Nuclear Import of Bovine Papillomavirus Type 1 E1 Protein Is Mediated by Multiple Alpha Importins and Is Negatively Regulated by Phosphorylation near a Nuclear Localization Signal

Bian, Xue Lin; Rosas-Acosta, Germán; Wu, Yu Chieh; Wilson, Van G.

doi:10.1128/jvi.01850-06

Cited by 24 publications

(31 citation statements)

References 70 publications

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“…Mutagenesis of C. jejuni UNGM-The R59H, R168K, and R59H/R168K mutants were prepared using the QuikChange XL II protocol by Stratagene (51). In the case of the R59H and R168K, mutants were prepared from the pWM1007 plasmid containing the glf gene isolated from E. coli DH5␣ cells using R59H_F and R59H_R or R168K_F and R168K_R (supplemental Table S1) as primer pairs, respectively.…”

Section: Methodsmentioning

confidence: 99%

Characterization of a Bifunctional Pyranose-Furanose Mutase from Campylobacter jejuni 11168

Poulin

Nothaft

Hug

et al. 2010

Journal of Biological Chemistry

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UDP-galactopyranose mutases (UGM) are the enzymes responsible for the synthesis of UDP-galactofuranose (UDP-Galf) from UDP-galactopyranose (UDP-Galp). The enzyme, encoded by the glf gene, is present in bacteria, parasites, and fungi that express Galf in their glycoconjugates. Recently, a UGM homologue encoded by the cj1439 gene has been identified in Campylobacter jejuni 11168, an organism possessing no Galf-containing glycoconjugates. However, the capsular polysaccharide from this strain contains a 2-acetamido-2-deoxy-D-galactofuranose (GalfNAc) moiety. Using an in vitro high performance liquid chromatography assay and complementation studies, we characterized the activity of this UGM homologue. The enzyme, which we have renamed UDP-N-acetylgalactopyranose mutase (UNGM), has relaxed specificity and can use either UDP-Gal or UDP-GalNAc as a substrate. Complementation studies of mutase knock-outs in C. jejuni 11168 and Escherichia coli W3110, the latter containing Galf residues in its lipopolysaccharide, demonstrated that the enzyme recognizes both UDP-Gal and UDP-GalNAc in vivo. A homology model of UNGM and site-directed mutagenesis led to the identification of two active site amino acid residues involved in the recognition of the UDP-GalNAc substrate. The specificity of UNGM was characterized using a two-substrate co-incubation assay, which demonstrated, surprisingly, that UDP-Gal is a better substrate than UDP-GalNAc.In nature, hexose sugars are found predominantly in the thermodynamically favored pyranose ring form; however, hexose sugars in the furanose ring form are found in bacteria, fungi, and parasites (1, 2). For example, D-galactofuranose (Galf) 5 is a component in many microbial cell surface oligosaccharides (3,

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Section: Methodsmentioning

confidence: 99%

Characterization of a Bifunctional Pyranose-Furanose Mutase from Campylobacter jejuni 11168

Poulin

Nothaft

Hug

et al. 2010

Journal of Biological Chemistry

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“…The efficiency of nuclear import is often modulated by phosphorylation (for a review, see reference 38) or sumoylation (66). BPV-1 E1 has a bipartite NLS near the amino terminus (41,42), and its nuclear translocation is mediated by importin ␣ (8). Cyclin/cylin-dependent kinases (cdk's) also regulate E1 protein subcellular localization and replication activity.…”

mentioning

confidence: 99%

Mitogen-Activated Protein Kinases Activate the Nuclear Localization Sequence of Human Papillomavirus Type 11 E1 DNA Helicase To Promote Efficient Nuclear Import

