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1 Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, 300 Cheoncheondong, Jangangu, Suwon, Gyeonggido 440-746, Korea
2 Department of Biological Science, Sungkyunkwan University, Suwon, Korea
Correspondence
Jin-Hyun Ahn
jahn{at}med.skku.ac.kr
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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These authors contributed equally to this work. 
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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). Among the approximately 150 proteins encoded by HCMV, the 72 kDa major immediate-early (IE) protein IE1 is the first and most abundantly expressed in infected cells and is believed to perform key regulatory functions in the preparation of the cells for the production of progeny virions (Mocarski, 2001). Studies with an IE1-deleted mutant virus have shown that IE1 is required for virus growth in cell cultures at a low m.o.i., whereas it appears to be unnecessary at a high m.o.i. (Greaves & Mocarski, 1998; Mocarski et al., 1996). This growth defect of the IE1-deleted mutant virus has been attributed to a failure or delay in the accumulation of viral delayed-early gene products (Ahn & Hayward, 2000; Gawn & Greaves, 2002; Greaves & Mocarski, 1998; Mocarski et al., 1996).
During the early stages of infection, IE1 initially targets to subnuclear structures referred to as PML oncogenic domains (PODs) or nuclear domain 10 (ND10). This targeting of POD by IE1 is a very transient effect. The PODs are disrupted within 3–4 h after infection, leading to the relocalization of both IE1 and POD components, including PML and Sp100, into the nucleoplasm (Ahn et al., 1998; Ahn & Hayward, 1997; Korioth et al., 1996; Wilkinson et al., 1998). The POD-disrupting activity exhibited by IE1 is well-conserved among the IE proteins of other herpesviruses, such as herpes simplex virus 1 (HSV-1) ICP0 (Everett & Maul, 1994; Maul & Everett, 1994; Maul et al., 1993) and Epstein–Barr virus (EBV) Zta (Adamson & Kenney, 2001; Bell et al., 2000), which suggests its importance in the progression of viral growth. Among betaherpesvirus subfamily members, this activity is also conserved in mouse cytomegalovirus (MCMV) (Tang & Maul, 2003). However, in Human herpesvirus 6, the equivalent IE1 proteins also target to the PODs, but remain stably associated with them throughout lytic infection (Gravel et al., 2002; Stanton et al., 2002). PODs have also been suggested to be involved in numerous cellular functions, including tumour suppression, transcriptional regulation, apoptosis, antiviral responses and the maintenance of genome stability (for reviews, see Bernardi & Pandolfi, 2003; Dellaire & Bazett-Jones, 2004; Regad et al., 2001; Salomoni & Pandolfi, 2002; Zhong et al., 2000b). The PML protein has multiple forms and is often covalently modified by the small ubiquitin-like modifier (SUMO) (Johnson, 2004; Seeler & Dejean, 2003), which is required for the formation of mature PODs (Ishov et al., 1999; Lallemand-Breitenbach et al., 2001; Zhong et al., 2000a).
Recent studies have indicated that the disruption of PODs by both HCMV IE1 and HSV-1 ICP0 involves the loss of sumoylated forms of PML (Boutell et al., 2003; Lee et al., 2004; Müller & Dejean, 1999). However, these two viral proteins appear to exploit different mechanisms. Unlike the reaction observed with ICP0, the PML desumoylation induced by IE1 is relatively resistant to the proteasome inhibitor MG132 and also does not result in any reduction in the levels of unmodified PML (Lee et al., 2004). Recently, it has been shown that ICP0 does not have intrinsic SUMO protease activity in vitro, but that ICP0-mediated PML degradation requires lysine 160, which is one of the principal targets of PML for sumoylation (Boutell et al., 2003). IE1 has also been found to bind to the N-terminal region of PML containing the RING-finger domain (Ahn et al., 1998), which is involved in both protein oligomerization and in the formation of supramolecular structures (Kentsis et al., 2002). In addition, a physical interaction of IE1 with PML has been demonstrated to be required at least for IE1-induced PML desumoylation in vivo (Lee et al., 2004). However, how IE1 induces PML desumoylation and POD disruption has yet to be elucidated clearly.
