Clinical Relevance of Ubiquitin-Proteasome System Profiling in Acute Leukemias.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2633-2633
Author(s):  
Wanlong Ma ◽  
Hagop M. Kantarjian ◽  
XI Zhang ◽  
Xiuqiang Wang ◽  
Zeev Estrov ◽  
...  
Keyword(s):  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Ubiquitin Proteasome System ◽  
Enzymatic Activities ◽  
Acute Leukemias ◽  
Clinical Behavior ◽  
Protein Levels ◽  
Control Subjects ◽  
Ubiquitin Proteasome ◽  
Proteasome Protein

Abstract Abstract 2633 Poster Board II-609 The ubiquitin-proteasome system (UPS) plays a major role in the homeostasis of cellular proteins: chymotrypsin-like (Ch-L), caspase-like (Cas-L), and trypsin-like (Tr-L). The UPS system is involved in most critical cellular processes and its role in oncogenesis is well established. Here we compared UPS protein levels (proteasome and ubiquitin) and proteasome enzymatic activities in peripheral blood plasma from newly diagnosed patients with acute myeloid leukemia (AML; n=147), acute lymphoblastic leukemia (ALL; n=34?), or myelodysplastic syndrome (MDS ; n=27), and 99 apparently healthy control subjects. Proteasome and ubiquitin were measured using immunoassays based on electro-chemiluminescence technology. The proteasome enzymatic activities were measure using a standard enzymatic assay utilizing fluorogenic peptide-AMC substrate. By normalizing these enzymatic activities to the levels of proteasome protein in plasma, we also determined specific proteasome enzyme activities (Ch-L/p, Cas-L/p, and Tr-L, respectively). AML, ALL, and MDS patients all had high levels of proteasome protein, ubiquitin, and enzymatic activities relative to control subjects. However, each specific enzyme activity in general was lower in all 3 disease groups than in control subjects, which suggests that each proteasome has lower enzymatic activity in blast cells but we cannot rule out that the number of proteasomes might be increased. Despite these similarities, AML, ALL, and MDS each exhibited a specific profile of UPS protein and enzymatic activities . Proteasome protein levels and enzymatic activities also correlated with clinical behavior. In AML, proteasome protein level was a strong predictor of survival as a both a continuous (P<0.00001) and a dichotomous (P=0.04) variable, independent of cytogenetics, performance status, and age. In ALL, Ch-L/p showed a significant negative correlation with survival (P=0.0015). In conclusion, these data demonstrate that each disease has a unique UPS profile and confirm that the UPS system plays a major role in the leukemic process. In addition, profiling the UPS in leukemias using plasma provides valuable biomarkers that can be used to help predict clinical behavior and may ultimately help manage disease. Disclosures: No relevant conflicts of interest to declare.

Use of Ubiquitin-Proteasome System Profiling for Differentiating Between Various Leukemic Processes.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4712-4712
Author(s):  
Ke Zhang ◽  
Hagop M. Kantarjian ◽  
Wanlong Ma ◽  
XI Zhang ◽  
Xiuqiang Wang ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Blast Crisis ◽  
Chronic Phase ◽  
Direct Analysis ◽  
Cell Types ◽  
Ubiquitin Proteasome System ◽  
Lymphocytic Leukemia ◽  
Ubiquitin Proteasome

