Figure 2.

TFEB serine 122 is regulated by MTORC1. (A) Validation of the p-TFEB (S122) antibody. HeLa cells were transfected with the indicated MYC-tagged TFEB constructs followed by MYC-IP and western blot. λ, lambda-phosphatase treatment of immunoprecipitates. HeLa cells were treated with Torin1 (250 nM) (B), or starved for amino acids (C), serum (D), or glucose (E). (F, G) HeLa cells were transfected with active (RRAGB^GTP/D^GDP) or inactive (RRAGB^GDP/D^GTP) RRAG-GTPases and its effects on TFEB localization and phosphorylation were assessed. (H) MTOR in vitro kinase assay of epitope tag immunoprecipitates from cells transfected with empty vector (EV) vs. MYC-TFEB and incubated with or without recombinant MTOR. Scale bar: 10 μm.