In the present study, we characterized the effects of IE1 in induction of desumoylation of PML. We obtained evidence to suggest that IE1 neither inhibits sumoylation of PML nor possesses intrinsic SUMO protease activity against PML in vitro. Furthermore, we determined that IE1 is also capable of interfering with the formation of SUMO-independent PML aggregates, suggesting that inhibition of PML oligomerization by IE1 may play an important role in inducing PML desumoylation in vivo.
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METHODS |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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).
Expression vectors and plasmids.
Plasmids encoding GST–SAE2/SAE1 and His–Ubc9 were provided by Ronald T. Hay (University of St Andrews, UK) and Chin Ha Chung (Seoul National University, Korea), respectively. pJHA354, which expresses GST–SUMO-1GG (an active form), has been described previously (Ahn et al., 2001). pT-E1E2S1, which encodes the E1 and E2 enzymes of SUMO conjugation, as well as an active form of SUMO-1, was provided by Hisato Saitoh (University of Kumamoto, Japan) and was used to introduce a synthetic SUMO-1 conjugation pathway into Escherichia coli (Uchimura et al., 2004a). Plasmids encoding HA–PML IV (pUS109) and HA–PML VI (pUS112) have been described previously (Lee et al., 2004). Plasmids expressing FLAG–Axam (pHR16), FLAG–SuPr-1 (pHR28) and FLAG–SuPr-1(C466S) (pHR27) were generated on a background of pSG5 (Green et al., 1988), using Gateway Technology (Invitrogen). Plasmids expressing FLAG–SENP1 and FLAG–SENP1(C603S) (Bailey & O'Hare, 2004) were provided by Peter O'Hare (Marie Curie Research Institute, UK). Plasmids expressing myc–IE1 (pRYK107), myc–PML IV (pJHA481) and myc–SENP1 (pHR52) were generated on a background of pCS3-MT (Roth et al., 1991), using Gateway Technology. The HA–PML IV proteins containing the K160R or K490R mutation, as well as the FLAG–PML VI-3M protein, which contains the K65/160/490R mutation, were generated according to the Stratagene QuickChange site-directed mutagenesis protocol.
In vitro sumoylation assay.
Recombinant GST–SAE2/SAE1 and GST–SUMO-1GG were expressed and purified from E. coli as described previously (Tatham et al., 2001). His–Ubc9 was expressed and purified using Ni-NTA columns (Invitrogen) in accordance with the manufacturer's instructions. Myc-tagged PML IV protein was prepared using the TNT Quick Coupled Transcription/Translation System (Promega). GST–IE1 protein was purified from E. coli. Sumoylation reactions were conducted in a 30 µl volume containing a substrate protein (in vitro-translated myc–PML IV, GST–IE1, or both), 600 ng GST–SAE2/SAE1, 150 ng His–Ubc9 and 500 ng GST–SUMO-1GG in buffer [50 mM Tris/HCl (pH 7.5), 5 mM MgCl2, 3 mM ATP]. The reaction was incubated for 1 h at 37 °C. After termination of the reaction using SDS sample buffer containing 
-mercaptoethanol, the reaction products were fractionated by 8 % SDS-PAGE. The sumoylated forms of myc–PML IV or GST–IE1 were detected by immunoblotting assays, using anti-myc or anti-IE1 antibodies.
Generation of SUMO-1-modified proteins in E. coli.
To generate the SUMO-1-conjugated PML or IE1 protein in E. coli, both a plasmid (AmR) encoding GST–PML VI or GST–IE1 and a plasmid (CmR) encoding SUMO-1 conjugation components (E1, E2 and SUMO-1GG) were introduced into E. coli BL21 (DE3) and the GST fusion proteins were expressed and purified, as described previously (Uchimura et al., 2004a). For GST–PML VI, both the unmodified and the SUMO-1-modified forms of GST–PML VI often accumulated in insoluble inclusion bodies. To obtain the soluble forms of GST–PML VI, the cell pellets were suspended in 9 vols cell lysis buffer II [50 mM Tris/HCl (pH 8.0), 10 mM EDTA (pH 8.0), 10 mM NaCl, 0.5 % Triton X-100] and incubated for 5 min at room temperature. After 15 min of centrifugation at 15 000 r.p.m. (Beckman JA-25.50) at 4 °C, the pellets were resuspended in inclusion body solubilization buffer I [50 mM Tris/HCl (pH 8.0), 1 mM EDTA (pH 8.0), 100 mM NaCl, 8 M urea, 0.1 mM PMSF] and incubated for 1 h at room temperature. Nine volumes of inclusion body solubilization buffer II [50 mM KH2PO4 (pH 10.7), 1 mM EDTA (pH 8.0), 50 mM NaCl] was added and the reaction incubated for 30 min at room temperature. After centrifugation, the supernatant containing the soluble GST–PML VI protein was collected. The sample was adjusted to a pH of 8.0 and then incubated for an additional 30 min at room temperature. Prior to use, samples were dialysed using PBS (pH 7.4).
