-mediated knockdown of activating transcription factor 4 (ATF4) and overexpression of exogenous ATF4 cDNA indicated that ATF4 up-regulates LAMP3 mRNA levels. Finally, ChIP assays verified an ATF4-binding site in the LAMP3 gene promoter, and a dual-luciferase assay confirmed that this ATF4-binding site is indeed required for transcriptional up-regulation of LAMP3 These results reveal that ATF4 directly regulates LAMP3, representing the first identification of a gene for a lysosomal component whose expression is directly controlled by ATF4. This finding may provide a key link between stresses such as accumulation of unfolded proteins and modulation of autophagy, which removes them.