Abstract Abstract 4712 The ubiquitin-proteasome system (UPS) plays a major role in cell homeostasis in normal and neoplastic states. Expression and function of the UPS system vary with the specific characteristics of individual cell types, suggesting that determination of UPS “signatures” could be useful in identifying various cell populations. Since direct analysis of cancer cells is often problematic, even in hematologic diseases, we explored the potential of using UPS signatures in plasma to differentiate between various leukemias. We first analyzed plasma UPS profiles of patients with acute myeloid leukemia (AML; n=111), acute lymphoblastic leukemia (ALL; n=29), advanced myelodysplastic syndrome (MDS; n=20), chronic lymphocytic leukemia (CLL; n=118), or chronic myeloid leukemia (CML; n=128; 46 in accelerated/blast crisis [ACC/BL], 82 in chronic phase), and 85 healthy control subjects. Plasma levels of proteasome, ubiquitin (poly-ubiquitin), and the 3 proteasome enzymatic activities (chymotrypsin-like [Ch-L], caspase-like [Cas-L], trypsin-like [Tr-L]) were measured. Specific activities were calculated by normalizing each of the 3 enzyme activities to the levels of proteasome protein in plasma (Ch-L/p, Cas-L/p, and Tr-L/p). These 8 variables were used in multivariate logistic regression models to differentiate between leukemic processes. UPS signatures provided clear differentiation between patients with a leukemic process and normal controls (AUC=0.991), using 6 different variables (Tr-L/P, Ch-L, Ch-L/p, Cas-L, Cas-L/P, ubiquitin). Distinguishing between acute (AML, ALL, MDS) and chronic (CML, CLL) processes was less efficient (AUC=0.853 using Tr-L, Tr-L/P, Cas-L/P, Ch-L/P, proteasome, Ch-L), likely due to the high proportion (36%) of CML patients in ACC/BL phase. However, UPS signatures generally yielded powerful differentiation between individual leukemias (Table). MDS was not well differentiated from AML (AUC=0.791), reflecting the significant biological overlap of these diseases. These data support the potential usefulness of the UPS profile to aid in the differential diagnosis of various leukemias. Disclosures: No relevant conflicts of interest to declare.

Gfi1 ubiquitination and proteasomal degradation is inhibited by the ubiquitin ligase Triad1

Blood ◽  
2007 ◽  
Vol 110 (9) ◽  
pp. 3128-3135 ◽  
Author(s):  
Jurgen A. F. Marteijn ◽  
Laurens T. van der Meer ◽  
Liesbeth van Emst ◽  
Simon van Reijmersdal ◽  
Willemijn Wissink ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Ubiquitin Ligase ◽  
Ubiquitin Proteasome System ◽  
Proteasomal Degradation ◽  
Binding Domain ◽  
U937 Cells ◽  
Dna Binding Domain ◽  
Protein Levels ◽  
Ubiquitin Ligase Activity ◽  
Ubiquitin Proteasome

Abstract Growth factor independence 1 (Gfi1) is a transcriptional repressor essential for the function and development of many different hematopoietic lineages. The Gfi1 protein expression is regulated by the ubiquitin-proteasome system. In granulocytes, Gfi1 is rapidly degraded by the proteasome, while it is more stable in monocytes. How the ubiquitination and degradation of Gfi1 is regulated is unclear. Here, we show that the ubiquitin ligase Triad1 interacts with the DNA-binding domain of Gfi1. Unexpectedly, we found that Triad1 inhibited Gfi1 ubiquitination, resulting in a prolonged half-life. Down-regulation of endogenous Triad1 by siRNAs resulted in increased Gfi1 ubiquitination. In U937 cells, Triad1 caused an increase in endogenous Gfi1 protein levels and slowed cell proliferation in a similar manner when Gfi1 itself was expressed. A Triad1 mutant that lacks the Gfi1-binding domain did not affect Gfi1 levels and proliferation. Because neither proteasome-ubiquitin nor Triad1 ubiquitin ligase activity was required for the inhibition of Gfi1 ubiquitination, these data suggest that Triad1 competes for Gfi1 binding with as yet to be identified E3 ubiquitin ligases that do mark Gfi1 for proteasomal degradation. The finetuning of Gfi1 protein levels regulated by Triad1 defines an unexpected role for this protein in hematopoiesis.

The Serum Fas and Fas Ligand levels in Childhood Acute Leukemias.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4724-4724
Author(s):  
Alev Kiziltas ◽  
Bulent Antmen ◽  
Ilgen Sasmaz ◽  
Yurdanur Kilinc ◽  
Mustafa Yilmaz ◽  
...  
Keyword(s):  
Fas Ligand ◽  
Conflicts Of Interest ◽  
Cytotoxic T Cells ◽  
Lymphoblastic Leukemia ◽  
Serum Levels ◽  
Control Group ◽  
Extrinsic Pathway ◽  
Acute Leukemias ◽  
Mean Values ◽  
The Mean