Production of GST–IE1 in insect cells using recombinant baculovirus.
GST–IE1 protein was also generated in insect cells, using the BAC-to-BAC baculovirus expression system (Invitrogen). In brief, the transfer vector pHJK35, which encodes recombinant GST–IE1, was generated on a background of pDEST20 (Invitrogen) using Gateway Technology and introduced into DH10Bac E. coli harbouring a baculovirus bacmid. Recombinant bacmid DNA containing the GST–IE1 expression cassette was then isolated and verified via PCR analysis. Sf9 cells were transfected with the recombinant bacmid using Cellfectin reagent (Invitrogen). Recombinant baculovirus particles were harvested from the transfected cells and amplified. To conduct the large-scale production of GST–IE1, Hi-Five cells were infected with recombinant baculovirus at an m.o.i. of 3. At 96 h post-infection, the cells were harvested. Cell pellets were resuspended in lysis buffer [50 mM Tris/HCl (pH 7.5), 100 mM NaCl, 5 mM MgCl2, 0.1 % NP-40, 10 % glycerol (v/v), 1 mM PMSF, 1 mM dithiothreitol, 1x protease inhibitor cocktail] and the GST–IE1 protein was purified.
Transient DNA transfection.
293T cells were transfected via the N,N-bis-(2-hydroxyethyl)-2-aminoethanesulfonic acid-buffered saline version of the calcium phosphate method, as described previously (Lee et al., 2004). Vero cells were transfected using Metafectin reagent (Biontex).
Antibodies and indirect immunofluorescence assay (IFA).
Mouse monoclonal antibody (mAb) 8131, which detects epitopes present in both IE1 and IE2 (exons 2 and 3), was obtained from Chemicon. Anti-HA rat mAb (3F10), either conjugated with peroxidase or labelled with fluorescein, and anti-myc mouse mAb 9E10 were purchased from Roche. Anti-FLAG mouse mAb M2 was obtained from Sigma. Rabbit anti-PML polyclonal Ab (pAb), referred to as PML(C), and anti-SUMO pAb have been described previously (Ahn et al., 2001).
For the IFA, cells were fixed with 1 % paraformaldehyde and permeabilized with 0.2 % Triton X-100. All of the subsequently performed procedures have been described previously (Lee et al., 2004). All slides were examined and photographed using a Zeiss Axiophot microscope.
Immunoblot analysis.
Cells were washed with PBS and total extracts were prepared via incubation of the cells with RIPA buffer [50 mM Tris/HCl (pH 8.0), 150 mM NaCl, 1.0 % NP-40, 0.5 % sodium deoxycholate, 0.1 % SDS) containing protease cocktail. For the sumoylation assays, samples were prepared via direct boiling in SDS loading buffer. The clarified cell extracts were then separated by 8 % SDS-PAGE, followed by a standard procedure using an enhanced chemiluminescence system (Amersham).
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RESULTS |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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). These findings indicated that both PML and IE1 could be modified efficiently by SUMO-1 in vitro, in an E3-independent manner.
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). This suggested that IE1 neither interferes with PML sumoylation, nor directly causes PML desumoylation. However, this result does not preclude the possibility that the presence of excess amounts of E1 and E2 enzymes for sumoylation renders it difficult to detect IE1-associated desumoylation activity.
