The acute promyelocytic leukemia-specific PML-RARα fusion protein inhibits differentiation and promotes survival of myeloid precursor cells

Formation of PML/RARα high molecular weight nuclear complexes through the PML coiled-coil region is essential for the PML/RARα-mediated retinoic acid response

“…Experimental approach and protein stability of the PML/RARa mutants U937 cells di erentiate poorly in response to RA, as determined by the low percentage (about 30 ± 40%) of cells which acquire the capacity to reduce nitroblue tetrazolium (NBT) (Grignani et al, 1993). A series of PML/RARa mutants (Figures 2 ± 4 and 6) were expressed in the U937 cells using zinc (Zn)-inducible mouse metallothionine (MT) promoter and appropriate cell clones were selected on the basis of two criteria: (1) Zn-induced protein expression comparable to that of PML/RARa in the control U937-PML/RARa expressing clone (PR9 clone); (2) capacity of each clone to undergo RA-induced di erentiation in the absence of Zn induction at a level comparable to that of the parental U937 cells.…”

“…Expression of PML/RARa into di erent haematopoietic cell lines revealed that the fusion protein is capable of mediating the di erentiative response to RA (Grignani et al, 1993;Ruthardt et al, 1997). Here, we investigated the molecular mechanisms underlying the biological activity of the fusion protein on RAsignalling.…”

“…Here, we investigated the molecular mechanisms underlying the biological activity of the fusion protein on RAsignalling. As model system, we have chosen the U937 cell line, whose RA-sensitivity is markedly increased by PML/RARa expression (Grignani et al, 1993). We (7) or Zn expressed in U937 cells PML/RARa mutant proteins deleted in relevant functional regions and studied their biological e ects.…”

“…The ectopic expression of PML/RARa into haemopoietic precursor cell lines induces block of di erentiation triggered by physiological stimuli, and prolongs survival (Grignani et al, 1993;Rogaia et al, 1995). In vivo, this may cause accumulation of haemopoietic precursors that are unable to respond to di erentiative physiological stimuli.…”

“…However, the degradation of PML/ RARa in RA-treated APL cells takes about 24 h, suggesting that the commitment to the di erentiation process occurs in the presence of PML/RARa expression and that the fusion protein might exert a direct e ect on the RA-signalling pathway in the presence of RA (Casini and Pelicci, 1998). Direct evidence for this hypothesis came from: (1) the demonstration that inhibition of PML/RARa degradation does not prevent RA-induced di erentiation ; (2) the identi®cation of RAresistant APL mutant cell clones with constitutive degradation of PML/RARa ; and (3) the demonstration that ectopic expression of PML/ RARa in cells that are poorly sensitive or resistant to RA restores RA-sensitivity (Grignani et al, 1993;Ruthardt et al, 1997). Notably, PML/RARa enhanced RA-response in U937 cells is also associated with the degradation of the fusion protein .…”