Lin

Deng

et al. 2007

J Virol

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Human and animal papillomavirus DNA replicates as multicopy nuclear plasmids. Replication requires two viral proteins, the origin-recognition protein E2 and the replicative DNA helicase E1. Using genetic, biochemical, and immunofluorescence assays, we demonstrated that efficient nuclear import of the human papillomavirus (HPV) type 11 E1 protein depends on a codominant bipartite nuclear localization sequence (NLS) and on phosphorylation of the serine residues S89 and S93 by the mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinase, and c-Jun N-terminal protein kinase. The NLS and the MAPK substrates are located within a 50-amino-acid-long peptide near the amino terminus, previously designated the localization regulatory region (LRR). The downstream NLS overlaps the cyclin-binding motif RRL, which is necessary for phosphorylation by the cyclin-dependent kinases to inactivate a dominant nuclear export sequence, also in the LRR. Alanine mutations of the MAPK substrates significantly impaired nuclear import, whereas phosphomimetic mutations partially restored nuclear import. We further identified two MAPK docking motifs near the C terminus of E1 that are conserved among E1 proteins of many HPVs and bovine papillomavirus type 1. Mutations of these MAPK docking motifs or addition of specific MAPK inhibitors significantly reduced nuclear import. Interestingly, a fraction of the NLS-minus E1 protein was cotransported with the E2 protein into the nucleus and supported transient viral DNA replication. In contrast, E1 proteins mutated in the MAPK docking motifs were completely inactive in transient replication, an indication that additional properties were adversely affected by those changes.Infections by human papillomaviruses (HPVs) can cause benign, hyperproliferative lesions of cutaneous or mucosal epithelium. The virus has a double-stranded circular DNA genome approximately 7,900 bp in length, which replicates as extrachromosomal nuclear plasmids. A low copy number of the viral DNA is maintained in the cycling basal and parabasal keratinocytes of squamous epithelium. Viral DNA amplification to produce progeny virions occurs only in postmitotic, suprabasal cells undergoing terminal differentiation (for a review, see reference 15). Initiation of replication from the origin (ori) of various HPV genotypes and bovine papillomavirus type 1 (BPV-1) depends on the virus-encoded ori binding protein E2 and the replicative DNA helicase E1 (for reviews, see references 16 and 63). The ori consists of several E2 protein binding sites flanking a cluster of E1 protein binding sites. The structures and functions of the E1 and E2 proteins of human and animal papillomaviruses are largely conserved, but significant differences are also noted. In brief, the 42-kDa E2 protein binds as dimers to the palindromic ori sequences, ACCGNNNNCGGT, and recruits the 70-kDa E1 protein via an interaction between the carboxyl terminus of E1 and the amino terminus of E2 (16). E1 then assembles into a dihexameric helicase (28,44...

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“…Previous work from our laboratory has demonstrated that E1 is SUMOylated, that PIAS proteins can serve as SUMO ligases for this reaction and that SUMOylation of E1 is necessary for proper nuclear localization [9,10]. The mechanism by which SUMOylation affects E1 nuclear localization is still unclear as SUMOylation does not appear to affect nuclear import [11] or export (G. Rosas-Acosta and V.G. Wilson, unpublished work) of E1 directly.…”

Section: Papillomavirus and Sumoylationmentioning

confidence: 98%

Papillomaviruses and the host SUMOylation system

Deyrieux

Wilson

2007

Biochemical Society Transactions

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SUMOylation of viral proteins is widespread and serves to modify or regulate the properties of those proteins. Papillomaviruses are a large group of small DNA viruses that infect the skin, leading to benign lesions (warts) that in some cases can progress to malignancy. The papillomavirus life cycle is intimately connected with the differentiation process of stratified epithelium, and several viral early proteins function to modulate the host cell environment. One of the critical early proteins is the E2 protein, which functions in both viral replication and transcription. In the present paper, we demonstrate that E2 proteins are SUMOylated and that overexpression of SUMOylation results in a dramatic increase in intracellular levels of the E2 protein. We have shown previously that there is increased SUMOylation during keratinocyte differentiation, suggesting that the levels of E2 protein may be tied to changes in the cellular SUMOylation state during differentiation. In addition to itself being regulated by SUMOylation, E2 appears to influence the SUMOylation state of one of its binding partners, the cellular transcription factor, C/EBP (CCAAT/enhancer-binding protein). Overall, these observations indicate a complex interplay between this viral protein and the host SUMOylation system.

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Nuclear Import of Bovine Papillomavirus Type 1 E1 Protein Is Mediated by Multiple Alpha Importins and Is Negatively Regulated by Phosphorylation near a Nuclear Localization Signal

Cited by 24 publications

References 70 publications

Characterization of a Bifunctional Pyranose-Furanose Mutase from Campylobacter jejuni 11168

Characterization of a Bifunctional Pyranose-Furanose Mutase from Campylobacter jejuni 11168

Mitogen-Activated Protein Kinases Activate the Nuclear Localization Sequence of Human Papillomavirus Type 11 E1 DNA Helicase To Promote Efficient Nuclear Import

Papillomaviruses and the host SUMOylation system

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