SENP1 desumoylates both PML and IE1 in vitro: the use of an E. coli sumoylation system to characterize desumoylation activity
To address more directly the question of whether IE1 itself exerts a desumoylation effect on PML, we employed an improved assay system for the study of in vitro desumoylation activity. Recently, a binary vector system was established, which introduces a synthetic SUMO modification system into E. coli (Uchimura et al., 2004a, b). In this system, E. coli is transformed with two plasmids, one encoding the E1 and E2 enzymes for sumoylation as well as an active form of SUMO-1 (or SUMO-2) and the other encoding a substrate (Fig. 2a). With this system, we generated both unmodified and SUMO-1-modified forms of both GST–IE1 and GST–PML VI in E. coli and purified them. As controls, the wild-type and catalytically inactive C603S mutant forms of SENP1, a SUMO protease, were also purified from E. coli as GST fusion forms. When the SUMO-1-modified forms of GST–IE1 or GST–PML VI were incubated with SENP1, the wild-type SENP1 was found to completely desumoylate both IE1 and PML VI (Fig. 2b, lanes 2 and 5), whereas the C603S mutant did not (Fig. 2b, lanes 3 and 6). This result clearly showed that SENP1 is capable of desumoylating both PML VI and IE1 in vitro. Thus, the use of bacterially generated sumoylated substrate proteins and SUMO proteases provides an ideal experimental system for the study of desumoylation activity under cell- and SUMO conjugation enzyme-free conditions.
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, top and middle panels). To exclude potential problems inherent to the different post-translational modification systems used in E. coli compared with eukaryotic cells, we also utilized GST–IE1 generated in insect cells infected with a recombinant baculovirus that expressed GST–IE1. However, even this eukaryotic version of GST–IE1 did not desumoylate PML (Fig. 3a, bottom panel). Moreover, when eightfold higher amounts (6.4 pmole) of bacterially purified GST–SENP1 or GST–IE1 were employed, GST–SENP1 desumoylated PML almost completely within 10 min, whereas GST–IE1 did not do so even after 80 min (Fig. 3b). The use of comparable amounts of GST–SENP1 and GST–IE1 proteins in these assays was confirmed by Coomassie blue staining (Fig. 3c). Collectively, our results demonstrated that neither IE1 protein generated from E. coli nor IE1 generated from insect cells exhibits intrinsic PML-desumoylating properties in vitro.
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). This result showed that, unlike ICP0, IE1 does not require lysine 160 to induce PML desumoylation and is consistent with the idea that HCMV and HSV-1 may exploit different mechanisms to reduce the levels of sumoylated PML.
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). These results also suggested that IE1-induced PML desumoylation is not PML isoform specific.
We then conducted a comparison of the effects of ectopic expression of IE1 and SUMO proteases, including SENP1 and SuPr-1, on the formation of PML foci. When Vero cells were transfected with SENP1, we observed that the number of PML foci decreased significantly. However, these few remnant PML foci became larger, exhibited brighter PML signals and were co-localized with SENP1 (Fig. 5a–c). Consistent with a previous observation (Nefkens et al., 2003), these novel PML foci (unlike standard PODs) also did not contain detectable levels of SUMO-1 by IFA (Fig. 5d–f), which suggests that the PML foci found in the SENP1-transfected cells were comprised of non-sumoylated PML proteins. However, when the cells were transfected with both SENP1 and IE1, all PML foci were completely disrupted and both SENP1 and IE1 proteins were diffusely localized (Fig. 5g–l). We also observed a similar effect of SuPr-1 expression on PML (Fig. 5m–o) and SUMO-1 signals (data not shown), except that IE1 had no effect on SuPr-1 localization (Fig. 5p–r). These results suggested that, unlike SENP1 and SuPr-1, IE1 is still able to disrupt even those PML foci that are composed of non-sumoylated forms of PML.
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). A similar mutant PML-3M protein has been demonstrated to retain its ability to target to the nuclear matrix and to become localized as foci in PML–/– cells (Lallemand-Breitenbach et al., 2001), thereby indicating that the sumoylation-deficient PML is nevertheless able to form at least a limited number of POD-like aggregates via protein dimerization and oligomerization. In accordance with these observations, when Vero cells were transfected with the mutant PML-3M protein, this protein also became localized in nuclear foci (Fig. 6a–c). When the cells were co-transfected with PML-3M and SENP1, these PML foci were not affected by SENP1 (Fig. 6d–f). However, when the cells were co-transfected with PML-3M and IE1, all PML foci were disrupted by IE1 and both the PML-3M and IE1 proteins became diffusely localized within the nucleus (Fig. 6g–i). This result indicated that, unlike SENP1, IE1 also interferes with the formation of PML aggregates generated in a SUMO-independent manner.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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; Lallemand-Breitenbach et al., 2001; Zhong et al., 2000a), it has been suggested that IE1 might disrupt PODs via the induction of PML desumoylation. This suggestion is bolstered by our previous demonstration that IE1 is responsible for the induction of PML desumoylation in virus-infected cells and that the ability of IE1 mutants to desumoylate PML is, indeed, associated with their ability to disrupt PODs (Lee et al., 2004). However, the direct investigation of this IE1 activity has been impaired by an inability to purify the mature PODs or the sumoylated forms of PML, primarily because of the activity of desumoylating enzymes. In this regard, the use of an E. coli sumoylation system for the purification of sumoylated forms of PML allowed us to establish ideal experimental conditions in which the activity of IE1 against sumoylated forms of PML in vitro could be evaluated. In this study, we have shown that IE1 exhibits neither an inhibitory effect on sumoylation of PML nor desumoylating activity against PML in vitro and that IE1 can disrupt even SUMO-independent PML aggregates.