Abstract Abstract 4724 Aim Abnormalities and alterations in apoptosis mechanism may lead to cancer development. Cystean proteases enzymes, called caspases, appear to be involved in both the initial signaling events. There are many proteins that trigger intrinsic and extrinsic pathway and induce apoptosis signals. Fas and its specific ligand that known as Fas Ligand are the best defined dead receptors and have functions in apoptosis regulation with many tumor types. Fas binds the ligand on the cytotoxic T cells and start apoptosis. Objectives of this study were to determine serum levels of Fas and Fas Ligand at the time of diagnosis in childhood acute leukemias that may be play important role in apoptosis mechanism. Patients and Methods In this study, we investigated serum Fas and Fas Ligand levels by using ELISA method in childhood acute leukemias. Twenty-nine cases with acute lymphoblastic leukemia and twenty-three cases with acute myeloblastic leukemia at the ages of 1-18 years are included this study. The age distrubition of the control group varied 1-15 years consisted of twenty-seven children. We investigated serum Fas and Fas Ligand levels at the time of diagnosis from peripheral blood samples. Results The comparison of the mean values of Fas and Fas Ligand levels in acute leukemia patients groups and control group have shown important difference as statistically (p<0,05). The mean values of Fas and Fas Ligand levels were higher in ALL and AML patients. The comparison of the mean values of Fas and Fas ligand levels in ALL and AML patients have shown no difference (p>0,05). The comparison of the Fas levels in ALL patients according to immunophenotypes; CALLA(+) B-ALL have higher mean level than T-ALL and shown important difference as statistically (p<0,05). The comparison of the mean values of Fas level at the diagnosis in ALL patients who had relapsed and patients who had remission have shown important difference (p<0,05). The mean values of Fas level were found higher in relapsed ALL patients. In these results showed that Fas and Fas ligand may play important role in apoptosis mechanism. Disclosures: No relevant conflicts of interest to declare.

Inherited and Acquired RUNX1 Mutations in Hematologic Disorders

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. SCI-5-SCI-5
Author(s):  
Nancy A. Speck ◽  
Xiongwei Cai ◽  
Jingping Ge ◽  
Philip J. Mason
Keyword(s):  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Conflicts Of Interest ◽  
P53 Protein ◽  
Lymphoblastic Leukemia ◽  
Genotoxic Stress ◽  
Myelogenous Leukemia ◽  
Phenotypic Properties ◽  
Hematopoietic Stem ◽  
Protein Levels

Abstract RUNX1, a DNA binding subunit of core binding factors, is frequently mutated or rearranged in hematopoietic malignancies, including acute myelogenous leukemia (AML), chronic myelomonocytic leukemia, acute lymphoblastic leukemia, and myelodysplastic syndrome (MDS). Mutations in RUNX1 can be early events in leukemia, and generate a long-lived pre-leukemic stem cell (pre-LSC). Additionally, it has been reported that loss of function RUNX1 mutations are particularly frequent in radiation-associated MDS and AML, suggesting that pre-existing RUNX1 mutations in a pre-LSC may predispose patients to MDS/AML following DNA damage. Discussion will focus on the phenotypic properties of Runx1-deficient pre-LSCs, and the mechanisms by which Runx1 deficiency contributes to these phenotypes. Pan-hematopoietic Runx1 loss in mice causes a G1 to S-cell cycle delay and decreases apoptosis of pre-LSCs. Runx1-deficient pre-LSCs are radiation- and chemotherapy-resistant, and this correlates with decreased p53 protein levels and an attenuated p53 pathway response. Both p53 protein levels and apoptosis are increased following treatment with Nutlin-3. Runx1-deficient pre-LSCs are smaller, consume less glucose, and produce less ATP than normal hematopoietic stem cells (HSCs). Runx1-deficient stem and progenitor cells have lower overall ribosomal content and skewing in the relative amounts of rRNA and mRNA encoding ribosomal proteins. Analysis of AKT pathway components suggests that the decreased ribosome biogenesis is unlikely to be primarily caused by lower AKT signaling. We hypothesize that one or more of the above-mentioned properties (low p53 levels, decreased metabolism) render Runx1-deficient pre-LSCs less sensitive to genotoxic stress than normal HSCs, allowing a Runx1-deficient pre-LSC population to both perdure and expand in the bone marrow. Disclosures: No relevant conflicts of interest to declare.

