TFEB is a critical transcription factor that regulates lysosomal capacity and the autophagy pathway [26, 60, 61]. Furthermore, in this study, we also observed that p-mTOR and p-ERK1/2 were regulated by ATO . SexSpecific Neurotoxicity of Dietary Advanced Glycation End Products in APP/PS1 Mice and Protective Roles of Trehalose by Inhibiting Tau Phosphorylation via GSK3TFEB Author: HuanHuan Zhou, Lan Luo, XueDi Zhai, Liangkai Chen, Guiping Wang, LiQiang Qin, Zengli Yu, LiLi Xin, Zhongxiao Wan Source: PARP and -tubulin were used as nuclear and . Interestingly, FCL also increased the nuclear translocation of transcription factor EB (TFEB), a master regulator of autophagic and lysosomal genes, and the mRNA expressions of TFEB-targeted genes, such as SQSTM1, MAP1LC3B, and UVRAG. TFEB Nuclear Translocation Assay. A novel mTORC1 effector implicated in lysosome biogenesis, endocytosis and autophagy. Similarly, treatment translocation.16 with the MTOR (mechanistic target of rapamycin [serine/thre- While the response of TFEB and TFE3 to nutrient status, onine kinase]) inhibitor Torin-1 in nutrient replete cells as well as their role in regulating the autophagy-lysosome induced a similar level of TFE3 nuclear translocation (Fig. Knockdown of TFEB by using small inference RNA In addition, Parkin, through its effects on PARIS, plays a crucial role in overall mitochondrial homeostasis via cellular regulation of the PGC-1-TFEB signaling pathway [ 34 ]. In contrast, while 47R-expressing cells have a mild basal elevation of nuclear TFEB, the nuclear translocation of TFEB in response to starvation is significantly . TFEB nuclear export is mediated by CRM1 and is dependent on phosphorylation. The PERK-dependent activation of TFEB contributes to cellular adaptation to stress by inducing autophagy genes 3.We . In addition, as a cellular phenotype, the number of lysosomes was increased by ten-fold in the Ragulator-deficient clone compared to that of control cells. Carbon monoxide-induced TFEB nuclear translocation enhances mitophagy/mitochondrial biogenesis in hepatocytes and ameliorates inflammatory liver injury Carbon monoxide (CO) can confer protection against cellular stress, whereas the potential involvement of autophagy and lysosomal biogenesis remains incompletely understood. Moreover, proteasome impairment not only promotes TFEB accumulation but also facilitates its dephosphorylation and nuclear translocation. (A) Resulting fractions were then detected with antibody against TFEB.Starvation (STV) was used as positive control. Therefore, we suggest that CO increases mitophagy through TFEB nuclear translocation by PERK-calcinuerin activation. When TFEB was knocked down, lysosomal biogenesis and autophagy were impaired in ATO-treated macrophages. Activity of TFEB is inhibited upon its serine phosphorylation by mTOR. PERK is required for TFEB nuclear translocation by SB202190. This can be followed by monitoring the changes in TFEB localization using widefield fluorescence microscopy. In contrast, while 47R-expressing cells have a mild basal elevation of nuclear TFEB, the nuclear translocation of TFEB in response to starvation is significantly . In control cells, TFEB:GFP is predominantly localized to the cytoplasm, whereas induction of autophagy by 3 hr starvation leads to robust nuclear translocation of TFEB (Figure 7A-B). Here we reported that the half-life of TFEB is around 13.5 h in neuronal-like cells, and TFEB is degraded through proteasome pathway in both neuronal-like and non-neuronal cells. The overall mechanisms by which TFEB activity in the cell is regulated are not well elucidated. Here, we describe steps to induce lysosomal damage in HeLa cells. Moreover, proteasome impairment not only promotes TFEB accumulation but also facilitates its dephosphorylation and nuclear translocation. Amino acid limitation that inactivates mTORC1 promotes de-phosphorylation and nuclear translocation of Transcription Factor EB (TFEB), a key transcriptional regulator of lysosome biogenesis and. Briefly, images were acquired randomly from at least ten different fields per sample. Representative images and quantification (A,B) of the TFEB-GFP translocation assay in HeLa cells stably expressing TFEB-GFP treated with 50, 100, and 150 M of genistein, in control conditions or treated with U18666A (U18) 0.5 g/mL for 24 h at the same time. Both the latter events were prevented by TFEB knockdown. During starvation the transcriptional activation of catabolic processes is induced by the nuclear translocation and consequent activation of transcription factor EB (TFEB), a master modulator of autophagy and lysosomal biogenesis. Here, we report that ATO triggered the nuclear translocation of TFEB, which in turn promoted autophagy and autophagosome-lysosome