plasmid expressing tfeb-gfp pegfp-n1-tfeb  (Addgene inc)

Journal: Genes to cells : devoted to molecular & cellular mechanisms

Article Title: MTMR4, a phosphoinositide-specific 3'-phosphatase, regulates TFEB activity and the endocytic and autophagic pathways.

doi: 10.1111/gtc.12609

Figure Lengend Snippet: FIGURE 6 MTMR4 knockdown inhibits starvation-induced increase in dephosphorylated TFEB/total TFEB ratio. (a) Phosphorylation of TFEB and other proteins and total amounts of MTMR4 and MTMR3 in control and MTMR4-depleted cells. A549 cells transfected with control- or MTMR4-siRNAs were placed in growth medium (fed) with or without 100 μM chloroquine or in HBSS for 4 hr, and subjected to western blot analyses. S6K, S6 kinase. (b) Quantified data of total TFEB protein level (sum of the dephosphorylated and phosphorylated forms of TFEB) (left), phospho-TFEB level (normalized to GAPDH) (middle) and ratio of dephosphorylated TFEB/total TFEB (right) in control and MTMR4–depleted cells. (c) Time course of starvation-induced change in phosphorylation of TFEB and S6K. A549 cells were starved by culture in HBSS for the indicated time periods and subjected to western blot analyses. Stv, starved. (d) Phosphorylation of TFEB in control and MTMR4-depleted cells under both fed and starved conditions with or without cyclosporine A treatment. Control and MTMR4-depleted A549 cells were incubated in growth medium (fed) or HBSS (starved) with or without cyclosporine-A (10 μM) for 4 hr

Article Snippet: GFP- TFEB expression vector (# 38119) was provided by Dr. Shawn Ferguson through Addgene (Roczniak- Ferguson et al., 2012).

Techniques: Knockdown, Phospho-proteomics, Control, Transfection, Western Blot, Incubation

Journal: Genes to cells : devoted to molecular & cellular mechanisms

Article Title: MTMR4, a phosphoinositide-specific 3'-phosphatase, regulates TFEB activity and the endocytic and autophagic pathways.

doi: 10.1111/gtc.12609

Figure Lengend Snippet: FIGURE 8 PI(3)P-phosphatase MTMR4 regulates endocytic and autophagic pathways. MTMR4 is localized mainly in LEs and APs to regulate PI(3)P levels. MTMR4 regulates the fusion, fission, motility and maturation of endosomes, the autophagic pathway, and lysosomal biogenesis. MTMR4 is also required for activation of TFEB. This action of MTMR4 contributes to MTMR4 regulation of the endocytic and autophagic pathways

Article Snippet: GFP- TFEB expression vector (# 38119) was provided by Dr. Shawn Ferguson through Addgene (Roczniak- Ferguson et al., 2012).

Techniques: Activation Assay