Diminished proteasomal degradation results in accumulation of Gfi1 protein in monocytes

Blood ◽  
2006 ◽  
Vol 109 (1) ◽  
pp. 100-108 ◽  
Author(s):  
Jurgen A. F. Marteijn ◽  
Laurens T. van der Meer ◽  
Liesbeth Van Emst ◽  
Theo de Witte ◽  
Joop H. Jansen ◽  
...  
Keyword(s):  
Ubiquitin Proteasome System ◽  
Proteasomal Degradation ◽  
Myeloid Differentiation ◽  
Mrna Levels ◽  
Monocytic Differentiation ◽  
Granulocytic Differentiation ◽  
Monocytic Cell ◽  
Protein Levels ◽  
Ubiquitin Proteasome ◽  
Monocytic Lineage

Abstract Gfi1 is a transcriptional repressor essential during myeloid differentiation. Gfi1−/− mice exhibit a block in myeloid differentiation resulting in the accumulation of an immature myelo-monocytic cell population and the complete absence of mature neutrophils. Even though mRNA levels of Gfi1 appear to be very low in monocytes, Gfi1 might play a role in the monocytic lineage as Gfi1−/− mice exhibit diminished monocyte-derived dendritic cells and disturbed cytokine production by macrophages in response to LPS. We show here that Gfi1 protein levels are mainly regulated by the ubiquitin-proteasome system. Upon forced monocytic differentiation of U937 cells, Gfi1 mRNA levels dropped but protein levels increased due to diminished proteasomal turnover. Similarly, Gfi1 mRNA levels are low in primary monocytes whereas the protein is clearly detectable. Conversely, Gfi1 mRNA levels are high in granulocytes but the protein is swiftly degraded by the proteasome in these cells. Chromatin immunoprecipitation experiments showed that Gfi1 binds to the promoter of several granulocyte-specific genes in primary monocytes, including C/EBPα, neutrophil elastase, and Gfi1 itself. The binding of the repressor Gfi1 to these promoters correlated with low expression of these genes in monocytes compared with granulocytes. Our data fit a model in which Gfi1 protein levels are induced in primary monocytes, due to diminished proteasomal degradation, to repress genes that play a role in granulocytic differentiation.

scholarly journals Gfi1 Upregulates c-Myc Expression and Promotes c-Myc-Driven Cell Proliferation

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3769-3769
Author(s):  
Yangyang Zhang ◽  
Fan Dong
Keyword(s):  
Cell Proliferation ◽  
Transcription Factor ◽  
Transcriptional Repression ◽  
Cell Cycle Progression ◽  
Target Genes ◽  
Bone Marrow Cells ◽  
Conflicts Of Interest ◽  
Protein Levels ◽  
Enhanced Expression ◽  
Ubiquitin Proteasome

Gfi1 is a zinc-finger transcriptional repressor that plays an important role in hematopoiesis. When aberrantly activated, Gfi1 may function as a weak oncoprotein in the lymphoid system, but collaborate strongly with c-Myc in lymphomagenesis. c-Myc is a transcription factor that is frequently activated in human cancers including leukemia and lymphoma mainly due to its overexpression as a result of gene amplifications and chromosomal translocations. c-Myc overexpression may also result from stabilization of c-Myc protein, which is highly unstable and rapidly degraded through the ubiquitin-proteasome pathway. The mechanism by which Gfi1 collaborates with c-Myc in lymphomagenesis is incompletely understood. c-Myc activates gene expression by forming a heterodimeric complex with the partner protein Max, but may also repress target genes through interaction with transcription factor Miz-1. We previously showed that Gfi1 indirectly interacts with c-Myc through Miz-1 and collaborates with c-Myc to repress CDK inhibitors p21Cip1 and p15Ink4B. In this study, we show that Gfi1 augmented the level of c-Myc protein transiently expressed in Hela cells and the levels of MycER fusion protein stably expressed in the mouse pro-B Ba/F3 and myeloid 32D cells. The C-terminal ZF domains of Gfi1, but not its transcriptional repression and DNA binding activities, were required for c-Myc upregulation. Notably, although Miz-1 has been shown to stabilize c-Myc protein, the expression of c-Myc V394D mutant, which is defective in Miz-1 interaction, was still upregulated by Gfi1, suggesting that Gfi1-mediated c-Myc upregulation was independent of Miz-1 interaction. We further show that Gfi1 overexpression led to reduced polyubiquitination and increased stability of c-Myc protein. Interestingly, the levels of endogenous c-Myc mRNA and protein were augmented upon induction of Gfi1 expression in Ba/F3 and Burkitt lymphoma Ramos cells transduced with the doxycycline-inducible Gfi1 lentiviral construct, but reduced in Gfi1-knocked down human leukemic HL60 and U937 cells. Additionally, targeted deletion of Gfi1 resulted in reduced c-Myc expression in mouse lineage negative bone marrow cells, which was associated with a decline in the expression of c-Myc-activated target genes. The oncogenic potential of Myc derives from its ability to stimulate cell proliferation. Our results demonstrate that inducible expression of Gfi1 in Ba/F3 cells expressing MycER promoted Myc-driven cell cycle progression and proliferation. Thus, in addition to its role in c-Myc-mediated transcriptional repression, Gfi1 upregulates c-Myc expression at both mRNA and protein levels, leading to enhanced expression of c-Myc-activated genes and augmented cell proliferation driven by c-Myc. Together, these data may reveal a novel mechanism by which Gfi1 collaborates with c-Myc in lymphomagenesis. Disclosures No relevant conflicts of interest to declare.

