Involvement of PML-I in reformation of PML nuclear bodies in acute promyelocytic leukemia cells by leptomycin B

Acute promyelocytic leukemia (APL) is characterized by a reciprocal translocation between chromosomes 15 and 17, t(15;17), resulting in the expression of PML-RAR alpha fusion protein, which disrupts the normal PML nuclear bodies (PML-NBs) to micro-speckled pattern, leading to loss of their original functions. Moreover, reformation of PML-NBs in APL by arsenic is considered as one of the important step for APL treatment. Leptomycin B (LMB), a nuclear export inhibitor, is commonly used to inhibit the proteins exporting from the nucleus to the cytoplasm. In the present study, we found that LMB could induce the reformation of PML-NBs in leukemia NB4 cells as well as in APL blast cells from the patients, implying that nuclear shuttle proteins might be involved in the reformation of PML-NBs. Herein, we further found that LMB totally lost the ability to induce PML-NBs reformation when the endogenous PML gene was knocked out, indicating that endogenous PML protein is probably involved in the reformation of PML-NBs. More interestingly, among all PML isoforms (i.e., seven isoforms), reformation of PML-NBs was only observed when co-transfection of PML-RAR alpha with PML-I after LMB treatment. Similarly, deletion of nuclear export signal (NES) of PML-I could also reform PML-NBs, suggesting that the protein level of endogenous PML-I in nucleus is important for the reformation of PML-NBs that interfered by PML-RAR alpha fusion protein. Additionally, LMB has synergistic effect with iAs(III) on enhancing PML-RAR alpha fusion protein degradation, and it might provide new insight into APL treatment at clinical level in the near future.