Abstract

Abstract 1046

Autophagy is a self-digesting recycling mechanism for cellular organelles and macromolecules with cytoprotective functions. On the other hand, autophagy might also lead to caspase-independent, type II cell-death. Of interest, there is accumulating evidence for non-metabolic, cell specific functions of autophagy in mature hematopoietic cells but also during hematopoietic differentiation. We therefore aimed at investigating the involvement of autophagy in the molecular pathogenesis of acute myeloid leukemia (AML) and, in particular, in acute promyelocytic leukemia (APL). We found significantly increased autophagic activity during ATRA-induced neutrophil differentiation of NB4 and HL60 AML cells as evidenced by increased autophagosome formation (appearance of LC3-GFP dots), LC3-I to -II conversion and degradation of p62 (SQSTM1), which is degraded in the autophagosome. Most importantly, ATRA treatment induced a novel type of autophagy that was independent of the key autophagy gene Beclin-1 as opposed to autophagy induced by starvation or chemotherapeutics. Moreover, autophagy seen during neutrophil differentiation of AML cells was clearly less intense than, for example, starvation-induced autophagy as measured by the number of LC3-GFP dots per cell. We have previously shown that the autophagy gene ATG5 as well as death-associated protein kinase 2 (DAPK2) are crucial in all-trans retinoic (ATRA)-induced neutrophil differentiation. Since it was found earlier that overexpression of DAPK2 can induce autophagy in epithelial cells, we were asking whether there might be a direct link from DAPK2 to ATG5. Indeed, we found direct DAPK2-ATG5 binding when overexpressed in 293T HEK cells. Moreover, we were able to pull down endogenous DAPK2 together with ATG5 in ATRA-treated NB4 APL cells. Using DAPK2 overexpressing NB4 cells, we found increased DAPK2-ATG5 interaction with progressive neutrophil differentiation. Further, knocking down DAPK2 caused decreased ATG5-ATG12 complex formation during AML differentiation. Lastly, we found that pharmacological activation of autophagy in combination with ATRA significantly boosted AML differentiation, representing a potential new strategy for APL therapy. In summary, we show that autophagy is needed for neutrophil differentiation of AML cells, possibly via the newly identified DAPK2-ATG5 pathway, and would like to refer to this new type of Beclin-1-independent, non-canonical autophagy as myeloid differentiation-associated autophagy (MDAA).

Disclosures:

No relevant conflicts of interest to declare.

Author notes

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Asterisk with author names denotes non-ASH members.