Retrovirus-Mediated Expression of E2A-PBX1 Blocks Lymphoid Fate but Permits Retention of Myeloid Potential in Early Hematopoietic Progenitors.

The oncogenic transcription factor E2A-PBX1 is expressed consequent to chromosomal translocation 1;19 and is an important oncogenic driver in cases of pre-B-cell acute lymphoblastic leukemia (ALL). Elucidating the mechanism by which E2A-PBX1 induces lymphoid leukemia would be expedited by the availability of a tractable experimental model in which enforced expression of E2A-PBX1 in hematopoietic progenitors induces pre-B-cell ALL. However, hematopoietic reconstitution of irradiated mice with bone marrow infected with E2A-PBX1-expressing retroviruses consistently gives rise to myeloid, not lymphoid, leukemia. Here, we elucidate the hematopoietic consequences of forced E2A-PBX1 expression in primary murine hematopoietic progenitors. We show that introducing E2A-PBX1 into multipotent progenitors permits the retention of myeloid potential but imposes a dense barrier to lymphoid development prior to the common lymphoid progenitor stage, thus helping to explain the eventual development of myeloid, and not lymphoid, leukemia in transplanted mice. Our findings also indicate that E2A-PBX1 enforces the aberrant, persistent expression of some genes that would normally have been down-regulated in the subsequent course of hematopoietic maturation. We show that enforced expression of one such gene, Hoxa9, a proto-oncogene associated with myeloid leukemia, partially reproduces the phenotype produced by E2A-PBX1 itself. Existing evidence suggests that the 1;19 translocation event takes place in committed B-lymphoid progenitors. However, we find that retrovirus-enforced expression of E2A-PBX1 in committed pro-B-cells results in cell cycle arrest and apoptosis. Our findings indicate that the neoplastic phenotype induced by E2A-PBX1 is determined by the developmental stage of the cell into which the oncoprotein is introduced.

Experimental characterization of the mechanism of perfluorocarboxylic acids' liver protein bioaccumulation: the key role of the neutral species.

Perfluorocarboxylic acids (PFCAs) of chain length greater than seven carbon atoms bioconcentrate in the livers of fish. However, a mechanistic cause for the empirically observed increase in the bioconcentration potential of PFCAs as a function of chain length has yet to be determined. To this end, recombinant rat liver fatty acid-binding protein (L-FABP) was purified, and its interaction with PFCAs was characterized in an aqueous system at pH 7.4. Relative binding affinities of L-FABP with PFCAs of carbon chain lengths of five to nine were established fluorimetrically. The energetics, mechanism, and stoichiometry of the interaction of perfluorooctanoic acid (PFOA) with L-FABP were examined further by isothermal titration calorimetry (ITC) and electrospray ionization combined with tandem mass spectrometry (ESI-MS/MS). Perfluorooctanoic acid was shown to bind to L-FABP with an affinity approximately an order of magnitude less than the natural ligand, oleic acid, and to have at least 3:1 PFOA:L-FABP stoichiometry. Two distinct modes of PFOA binding to L-FABP were observed by ESI-MS/MS analysis; in both cases, PFOA binds solely as the neutral species under typical physiological pH and aqueous concentrations of the anion. A comparison of their chemical and physical properties with other well-studied biologically relevant chemicals showed that accumulation of PFCAs in proteins as the neutral species is predictable. For example, the interaction of PFOA with L-FABP is almost identical to that of the acidic ionizing drugs ketolac, ibuprofen, and warfarin that show specificity to protein partitioning with a magnitude that is proportional to the K(OW) (octanol-water partitioning) of the neutral species. The experimental results suggest that routine pharmacochemical models may be applicable to predicting the protein-based bioaccumulation of long-chain PFCAs.