Cooperation between the RING+B1-B2 and coiled-coil domains of PML is necessary for its effects on cell survival

Abstract

PML/RARa is the abnormal protein product of the Acute Promyelocytic Leukemia-speci®c 15;17 translocation. Both the PML and RARa components are required for the PML/RARa biological activities, namely its capacity to block di erentiation and to increase survival of haematopoietic precursors. The physiological role of PML and its contribution to the function of the fusion protein are unknown. PML localizes to the cytoplasm and within speci®c nuclear bodies (NBs). In vitro, overexpression of PML correlates with suppression of cell transformation. The PML aminoterminal portion retained within the PML/RARa protein contains the RING ®nger, two newly de®ned cystein/histidine-rich motifs called Bboxes (B1 and B2) and a coiled-coil region. We report here that PML has a growth suppressive activity in all the cell lines tested, regardless of their transformed phenotype, and that the cellular basis for the PML growth suppression is induction of apoptotic cell death. Analysis of various nuclear and cytoplasmic PML isoforms showed that the PML growth suppressive activity correlates with its nuclear localization. Analysis of the localization and growth suppressive activity demonstrated that: (i) the Ring+B1-B2 and coiled-coil regions are both indispensable and su cient to target PML to the NBs; (ii) individual deletions of the various PML domains have no e ect on its growth suppressor activity; (iii) the Ring+B1-B2 region exerts a partial growth suppressor activity but its fusion with the coiled-coil region is su cient to recapitulate the suppressive function of wild type PML. These results indicate that PML is involved in cell survival regulation and that the PML component of the fusion protein (Ring+B1-B2 and coiled-coil regions) retains intact biological activity, thereby suggesting that the e ects of PML/RARa on survival derive from the activation of the incorporated PML sequence.

Cloning and Characterization of a Novel RING-B-box-Coiled-coil Protein with Apoptotic Function

“…The apoptotic activity of PML appears to be mediated through its RING domain (24). In addition, deletion of the coiled-coil motif in PML yields a diffuse nuclear localization pattern and results in no growth suppression (50). Although MAIR is a cytoplasmic protein rather than a nuclear protein, corresponding regions of MAIR appear to be required for the apoptosis-inducing activity.…”

Deregulated TGF-β signaling in leukemogenesis

“…Another study concluded that nuclear localization of PML is required for its growth-suppressive function. In this study, a PML cytoplasmic isoform did not yield an appreciable effect on cell growth inhibition when compared with a PML nuclear isoform (Fagioli et al, 1998). Owing to lack of any known functions for the cytoplasmic isoforms by these studies, research on PML in this decade has focused exclusively on the nuclear isoforms.…”

PRIMA-1MET induces nucleolar accumulation of mutant p53 and PML nuclear body-associated proteins

“…PML was originally identified as the fusion partner with the retinoic acid receptor RARalpha in leukemic blasts from patients suffering from acute promyelocytic leukemia (APL), leading to disrupted PML-NBs (Melnick and Licht, 1999). Accumulating evidence indicates that PML-NBs play regulatory roles in cellular processes such as cell growth (Fagioli et al, 1998;Cohen et al, 2001), chromatin remodeling (Amin et al, 2001), cellular differentiation (Drouin et al, 2001), protein degradation (Lallemand-Breitenbach et al, 2001), transcriptional regulation (Li et al, 2000;Zhong et al, 2000), antiviral response (Everett, 2001;Regad and Chelbi-Alix, 2001), cellular senescence (Ferbeyre et al, 2000;Pearson et al, 2000;Bischof et al, 2002;Langley et al, 2002), tumor suppression (reviewed by Salomoni and Pandolfi (2002)), DNA repair (Bischof et al, 2001;Carbone et al, 2002) and apoptosis (Wang et al, 1998;Fogal et al, 2000;Guo et al, 2000;D'Orazi et al, 2002;Hofmann et al, 2002).…”