Recent Advances in the Discovery of Novel Peptide Inhibitors Targeting 26S Proteasome

2019 ◽  
Vol 18 (12) ◽  
pp. 1656-1673
Author(s):  
Xinjie Gu ◽  
Shutao Ma
Keyword(s):  
Cancer Therapy ◽  
Proteasome Inhibitors ◽  
Ubiquitin Proteasome System ◽  
Peptide Inhibitors ◽  
26S Proteasome ◽  
Design Strategies ◽  
Intracellular Protein ◽  
Primary Tumors ◽  
Protein Levels ◽  
Ubiquitin Proteasome

Background: The 26S proteasome is a proteolytic complex of multimeric protease, which operates at the executive end of the Ubiquitin-Proteasome System (UPS) and degrades the polyubiquitylated proteins. Methods: After a brief introduction of 26S proteasome and Ubiquitin-Proteasome System (UPS), this review focuses on the structure and function of the 26S proteasome in intracellular protein level regulation. Then, physiological regulation mechanisms and processes are elaborated. In addition, the advantages and defects of approved 26S proteasome inhibitors were discussed. Finally, we summarized the novel peptide 26S proteasome inhibitors according to their structural classifications, highlighting their design strategies, inhibitory activity and Structure-Activity Relationships (SARs). Results: Cellular function maintenance relies on the proteasome metabolizing intracellular proteins to control intracellular protein levels, which is especially important for cancer cells to survive and proliferate. In primary tumors, proteasomes had a higher level and more potent activity. Currently, the approved small peptide inhibitors have proved their specific 26S proteasome inhibitory effects and considerable antitumor activities, but with obvious defects. Increasingly, novel peptide inhibitors are emerging and possess promising values in cancer therapy. Conclusion: Overall, the 26S proteasome is an efficient therapeutic target and novel 26S proteasome inhibitors hold potency for cancer therapy.

Flow Cytometric Leukemic Blasts Detection in Cerebrospinal Fuid of Children with Acute Leukemias

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1449-1449
Author(s):  
Alexander Popov ◽  
Tatiana Verzhbitskaya ◽  
Grigory Tsaur ◽  
Alexander Tomilov ◽  
Olga Khlebnikova ◽  
...  
Keyword(s):  
Risk Factors ◽  
Flow Cytometry ◽  
Tumor Cells ◽  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Prognostic Significance ◽  
Low Flow ◽  
Prognostic Impact ◽  
Acute Leukemias ◽  
Childhood Acute Leukemia