These findings provided useful information with regard to models to explain the ability of IE1 to induce POD disruption and PML desumoylation. Previously, we proposed several mechanisms that might be responsible for IE1-induced PML desumoylation (Lee et al., 2004). First, IE1 may prevent the formation of sumoylated forms of PML, either by competing with PML for SUMO or via the direct inhibition of PML sumoylation by conjugation enzymes. In particular, the fact that IE1 is also a SUMO substrate (Nevels et al., 2004; Sadanari et al., 2005; Spengler et al., 2002; Xu et al., 2001) appeared to support the SUMO competition hypothesis. However, this scenario now seems unlikely for HCMV. Previously, the characterization of two mutant IE1 proteins indicated that a sumoylation-deficient mutant IE1(K450R) protein still retained its ability to disrupt PODs (and desumoylate PML) (Lee et al., 2004; Nevels et al., 2004; Spengler et al., 2002) and that a deleted IE1(
290–320) protein could be sumoylated as efficiently as the wild-type protein, but did not retain the ability to induce PML desumoylation (Lee et al., 2004). The data presented in this study demonstrate conclusively that purified IE1 does not interfere with PML desumoylation in in vitro sumoylation reactions.
IE1 has also been suggested to actively promote the removal of SUMO from PML, either via its own direct intrinsic SUMO protease activity or by the ability to recruit SUMO proteases to PODs indirectly, thereby facilitating PML desumoylation. Importantly, the use of the recently developed E. coli sumoylation system enabled us to generate target sumoylated proteins without contamination from SUMO protease activity, which is absent in E. coli. By using the purified sumoylated substrates, we were able to prevent contamination from excess SUMO modification components, which can be problematic in assays for desumoylating activity. Using this assay system, we determined that bacterially purified SENP1 efficiently desumoylates PML, whereas IE1, whether generated from E. coli or from insect cells, causes no PML desumoylation. As the PML protein used in this assay was not produced in eukaryotic cells, it cannot be excluded that modification of PML, such as phosphorylation, contributes to the induction of desumoylation by IE1. However, our results constitute clear evidence that IE1 itself does not possess a SUMO protease activity against bacterially produced PML in vitro.
The notion that IE1 may induce PML desumoylation indirectly via the recruitment of SUMO proteases to PODs originates with the observation that HSV-1 ICP0 co-localizes with SENP1 in HSV-1-infected cells and promotes the recruitment of SENP1 to PODs in co-transfection experiments, although this study did not address whether or not ICP0 interacted with SENP1 (Bailey & O'Hare, 2002). We were not able to demonstrate an interaction between IE1 and SUMO proteases, such as SENP1, Axam and SuPr-1, in either in vitro- or in vivo-binding assays (data not shown). Nevertheless, it remains possible that IE1 may interact with other SUMO proteases or that these interactions are too transient to be detected. However, several lines of experimental evidence now strongly suggest that HCMV IE1 employs a different mechanism to disrupt PODs than HSV-1 ICP0. Unlike the reaction observed with HSV-1 ICP0, the disruption of PODs in HCMV-infected cells occurs in a proteasome-independent manner (Xu et al., 2001). IE1 effected a reduction of the sumoylated forms of PML, but did not alter the levels of unmodified PML, whereas ICP0 caused a loss of both the sumoylated and the unmodified forms of PML (Boutell et al., 2003; Lee et al., 2004). Furthermore, as was demonstrated in this study, unlike ICP0-induced PML degradation (Boutell et al., 2003), IE1-induced PML desumoylation does not require PML lysine 160.
