Acute promyelocytic leukemia (APL) is characterized by an arrest of the terminal differentiation of myeloid cells into granulocytes.1
The genetic hallmark of this disease is a reciprocal chromosomal translocation that joins the RAR (retinoic acid receptor) α gene to the PML (promyelocytic leukemia) gene.2
The resulting chimeric protein, PML-RARα, is thought to be responsible for the arrest of granulopoiesis by directly inhibiting the transcription of the retinoic acid target gene, a mechanism that could account for its leukemogenic properties.3–5
Administration of pharmacological doses of All-trans retinoic acid (ATRA) to APL patients produces a clinical remission of the disease by inducing the maturation of promyelocytes and the degradation of the PML-RARα protein, leading to the eradication of the leukemic stem cells.5–8
However, some of these patients relapse due to the development of ATRA resistance by leukemic cells. Moreover, one of the main complications of ATRA treatment of APL patients is the ATRA syndrome, which is caused by the accumulation of mature granulocytes and can lead to a fatal outcome in APL patients.9
Being able to identify the molecular pathways that can enhance the effectiveness of ATRA-induced both differentiation of APL cells and death of mature cells will therefore be helpful in designing new strategies for improving the treatment of patients affected by APL.
Macroautophagy (referred to as autophagy hereafter) is a lysosomal process that induces the degradation of cytoplasmic constituents such as proteins and organelles, thus leading to cell renovation. In response to environmental and hormonal changes, autophagy is induced to provide cells with the nutrients and energy necessary for cell survival and cellular remodeling.10–12
Paradoxically, under certain circumstances, autophagy contributes to programmed cell death.13
During the autophagy process, a cell sequesters cytoplasmic material within double-membrane vesicles known as autophagosomes, and delivers them to lysosomes for degradation by lytic enzymes. At the molecular level, the Atg proteins (the products of autophagy-related genes) orchestrate the sequential steps of generating autophagosomes.14
An early event in the initiation of autophagy is the binding of Beclin 1 (a mammalian ortholog of yeast Atg6) to the class III PtdIns 3-kinase (hVps34) which promotes the recruitment of other Atg proteins to the phagophore membrane.15,16
Beclin 1 possesses a BH3 only-domain which is necessary for its binding to Bcl-2 and Bcl-XL
. This binding represses Beclin 1-dependent autophagy in several experimental settings.17–19
Recent studies have established the role of autophagy during mammalian development and cell differentiation, processes that require an extensive cellular remodeling by a rapid change in intracellular contents, such as organelles and proteins.20,21
For example, differentiation of T-lymphocytes and erythrocytes requires a selective removal of mitochondria by autophagy.22–24
Along this line, mice lacking the essential autophagy-related gene Atg7
in the hematopoietic system develop severe anemia and lymphopenia as a result of defective removal of mitochondria.23
are involved in normal adipocyte differentiation suggesting an essential function of autophagy in adipogenesis.25,26
Autophagy is also an important event for megakaryocytic maturation of the chronic myelogenous leukemia,27
and differentiation of neuroblastoma and glioma stem progenitor cells, as well.28,29
Moreover, autophagy is required for the pre-implantation development of mouse embryos,30
and its deficiency in murine models lacking the key autophagy-regulatory genes (e.g., Ambra1, Atg5
) induces the accumulation of abnormal proteins during the development of the nervous system20
suggesting its essential role in developmental processes.
However, the function of autophagy-regulatory pathways in ATRA-induced neutrophil/granulocyte differentiation of APL cells is still not well documented. In the present study, we investigated the autophagic responses during the course of ATRA-induced maturation of an APL-derived cell line, NB4 cells. In particular, we explored the involvement of Beclin 1 in regulating NB4 cell death and differentiation, two processes that occur during ATRA treatment of APL cells.