Abstract Abstract 1449 Central nervous system (CNS) involvement is one of the important risk factors in childhood acute leukemia (AL). Tumor cells detection in cerebrospinal fluid (CSF) is one of the main signs of CNS lesion. Traditionally blasts presence in CSF is assessed by conventional cytomorphology (CM) of cytospin slides. However, sensitivity of this method is relatively low. Flow cytometry (FC) having a higher sensitivity could provide better diagnostic applicability for CSF blasts detection. Aim. To compare results of tumor cells detection in CSF of children with AL by flow FC and CM. Methods. 183 samples from 52 boys and 31 girls aged from 5 months to 15 years with different types of acute lymphoblastic leukemia (ALL) (77 patients), acute myeloid leukemia (AML) (5 patients) and acute biphenotypic leukemia (1 patient) were investigated. 17 positive samples obtained by traumatic lumbar puncture were excluded from analysis because tumor blasts were also detected in peripheral blood. Comparison between FC and CM data was performed in 166 samples. Among these samples 61 was taken at the time of initial diagnostics, 34 – during AL follow-up, 17 – at relapse and 54 – during relapse monitoring. Monoclonal antibodies panels were constructed according to immunophenotype of tumor cells in bone marrow. Results. In 24 out of 166 samples (14.5%) tumor cells were detected by CM. In all these cases blasts were also found by FC, while FC allowed finding blasts in other 35 samples. Thus the total number of FC-positive samples was 59 out of 166 (35.5%). This frequency was significantly higher than rate of CM-positive cases (g < 0.0001). Among initial diagnostics samples there were 20 FC-positive and only 10 CM-positive patients (32.8% vs. 16.1%, p=0.0585). At relapse 9 (52.9%) patients were FC-positive, while 6 (35.3%) were CM-positive (p=0.4897). In both B-lineage and T-lineage ALL, analyzed separately, FC detected blasts in CSF frequently than CM (p=0.0098 and p=0.0002 respectively). Absolute blast count in 1 ml in CSF samples, positive by both methods was significantly higher than in samples, positive only by FC (median = 418, range 8–158171 and median = 34, range 5–2762 respectively, g = 0.0002). Thus FC allows detecting tumor cells in CSF much more frequently than conventional CM, which could be explained mainly by higher FC sensitivity. Moreover FC is applicable also for qualitative and quantitative monitoring of CNS lesion. Nevertheless prognostic impact of FC CSF investigation is questionable. Among 13 patients in whom discordant results were obtained in initial diagnostics samples and at relapse, only for one patient risk stratification could have been changed. For all other patients there were other risk factors, that decreased significance of FC leukemic blast detection in CSF. Conclusion. Flow cytometry allows more frequent detection of tumor blasts in CSF of children with AL, while prognostic significance of these findings is still unclear and needs to be confirmed in large prospective trials. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Sestrin2 Protects Dopaminergic Cells against Rotenone Toxicity through AMPK-Dependent Autophagy Activation

2015 ◽  
Vol 35 (16) ◽  
pp. 2740-2751 ◽  
Author(s):  
Yi-Sheng Hou ◽  
Jun-Jie Guan ◽  
Hai-Dong Xu ◽  
Feng Wu ◽  
Rui Sheng ◽  
...  
Keyword(s):  
Ubiquitin Proteasome System ◽  
Dependent Manner ◽  
Dopaminergic Cells ◽  
Protein Levels ◽  
Autophagy Induction ◽  
Ubiquitin Proteasome ◽  
Autophagic Degradation ◽  
Ampk Activity ◽  
Amp Activated Protein Kinase ◽  
Autophagy Activity

Dysfunction of the autophagy-lysosomal pathway (ALP) and the ubiquitin-proteasome system (UPS) was thought to be an important pathogenic mechanism in synuclein pathology and Parkinson's disease (PD). In the present study, we investigated the role of sestrin2 in autophagic degradation of α-synuclein and preservation of cell viability in a rotenone-induced cellular model of PD. We speculated that AMP-activated protein kinase (AMPK) was involved in regulation of autophagy and protection of dopaminergic cells against rotenone toxicity by sestrin2. The results showed that both the mRNA and protein levels of sestrin2 were increased in a TP53-dependent manner in Mes 23.5 cells after treatment with rotenone. Genetic knockdown of sestrin2 compromised the autophagy induction in response to rotenone, while overexpression of sestrin2 increased the basal autophagy activity. Sestrin2 presumably enhanced autophagy in an AMPK-dependent fashion, as sestrin2 overexpression activated AMPK, and genetic knockdown of AMPK abrogated autophagy induction by rotenone. Restoration of AMPK activity by metformin after sestrin2 knockdown recovered the autophagy activity. Sestrin2 overexpression ameliorated α-synuclein accumulation, inhibited caspase 3 activation, and reduced the cytotoxicity of rotenone. These results suggest that sestrin2 upregulation attempts to maintain autophagy activity and suppress rotenone cytotoxicity through activation of AMPK, and that sestrin2 exerts a protective effect on dopaminergic cells.

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