BAG-1 IN ACUTE LEUKEMIA: HUNDRED FACES OF A SINGLE PROTEIN

Introduction. Bcl-2 associated athanogene-1 (Bag-1), a founding member of BAG protein family, is a multifunctional protein which has a role in a wide range of cellular processes including apoptosis, cell survival, transcription, cell motility and proliferation. The involvement of Bag-1 in different cellular pathways can be in part examined by the sub-cellular compartmentalisation of the three major Bag-1 isoforms (Bag-1L, Bag-1M and Bag-1S), generated by alternative translation from a single mRNA, and in part by its interaction with a large number of disparate proteins, including Bcl-2, Raf-1, nuclear hormone receptors, subunits of the ubiquitinylation proteasome complex, Hsc70 and Hsp70. Aims. The elevated level of Bag-1 protein has been confirmed as a considerable index in several malign diseases. To determine significance Bag-1 might have in the processes of leukemogenesis a sequence of human leukemic cell lines and pediatric bone marrow samples with confirmed acute myeloid leukemia were included in our research. Our goal was to clarify the molecular mechanisms potentially in charge for Bag-1 action, in either leukemic cell lines or primary cell cultures. Methods. In vitro studies were based on a small-interfering RNA (siRNA) approach and the results were validated using standard techniques for mRNA and protein expression study. Assays for cell cycle and apoptosis detection were performed. Results. The protein study revealed elevated Bag-1 levels in human leukemic cell lines of both myeloid (ML2, THP1, NOMO1, NB4, MV4;11 and HL60) and lymphoid (REH, RS4;11, 697 and JURKAT) origin. A different expression pattern of Bag-1 protein isoforms was noted for two considered groups of patients, AML or ALL, with changes in protein expression profile at different point of the disease. After Bag-1 was knock-down, a modest effect on cell death or cell cycle profile was observed for the human cell lines while primary cultures showed to be more sensitive to Bag-1 silencing. However, significant decrease was confirmed at the expression level of a wide range of proteins, specially the ones involved in the regulation of apoptosis (Bcl-2, PARP, Caspase-3), cell cycle (p27, CDK2, Cyclin D1) and autophagy (LC3, p62), without affecting the mRNA levels. When double silencing experiments of Bag-1 and Bag-3 (a family co-member) were performed, the effect on cell death and cell cycle arrest were found enforced, suggesting a connection between two proteins to be significant for cells faith in leukemia. Conclusions. Results indicate that the role of Bag-1 in cell death prevention might be more related to lymphoid leukemia, while it might be considered more significant for cell differentiation in myeloid cells. At the same time, elevated Bag-1 protein expression levels in acute leukemia indicates its possible significance for AML and ALL growth. A different expression profile of Bag-1L, Bag-1M and Bag-1S isoforms in myeloid with respect to lymphoid leukemia could lead to the hypothesis that Bag-1 might play a role in leukemia switch, triggering to either ALL or AML phenotype.