fusion. These findings indicate that mTORC1 essentially . Immunohistochemistry showing strong nuclear staining for TFEB is highly specific for t (6;11) Less common is fusion Xp11 of TFE3 and ASPL Immunohistochemistry showing strong nuclear staining with antibody to the C terminus of TFE3 is highly sensitive and specific for Xp11 Break apart FISH may be useful and less sensitive to fixation effects TFEB is also a target of the protein kinase AKT/PKB. Following lysosomal damage, activation and nuclear translocation of transcription factor EB (TFEB) is the key event to maintain lysosomal homeostasis. We observed that TFEB is mainly localized to the cytoplasm with focal concentration on lysosomes under basal cell growth conditions but that it translocates to the nucleus when lysosome function is inhibited. in TFEB phosphorylation was independent of mTORC1. (a) Phosphorylation is a well-recognized post-translational modification that regulates TFEB nuclear translocation. Author information Article notes . Interestingly, FCL also increased the nuclear translocation of transcription factor EB (TFEB), a master regulator of autophagic and lysosomal genes, and the mRNA expressions of TFEB-targeted genes, such as SQSTM1, MAP1LC3B, and UVRAG. TFEB is also a target of the protein kinase AKT/PKB. Conditions that promote TFEB nuclear translocation Initial evidence for the shuttling of TFEB between the cytoplasm and the nucleus was obtained in cells treated with sucrose, which is endocytosed and accumulated in the lysosomes due to their lack of invertase enzymes, and thus provides a cellular model of lysosome storage (Sardiello et al, 2009 ). GSK3 phosphorylates Ser-134 and Ser-138 and inhibits nuclear translocation of TFEB . Both the latter events were prevented by TFEB knockdown. We further found that during PIKfyve inhibition, the dephosphorylation of TFEB as well as its nuclear translocation is prevented by blocking protein phosphatase 2 A (PP2A) activity, either by knockdown or via the addition of okadaic acid, an inhibitor of PP2A. In this case, activation of TFEB and/or TFE3 appears to be mTOR-independent and is achieved through a protein kinase RNA-like endoplasmic reticulum kinase (PERK, also known as EIF2AK3)-dependent mechanism that induces activation of calcineurin and nuclear translocation of TFEB and/or TFE3 (Martina et al., 2016). Samuel Pea-Llopis and James Brugarolas. Figure 3. We found that laminar shear stress induced TFEB nuclear translocation, which induces TFEB transcription in an autoregulatory loop . 1. Genistein promotes transcription factor EB (TFEB) nuclear translocation to the nucleus in Niemann-Pick type C (NPC) cells. mTOR or mTORC2-GSK3 facilitates TFEB nuclear export. (A, B) SH-SY5Y cells were treated with SB202190 at the indicated concentrations (5, 10, and 20 M) for 4 h and subjected to nuclear and cytosolic fractionation. Pharmacological inhibition of AKT/PKB activates TFEB, promotes lysosome . CO increases TFEB nuclear translocation via the PERK-Ca 2+-calcineurin pathway. During starvation the transcriptional activation of catabolic processes is induced by the nuclear translocation and consequent activation of transcription factor EB (TFEB), a master modulator of autophagy and lysosomal biogenesis. Several kinases targeting TFEB have been identified. We identified 14-3-3 proteins as binding partners of TFEB that prevent its nuclear accumulation under conditions of lysosome sufficiency. Under cellular stress, such as low levels of ATP or glucose, mTORC1 is inhibited and TFEB then translocates to the nucleus to transcribe its target genes [ 20 ]. TFEB translocates to the nucleus and displays transcriptional activity in a PINK1-Parkin-dependent manner [ 33 ]. 4A), and a reduction in the diffuse cytoplasmic pool (Fig. Nutrient depletion induces TFEB dephosphorylation and subsequent nuclear translocation via the phosphatase calcineurin. In addition, trehalose, a known autophagy activator, was shown to inhibit Akt activity, thus promoting TFEB nuclear translocation. Similarly, treatment translocation.16 with the MTOR (mechanistic target of rapamycin [serine/thre- While the response of TFEB and TFE3 to nutrient status, onine kinase]) inhibitor Torin-1 in nutrient replete cells as well as their role in regulating the autophagy-lysosome induced a similar level of TFE3 nuclear translocation (Fig. Although the pathogenic factors are diverse, the underlying pathological features of sensorineural hearing loss 3A). Nuclear TFEB binds to the CLEAR element within the promoter region of LC3 and LAMP1 genes contributing to the upregulation of their transcription. TFEB nuclear export is mediated by CRM1 and is modulated by nutrient availability via mTOR-dependent hierarchical multisite phosphorylation