Inhibition of damage-regulated autophagy modulator-1 (DRAM-1) impairs neutrophil differentiation of NB4 APL cells

Humbert, Magali; Mueller, Chantal; Fey, Martin F.; Tschan, Mario P.

doi:10.1016/j.leukres.2012.08.024

Cited by 20 publications

(22 citation statements)

References 20 publications

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“…We show that inhibition of autophagy, either pharmacologically or through targeted genetic knockdown of ATG7 , impedes ATRA-mediated differentiation. Thus, we propose that the process of autophagy is an important mediator of ATRA’s effects on APL cells, providing further support to recent data on leukemic cell differentiation [20–22, 31]. …”

Section: Discussionsupporting

confidence: 88%

Induction of autophagy is a key component of all-trans-retinoic acid-induced differentiation in leukemia cells and a potential target for pharmacologic modulation

Orfali

O’Donovan

Nyhan

et al. 2015

Experimental Hematology

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Acute myeloid leukemia (AML) is characterized by the accumulation of immature blood cell precursors in the bone marrow. Pharmacologically overcoming the differentiation block in this condition is an attractive therapeutic avenue, which has only achieved success in a subtype of AML, acute promyelocytic leukemia (APL). Attempts to emulate this success in other AML subtypes have thus far been unsuccessful. Autophagy is a conserved protein degradation pathway with important roles in mammalian cell differentiation, particularly within the hematopoietic system. In this study we demonstrate the functional importance of autophagy in APL cell differentiation. We show that autophagy is increased during ATRA-induced granulocytic differentiation of the APL cell line NB4, and that this is associated with increased expression of LC3-II and GATE-16 proteins involved in autophagosome formation. Autophagy inhibition, using either drugs (chloroquine/3-methyladenine) or short-hairpin (sh)RNA targeting the essential autophagy gene ATG7, attenuates myeloid differentiation. Importantly, we show that enhancing autophagy promotes ATRA-induced granulocytic differentiation of an ATRA-resistant derivative of the non-APL AML HL60 cell line (HL60-Diff-R). These data support the development of strategies to stimulate autophagy as a novel approach to promote differentiation in AML.

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Section: Discussionsupporting

confidence: 88%

Induction of autophagy is a key component of all-trans-retinoic acid-induced differentiation in leukemia cells and a potential target for pharmacologic modulation

Orfali

O’Donovan

Nyhan

et al. 2015

Experimental Hematology

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“…14,49 Interestingly, it has been shown that the induction of autophagy is required for the differentiation of acute myeloid leukemia (AML) cells. 50,51 The accelerated differentiation might be explained, at least partially, by the autophagic degradation of the driving oncoprotein. 52 The question arises, whether the induction of differentiation by autophagy regulators is differently regulated in AML cells than in healthy Figure 5 Autophagic activity declines during neutrophil differentiation and pharmacological induction of autophagy delays the process of neutrophil differentiation.…”

