No abstract

Acute promyelocytic leukemia (APL) is associated with the t(15;17) translocation, which generates a PML͞RAR␣ fusion protein between PML, a growth suppressor localized on nuclear matrix-associated bodies, and RAR␣, a nuclear receptor for retinoic acid (RA). PML͞RAR␣ was proposed to block myeloid differentiation through inhibition of nuclear receptor response, as does a dominant negative RAR␣ mutant. In addition, in APL cells, PML͞RAR␣ displaces PML and other nuclear body (NB) antigens onto nuclear microspeckles, likely resulting in the loss of PML and͞or NB functions. RA leads to clinical remissions through induction of terminal differentiation, for which the respective contributions of RAR␣ (or PML͞RAR␣) activation, PML͞ RAR␣ degradation, and restoration of NB antigens localization are poorly determined. Arsenic trioxide also leads to remissions in APL patients, presumably through induction of apoptosis. We demonstrate that in non-APL cells, arsenic recruits the nucleoplasmic form of several NB antigens onto NB, but induces the degradation of PML only, identifying a powerful tool to approach NB function. In APL cells, arsenic targets PML and PML͞RAR␣ onto NB and induces their degradation. Thus, RA and arsenic target RAR␣ and PML, respectively, but both induce the degradation of the PML͞ RAR␣ fusion protein, which should contribute to their therapeutic effects. The difference in the cellular events triggered by these two agents likely stems from RA-induced transcriptional activation and arsenic effects on NB proteins.

Analyzing the pathways by which retinoic acid (RA) induces promyelocytic leukemia͞retinoic acid receptor ␣ (PML͞RAR␣) catabolism in acute promyelocytic leukemia (APL), we found that, in addition to caspase-mediated PML͞RAR␣ cleavage, RA triggers degradation of both PML͞RAR␣ and RAR␣. Similarly, in non-APL cells, RA directly targeted RAR␣ and RAR␣ fusions to the proteasome degradation pathway. Activation of either RAR␣ or RXR␣ by specific agonists induced degradation of both proteins. Conversely, a mutation in RAR␣ that abolishes heterodimer formation and DNA binding, blocked both RAR␣ and RXR␣ degradation. Mutations in the RAR␣ DNA-binding domain or AF-2 transcriptional activation region also impaired RAR␣ catabolism. Hence, our results link transcriptional activation to receptor catabolism and suggest that transcriptional up-regulation of nuclear receptors by their ligands may be a feedback mechanism allowing sustained target-gene activation.

The PML protein is associated to nuclear bodies (NBs) whose functions are as yet unknown. PML and two other NBs-associated proteins, Sp100 And ISG20 are directly induced by interferons (IFN). PML and Sp100 proteins are covalently linked to SUMO-1, and ubiquitin-like peptide. PML NBs are disorganized in acute promyelocytic leukemia and during several DNA virus infections. In particular, the HSV-1 ICP0 protein is known to delocalize PML from NBs. Thus, NBs could play an important role in oncogenesis, IFN response and viral infections. Here, we show that HSV-1 induced PML protein degradation without altering its mRNA level. This degradation was time-and multiplicity of infectiondependent. Sp100 protein was also degraded, while another SUMO-1 conjugated protein, RanGAP1 and the IFN-induced protein kinase PKR were not. The proteasome inhibitor MG132 abrogated the HSV-1-induced PML and Sp100 degradation and partially restored their NB-localization. HSV-1 induced PML and Sp100 degradation constitutes a new example of viral inactivation of IFN target gene products.