Finally, it is also possible that IE1 simply induces the disaggregation of the PODs (presumably through physical binding to PML), with subsequent exposure of sumoylated PML to cellular SUMO proteases. This idea is generally supported by the data in the present study. In our transfection assays, IE1 disrupted both the remaining endogenous SUMO-independent PML foci generated after transfection of SUMO proteases and the exogenous PML foci generated by transfection of the sumoylation-deficient mutant PML protein. This clearly shows that IE1 can interfere with the formation of PML aggregates, even in a SUMO-independent manner. We previously demonstrated that IE1 interacts with the N-terminal region of PML harbouring the RING-finger domain (Ahn et al., 1998). Considering that the RING-finger domain of PML is involved with both protein oligomerization and the formation of supramolecular structures (Kentsis et al., 2002), we propose that IE1 binding to PML interferes with both PML oligomerization and the formation of supramolecular structures. This activity of IE1 may play an important role in triggering the disassembly of PODs in a SUMO-independent manner and in inducing the desumoylation of PML by SUMO proteases.
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ACKNOWLEDGEMENTS |
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REFERENCES |
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Ahn, J.-H. & Hayward, G. S. (1997). The major immediate-early proteins IE1 and IE2 of human cytomegalovirus colocalize with and disrupt PML-associated nuclear bodies at very early times in infected permissive cells. J Virol 71, 4599–4613.[Abstract]
Ahn, J.-H. & Hayward, G. S. (2000). Disruption of PML-associated nuclear bodies by IE1 correlates with efficient early stages of viral gene expression and DNA replication in human cytomegalovirus infection. Virology 274, 39–55.[CrossRef][Medline]
Ahn, J.-H., Brignole, E. J., III & Hayward, G. S. (1998). Disruption of PML subnuclear domains by the acidic IE1 protein of human cytomegalovirus is mediated through interaction with PML and may modulate a RING finger-dependent cryptic transactivator function of PML. Mol Cell Biol 18, 4899–4913.
Ahn, J. H., Xu, Y., Jang, W.-J., Matunis, M. J. & Hayward, G. S. (2001). Evaluation of interactions of human cytomegalovirus immediate-early IE2 regulatory protein with small ubiquitin-like modifiers and their conjugation enzyme Ubc9. J Virol 75, 3859–3872.
Bailey, D. & O'Hare, P. (2002). Herpes simplex virus 1 ICP0 co-localizes with a SUMO-specific protease. J Gen Virol 83, 2951–2964.
Bailey, D. & O'Hare, P. (2004). Characterization of the localization and proteolytic activity of the SUMO-specific protease, SENP1. J Biol Chem 279, 692–703.
Bell, P., Lieberman, P. M. & Maul, G. G. (2000). Lytic but not latent replication of Epstein–Barr virus is associated with PML and induces sequential release of nuclear domain 10 proteins. J Virol 74, 11800–11810.
Bernardi, R. & Pandolfi, P. P. (2003). Role of PML and the PML-nuclear body in the control of programmed cell death. Oncogene 22, 9048–9057.[CrossRef][Medline]
Boutell, C., Orr, A. & Everett, R. D. (2003). PML residue lysine 160 is required for the degradation of PML induced by herpes simplex virus type 1 regulatory protein ICP0. J Virol 77, 8686–8694.
Dellaire, G. & Bazett-Jones, D. P. (2004). PML nuclear bodies: dynamic sensors of DNA damage and cellular stress. Bioessays 26, 963–977.[CrossRef][Medline]
Everett, R. D. & Maul, G. G. (1994). HSV-1 IE protein Vmw110 causes redistribution of PML. EMBO J 13, 5062–5069.[Medline]
Gawn, J. M. & Greaves, R. F. (2002). Absence of IE1 p72 protein function during low-multiplicity infection by human cytomegalovirus results in a broad block to viral delayed-early gene expression. J Virol 76, 4441–4455.
Gravel, A., Gosselin, J. & Flamand, L. (2002). Human herpesvirus 6 immediate-early 1 protein is a sumoylated nuclear phosphoprotein colocalizing with promyelocytic leukemia protein-associated nuclear bodies. J Biol Chem 277, 19679–19687.