Section: Discussionmentioning

confidence: 99%

The generation of neutrophils in the bone marrow is controlled by autophagy

et al. 2014

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Autophagy has been demonstrated to have an essential function in several cellular hematopoietic differentiation processes, for example, the differentiation of reticulocytes. To investigate the role of autophagy in neutrophil granulopoiesis, we studied neutrophils lacking autophagy-related (Atg) 5, a gene encoding a protein essential for autophagosome formation. Using Cre-recombinase mediated gene deletion, Atg5-deficient neutrophils showed no evidence of abnormalities in morphology, granule protein content, apoptosis regulation, migration, or effector functions. In such mice, however, we observed an increased proliferation rate in the neutrophil precursor cells of the bone marrow as well as an accelerated process of neutrophil differentiation, resulting in an accumulation of mature neutrophils in the bone marrow, blood, spleen, and lymph nodes. To directly study the role of autophagy in neutrophils, we employed an in vitro model of differentiating neutrophils that allowed modulating the levels of ATG5 expression, or, alternatively, intervening pharmacologically with autophagy-regulating drugs. We could show that autophagic activity correlated inversely with the rate of neutrophil differentiation. Moreover, pharmacological inhibition of p38 MAPK or mTORC1 induced autophagy in neutrophilic precursor cells and blocked their differentiation, suggesting that autophagy is negatively controlled by the p38 MAPK-mTORC1 signaling pathway. On the other hand, we obtained no evidence for an involvement of the PI3K-AKT or ERK1/2 signaling pathways in the regulation of neutrophil differentiation. Taken together, these findings show that, in contrast to erythropoiesis, autophagy is not essential for neutrophil granulopoiesis, having instead a negative impact on the generation of neutrophils. Thus, autophagy and differentiation exhibit a reciprocal regulation by the p38-mTORC1 axis. Autophagy is an evolutionarily conserved mechanism, by which portions of cytoplasm are engulfed in a doublemembrane structure, known as the autophagosome, and delivered to lysosomes for subsequent degradation. Autophagy is dependent on autophagy-related (ATG) proteins. 1Autophagosome formation, elongation, and completion of enclosure require two ubiquitin-like conjugation systems: the first one generates the ATG5-ATG12 conjugate, which functions as a complex together with ATG16, and binds to the sequestering (pre-autophagosomal) phagophore. The second system conjugates an ATG8 homolog, LC3-I, with phosphatidylethanolamine to generate the lipidated LC3-II that associates with autophagosomes.2-4 The conversion of LC3-I into LC3-II represents a hallmark of autophagic activity and is widely used for the detection of autophagosome formation. Another frequently used marker for monitoring autophagic activity is p62, a protein, which is specifically degraded through autophagy. 5The vital role of autophagy in cell growth, development, and homeostasis has long been appreciated. Recent data also indicate its involvement in the differentiation of hematopoietic c...

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“…NB4 APL cells and their all- trans retinoic acid (ATRA)-resistant NB4-R2 subclone and HEK-293T cells were cultured as described [26]. ATRA-induced autophagy was blocked using Bafilomycin A1 (BML-CM110, Enzo Life Science, Lausen, Switzerland) at a concentration of 100nM.…”

Section: Methodsmentioning

confidence: 99%

“…Westernblot was performed as described [26]. Primary antibodies used were anti-LC3B (NB600-1384; Novus Biologicals, Cambridge, England and anti-GAPDH (MAB374; Milipore, Zug, Switzerland).…”

Section: Methodsmentioning

confidence: 99%

Inhibition of GATE-16 attenuates ATRA-induced neutrophil differentiation of APL cells and interferes with autophagosome formation

Brigger

Torbett

Chen

et al. 2013

Biochemical and Biophysical Research Communications

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Autophagy is an intracellular bulk degradation process involved in cell survival upon stress induction, but also with a newly identified function in myeloid differentiation. The autophagy-related (ATG)8 protein family, including the GABARAP and LC3 subfamilies, is crucial for autophagosome biogenesis. In order to evaluate the significance of the GABARAPs in the pathogenesis of acute myeloid leukemia, we compared their expression in primary AML patient samples, CD34+ progenitor cells and in granulocytes from healthy donors. GABARAPL1 and GABARAPL2/GATE-16, but not GABARAP, were significantly downregulated in particular AML subtypes compared to normal granulocytes. Moreover, the expression of GABARAPL1 and GATE-16 was significantly induced during ATRA-induced neutrophil differentiation of acute promyelocytic leukemia cells. Lastly, knocking down GABARAPL2/GATE-16 in APL cells attenuated neutrophil differentiation and decreased autophagic flux. In conclusion, low GABARAPL2/GATE-16 expression is associated with an immature myeloid leukemic phenotype and these proteins are necessary for neutrophil differentiation of APL cells.

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Inhibition of damage-regulated autophagy modulator-1 (DRAM-1) impairs neutrophil differentiation of NB4 APL cells

Cited by 20 publications

References 20 publications

Induction of autophagy is a key component of all-trans-retinoic acid-induced differentiation in leukemia cells and a potential target for pharmacologic modulation

Induction of autophagy is a key component of all-trans-retinoic acid-induced differentiation in leukemia cells and a potential target for pharmacologic modulation

The generation of neutrophils in the bone marrow is controlled by autophagy

Inhibition of GATE-16 attenuates ATRA-induced neutrophil differentiation of APL cells and interferes with autophagosome formation

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