of serines S142 and S138, which are localized in. Inhibition of mTOR or ERK, activation of PKC or activation of phosphatases calcineurin or PP2A induces TFEB nuclear translocation and enhances TFEB transcriptional activity. DAPI (blue) is included in the merge. Under basal conditions TFEB is found in the cytoplasm in an inactive, phosphorylated state (Settembre et al., 2011).However, under specific conditions, such as nutrient depletion or lysosomal stress (see below), TFEB moves to the nucleus, following dephosphorylation. Knock-in of nuclear translocation-deficient or inactive ACSS2 mutants in glioblastoma cells abrogates glucose . The ROS scavenger N-acetyl-1-cysteine (NAC) abolished ATO-induced nuclear translocation of TFEB, as well as changes in key molecules of the AKT/mTOR signaling pathway and downstream autophagy. In the nucleus, ACSS2 forms a complex with TFEB (transcription factor EB) and utilizes the acetate generated from histone deacetylation to locally produce acetyl-CoA for histone acetylation in the promoter regions of TFEB target genes. Furthermore, when TFEB is phosphorylated by mTOR complex 1 (mTORC1), it is retained in the cytoplasm ( 43 - 45 ). The mechanistic target of rapamycin (mTOR), as part of the mTORC1 complex, is a kinase that localizes to lysosomes. AKT/PKB phosphorylates TFEB at serine 467 and inhibits TFEB nuclear translocation. Nuclear transcription factor EB (TFEB) prevents atherosclerosis by activating macrophage autophagy and promoting lysosomal biogenesis. Nuclear transcription factor EB (TFEB) prevents atherosclerosis by activating macrophage autophagy and promoting lysosomal biogenesis. TFEB nuclear export is mediated by CRM1 and is dependent on phosphorylation. (A, B) SH-SY5Y cells were treated with SB202190 at the indicated concentrations (5, 10, and 20 M) for 4 h and subjected to nuclear and cytosolic fractionation. Accordingly, inhibition of GSK3 results in TFEB nuclear localization and autophagy induction (Marchand et . Once in the nucleus, TFEB is responsible for the transcription of autophagy-related genes and genes encoding lysosomal proteins [ 19 ]. 1C), system . However, how TFEB is inactivated upon nutrient refeeding is currently Figure 3. However, the exact role of PERK . PERK is required for TFEB nuclear translocation by SB202190. Conversely, activation of Here, we report that ATO triggered the nuclear translocation of TFEB, which in turn promoted autophagy and autophagosome-lysosome fusion. A recent study showed that TFEB is phosphorylated by AKT at serine residue S467 and that treating cells with an AKT inhibitor promotes TFEB nuclear translocation (Palmieri et al, 2017). PARP and -tubulin were used as nuclear and . The common pathogenic mutant, LRRK2G2019S, appears to hijack this pathway. TFEB bound to PD-L1 promoter in RCCs and inhibition of mTOR led to enhanced TFEB nuclear translocation and PD-L1 expression. Recently, dysfunctional TFEB nuclear translocation was reported to induce deficient lysosomal capacity and autophagy arrest, which are associated with the development of HD, PD, AD and other neurodegenerative diseases [27, 62, 63]. Starvation is known to induce calcineurin-mediated TFEB dephosphorylation and nuclear translocation 22, however how nutrient refeeding induces TFEB inactivation is currently unknown. Lysosomal stress induces TFEB nuclear translocation. In addition, the inhibition of TFEB with siRNA against TFEB abrogated the increase of mtDNA with CO, markers of mitochondrial biogenesis such as PGC1, NRF1, and TFAM, and the mitochondrial proteins COX II, COX IV, and cytochrome c. TFEB contains a nuclear export signal (NES), whose phosphorylation on nearby serines (S122, S134, S138, and S142) is a major determinant of cytoplasmic-nuclear shuttling . spiral ganglion neuron; TFEB Introduction Sensorineural hearing loss is a common sensory disease that seriously affects physical and mental health and quality of life in humans. 1C), system . Although phosphorylation plays a role in regulating the nuclear abundance of TFEB ( 11, 13 ), the cellular mechanisms that sense lysosomal status and transduce the signals that regulate TFEB localization remain unclear. Here we reported that the half-life of TFEB is around 13.5 h in neuronal-like cells, and TFEB is degraded through proteasome pathway in both neuronal-like and non-neuronal cells. Furthermore, FCL decreased the activities of cathepsin B and cathepsin D and affected the cellular acidification. of CD38, both by stabilizing TFEB and promoting its nuclear translocation via aberrant calcium signaling. ARTICLE HISTORY Received 3 June 2020 Next, in the presence of NAC, ATO failed to promote TFEB nuclear translocation, as well as to upregulate autophagy- and lysosome-related proteins.