Greaves, R. F. & Mocarski, E. S. (1998). Defective growth correlates with reduced accumulation of a viral DNA replication protein after low-multiplicity infection by a human cytomegalovirus ie1 mutant. J Virol 72, 366–379.
Green, S., Isseman, P. & Sheer, E. (1988). A versatile in vivo and in vitro eukaryotic expression vector for protein engineering. Nucleic Acids Res 16, 369.
Ishov, A. M., Sotnikov, A. G., Negorev, D., Vladimirova, O. V., Neff, N., Kamitani, T., Yeh, E. T. H., Strauss, J. F., III & Maul, G. G. (1999). PML is critical for ND10 formation and recruits the PML-interacting protein Daxx to this nuclear structure when modified by SUMO-1. J Cell Biol 147, 221–234.
Johnson, E. S. (2004). Protein modification by SUMO. Annu Rev Biochem 73, 355–382.[CrossRef][Medline]
Kamitani, T., Kito, K., Nguyen, H. P., Wada, H., Fukuda-Kamitani, T. & Yeh, E. T. H. (1998). Identification of three major sentrinization sites in PML. J Biol Chem 273, 26675–26682.
Kentsis, A., Gordon, R. E. & Borden, K. L. (2002). Control of biochemical reactions through supramolecular RING domain self-assembly. Proc Natl Acad Sci U S A 99, 15404–15409.
Korioth, F., Maul, G. G., Plachter, B., Stamminger, T. & Frey, J. (1996). The nuclear domain 10 (ND10) is disrupted by the human cytomegalovirus gene product IE1. Exp Cell Res 229, 155–158.[CrossRef][Medline]
Lallemand-Breitenbach, V., Zhu, J., Puvion, F. & 8 other authors (2001). Role of promyelocytic leukemia (PML) sumolation in nuclear body formation, 11S proteasome recruitment, and As2O3-induced PML or PML/retinoic acid receptor 
degradation. J Exp Med 193, 1361–1371.
Lee, H.-R., Kim, D.-J., Lee, J.-M., Choi, C. Y., Ahn, B.-Y., Hayward, G. S. & Ahn, J.-H. (2004). Ability of the human cytomegalovirus IE1 protein to modulate sumoylation of PML correlates with its functional activities in transcriptional regulation and infectivity in cultured fibroblast cells. J Virol 78, 6527–6542.
Maul, G. G. & Everett, R. D. (1994). The nuclear location of PML, a cellular member of the C3HC4 zinc-binding domain protein family, is rearranged during herpes simplex virus infection by the C3HC4 viral protein ICP0. J Gen Virol 75, 1223–1233.
Maul, G. G., Guldner, H. H. & Spivack, J. G. (1993). Modification of discrete nuclear domains induced by herpes simplex virus type 1 immediate early gene 1 product (ICP0). J Gen Virol 74, 2679–2690.
Mocarski, E. S. & Tan Courcelle, C. (2001). Cytomegalovirus and their replication. In Fields Virology, 4th edn, pp. 2629–2673. Edited by D. M. Knipe & P. M. Howley. Philadelphia, PA: Lippincott Williams & Wilkins.
Mocarski, E. S., Kemble, G. W., Lyle, J. M. & Greaves, R. F. (1996). A deletion mutant in the human cytomegalovirus gene encoding IE1491aa is replication defective due to a failure in autoregulation. Proc Natl Acad Sci U S A 93, 11321–11326.
Müller, S. & Dejean, A. (1999). Viral immediate-early proteins abrogate the modification by SUMO-1 of PML and Sp100 proteins, correlating with nuclear body disruption. J Virol 73, 5137–5143.
Nefkens, I., Negorev, D. G., Ishov, A. M., Michaelson, J. S., Yeh, E. T. H., Tanguay, R. M., Müller, W. E. G. & Maul, G. G. (2003). Heat shock and Cd2+ exposure regulate PML and Daxx release from ND10 by independent mechanisms that modify the induction of heat-shock proteins 70 and 25 differently. J Cell Sci 116, 513–524.
Nevels, M., Brune, W. & Shenk, T. (2004). SUMOylation of the human cytomegalovirus 72-kilodalton IE1 protein facilitates expression of the 86-kilodalton IE2 protein and promotes viral replication. J Virol 78, 7803–7812.
