The promyelocytic leukemia (PML) protein is expressed in the diffuse nuclear fraction of the nucleoplasm and in matrix-associated structures, known as nuclear bodies (NBs). PML NB formation requires the covalent modification of PML to SUMO. The noncovalent interactions of SUMO with PML based on the identification of a SUMO-interacting motif within PML seem to be required for further recruitment within PML NBs of SUMOylated proteins. RNA viruses whose replication takes place in the cytoplasm and is inhibited by PML have developed various strategies to counteract the antiviral defense mediated by PML NBs. We show here that primary fibroblasts derived from PML knockout mice are more sensitive to infection with encephalomyocarditis virus (EMCV), suggesting that the absence of PML results in an increase in EMCV replication. Also, we found that EMCV induces a decrease in PML protein levels both in interferon-treated cells and in PMLIIIexpressing cells. Reduction of PML was carried out by the EMCV 3C protease. Indeed, at early times postinfection, EMCV induced PML transfer from the nucleoplasm to the nuclear matrix and PML conjugation to SUMO-1, SUMO-2, and SUMO-3, leading to an increase in PML body size where the viral protease 3C and the proteasome component were found colocalizing with PML within the NBs. This process was followed by PML degradation occurring in a proteasome-and SUMO-dependent manner and did not involve the SUMOinteracting motif of PML. Together, these findings reveal a new mechanism evolved by EMCV to antagonize the PML pathway in the interferon-induced antiviral defense.The PML (promyelocytic leukemia) gene was originally identified through its fusion with the RAR␣ gene in the t(15;17) translocation found in acute promyelocytic leukemia (APL) (14). PML, also known as TRIM19, is expressed in the diffuse nuclear fraction of the nucleoplasm and in matrixassociated structures, known as nuclear bodies (NBs) (17, 30). PML functions as the organizer of PML NBs, which are dynamic structures harboring numerous transiently and permanently localized proteins (35). The RBCC/TRIM motif, which contains a C 3 HC 4 (RING finger) zinc-binding domain, two cysteine/histidine-rich motifs (the B boxes B1 and B2), an a helical coiled-coil region (RBCC), is embedded within the PML protein and is required for PML NB formation (28). Due to alternative splicing, many PML isoforms are synthesized, and they are classified into seven groups, designated PML I to PML VII (reviewed in reference 28). They share the N-terminal region (exons 1 to 3), which encodes the RBCC motif, whereas they differ in their C-terminal regions. Posttranslational modification of PML by SUMO (small ubiquitin-like modifier), a ubiquitin-like protein of 11 kDa, is another requirement for PML NB formation. SUMO is covalently coupled to PML through its lysines 65, 160, and 490 via a process called SUMOylation (29). The noncovalent interaction of SUMO with PML through a SUMO-interacting motif (SIM; also named SBD for SUMO binding domain) (46) has been sug...

Heavy metals are important regulators of cell apoptosis. Manganese (Mn 2 þ ) is a potent inducer of apoptosis in different cell types, but the precise mechanisms that mediate such effects are not well defined. We previously reported that Mn 2 þ was a potent apoptotic agent in human B cells, including lymphoma B cell lines. We show here that Mn 2 þ -induced cell death in human B cells is associated with caspase-8-dependent mitochondrial activation leading to caspase-3 activity and apoptosis. We used specific caspase-8 interfering shRNAs to reduce caspase-8 expression, and this also reduced Mn 2 þ -induced caspase-3 activation and apoptosis. Mn 2 þ -triggered caspase-8 activation is associated with a specific pathway, which is independent of Fas-associated death domain protein, and dependent on the sequential activation of p38-mitogen-activated protein kinase (p38 MAPK) and mitogen-and stress-response kinase 1 (MSK1). Inhibition of p38 activity using either pharmacological inhibitors or dominant-negative mutant forms of p38 blocked Mn 2 þ -mediated phosphorylation of MSK1 and blocked subsequent caspase-8 activation. However, specific inhibitors and the expression of a dominant-interfering mutant of MSK1 only inhibited caspase-8 activation, but not p38 activity. These findings suggest a novel model for the regulation of caspase-8 during Mn 2 þ -induced apoptosis based on the sequential activation of p38 MAPK, MSK1, caspase-8 and mitochondria, respectively.

EBV infects a large proportion of the human population worldwide and is one of the major viruses with human B lymphocyte tropism. It can immortalize human B lymphocytes and controls their resistance to apoptosis. EBV infection is associated with several lymphomas, including Burkitt’s lymphoma. In this report we show that EBV infection leads to the post-transcriptional down-regulation of expression of the proapoptotic protein Bim. This process involves the phosphorylation of BimEL by the constitutive EBV-activated kinase ERK1/2, followed by its degradation through the proteasome pathway. We also show that ectopic expression of BimEL in EBV-positive Burkitt’s lymphoma cells can enhance the sensitivity of these cells to serum deprivation-dependent apoptosis. Thus, EBV-mediated resistance to growth factor deprivation in human B lymphocytes is dependent on BimEL expression. Our data suggest that this regulatory pathway is an important contributor to the oncogenic potential of EBV.