Pass, R. F. (2001). Cytomegalovirus. In Fields Virology, 4th edn, pp. 2675–2705. Edited by D. M. Knipe & P. M. Howley. Philadelphia, PA: Lippincott Williams & Wilkins.
Regad, T., Saib, A., Lallemand-Breitenbach, V., Pandolfi, P. P., de Thé, H. & Chelbi-Alix, M. K. (2001). PML mediates the interferon-induced antiviral state against a complex retrovirus via its association with the viral transactivator. EMBO J 20, 3495–3505.[CrossRef][Medline]
Roth, M. B., Zahler, A. M. & Stolk, J. A. (1991). A conserved family of nuclear phosphoproteins localized to sites of polymerase II transcription. J Cell Biol 115, 587–596.
Sadanari, H., Yamada, R., Ohnishi, K., Matsubara, K. & Tanaka, J. (2005). SUMO-1 modification of the major immediate-early (IE) 1 and 2 proteins of human cytomegalovirus is regulated by different mechanisms and modulates the intracellular localization of the IE1, but not IE2, protein. Arch Virol 150, 1763–1782.[CrossRef][Medline]
Salomoni, P. & Pandolfi, P. P. (2002). The role of PML in tumor suppression. Cell 108, 165–170.[CrossRef][Medline]
Seeler, J.-S. & Dejean, A. (2003). Nuclear and unclear functions of SUMO. Nat Rev Mol Cell Biol 4, 690–699.[CrossRef][Medline]
Spengler, M. L., Kurapatwinski, K., Black, A. R. & Azizkhan-Clifford, J. (2002). SUMO-1 modification of human cytomegalovirus IE1/IE72. J Virol 76, 2990–2996.
Stanton, R., Fox, J. D., Caswell, R., Sherratt, E. & Wilkinson, G. W. G. (2002). Analysis of the human herpesvirus-6 immediate-early 1 protein. J Gen Virol 83, 2811–2820.
Tang, Q. & Maul, G. G. (2003). Mouse cytomegalovirus immediate-early protein 1 binds with host cell repressors to relieve suppressive effects on viral transcription and replication during lytic infection. J Virol 77, 1357–1367.[Medline]
Tatham, M. H., Jaffray, E., Vaughan, O. A., Desterro, J. M. P., Botting, C. H., Naismith, J. H. & Hay, R. T. (2001). Polymeric chains of SUMO-2 and SUMO-3 are conjugated to protein substrates by SAE1/SAE2 and Ubc9. J Biol Chem 276, 35368–35374.
Uchimura, Y., Nakamura, M., Sugasawa, K., Nakao, M. & Saitoh, H. (2004a). Overproduction of eukaryotic SUMO-1- and SUMO-2-conjugated proteins in Escherichia coli. Anal Biochem 331, 204–206.[Medline]
Uchimura, Y., Nakao, M. & Saitoh, H. (2004b). Generation of SUMO-1 modified proteins in E. coli: towards understanding the biochemistry/structural biology of the SUMO-1 pathway. FEBS Lett 564, 85–90.[CrossRef][Medline]
Wilkinson, G. W., Kelly, C., Sinclair, J. H. & Rickards, C. (1998). Disruption of PML-associated nuclear bodies mediated by the human cytomegalovirus major immediate early gene product. J Gen Virol 79, 1233–1245.[Abstract]
Xu, Y., Ahn, J.-H., Cheng, M., apRhys, C. M., Chiou, C.-J., Zong, J., Matunis, M. J. & Hayward, G. S. (2001). Proteasome-independent disruption of PML oncogenic domains (PODs), but not covalent modification by SUMO-1, is required for human cytomegalovirus immediate-early protein IE1 to inhibit PML-mediated transcriptional repression. J Virol 75, 10683–10695.
Zhong, S., Muller, S., Ronchetti, S., Freemont, P. S., Dejean, A. & Pandolfi, P. P. (2000a). Role of SUMO-1-modified PML in nuclear body formation. Blood 95, 2748–2752.
Zhong, S., Salomoni, P. & Pandolfi, P. P. (2000b). The transcriptional role of PML and the nuclear body. Nat Cell Biol 2, E85–E90.[CrossRef][Medline]
Received 22 December 2005;
accepted 31 March 2006.
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