scholarly journals Nuclear and Nucleolar Localization of Bovine Adenovirus-3 Protein V

2021 ◽  
Vol 11 ◽  
Author(s):  
Xin Zhao ◽  
Suresh K. Tikoo
Keyword(s):  
Amino Acids ◽  
Intracellular Localization ◽  
Protein A ◽  
Specific Protein ◽  
Nucleolar Localization ◽  
Bovine Adenovirus ◽  
Nuclear Localization Signals ◽  
Importin Α ◽  
Infected Cells ◽  
Import Receptor

The L2 region of bovine adenovirus-3 (BAdV-3) encodes a Mastadenovirus genus-specific protein, designated as pV, which is important for the production of progeny viruses. Here, we demonstrate that BAdV-3 pV, expressed as 55 kDa protein, localizes to the nucleus and specifically targets nucleolus of the infected cells. Analysis of deletion mutants of pV suggested that amino acids 81–120, 190–210, and 380–389 act as multiple nuclear localization signals (NLS), which also appear to serve as the binding sites for importin α-3 protein, a member of the importin α/β nuclear import receptor pathway. Moreover, pV amino acids 21–50 and 380–389 appear to act as nucleolar localization signals (NoLs). Interestingly, amino acids 380–389 appear to act both as NLS and as NoLS. The presence of NoLS is essential for the production of infectious progeny virions, as deletion of both NoLs are lethal for the production of infectious BAdV-3. Analysis of mutant BAV.pVd1d3 (isolated in pV completing CRL cells) containing deletion/mutation of both NoLS in non-complementing CRL cells not only revealed the altered intracellular localization of mutant pV but also reduced the expression of some late proteins. However, it does not appear to affect the incorporation of viral proteins, including mutant pV, in BAV.pVd1d3 virions. Further analysis of CsCl purified BAV.pVd1d3 suggested the presence of thermo-labile virions with disrupted capsids, which appear to affect the infectivity of the progeny virions. Our results suggest that pV contains overlapping and non-overlapping NoLS/NLS. Moreover, the presence of both NoLS appear essential for the production of stable and infectious progeny BAV.pVd1d3 virions.

scholarly journals Localization of the adenovirus E1Aa protein, a positive-acting transcriptional factor, in infected cells infected cells.

1983 ◽  
Vol 3 (5) ◽  
pp. 829-838 ◽  
Author(s):  
L T Feldman ◽  
J R Nevins
Keyword(s):  
Amino Acids ◽  
Mechanism Of Action ◽  
Hela Cells ◽  
Protein A ◽  
Keyhole Limpet Hemocyanin ◽  
Protein Product ◽  
Transcriptional Factor ◽  
Cellular Structures ◽  
Infected Cells

The function of the adenovirus E1Aa protein (the product of the 13S E1A mRNA) during a productive viral infection is to activate transcription of the six early viral transcription units. To study the mechanism of action of this protein, a peptide which was 13 amino acids long and had a sequence unique to the protein product of the adenovirus 13S E1A mRNA (pE1Aa) was coupled to keyhole limpet hemocyanin and used to raise an antibody in rabbits. The resulting antiserum was specific to this protein and did not react with the protein product of the 12S E1A mRNA, which shares considerable sequence with the E1Aa protein. This antiserum was used to probe for the E1Aa protein in situ by indirect immunofluorescence and in extracts of infected HeLa cells. We found that the protein was associated with large cellular structures both in the nucleus and in the cytoplasm. The nuclear form of the protein was analyzed further and was found to purify with the nuclear matrix.

scholarly journals Importin 7 and Importin α/Importin β Are Nuclear Import Receptors for the Glucocorticoid Receptor

2004 ◽  
Vol 15 (5) ◽  
pp. 2276-2286 ◽  
Author(s):  
Neal D. Freedman ◽  
Keith R. Yamamoto
Keyword(s):  
Glucocorticoid Receptor ◽  
Nuclear Import ◽  
Hormonal Control ◽  
Nuclear Localization Signals ◽  
Cell Extracts ◽  
Importin Α ◽  
Import Receptors ◽  
Unidentified Component ◽  
Import Receptor

The vertebrate glucocorticoid receptor (GR) is cytoplasmic without hormone and localizes to the nucleus after hormone binding. GR has two nuclear localization signals (NLS): NL1 is similar in sequence to the SV40 NLS; NL2 is poorly defined, residing in the ligand-binding domain. We found that GR displayed similar hormone-regulated compartmentalization in Saccharomyces cerevisiae and required the Sxm1 nuclear import receptor for NL2-mediated import. Two metazoan homologues of Sxm1, importin 7 and importin 8, bound both NL1 and NL2, whereas importin α selectively bound NL1. In an in vitro nuclear import assay, both importin 7 and the importin α-importin β heterodimer could import a GR NL1 fragment. Under these conditions, full-length GR localized to nuclei in the presence but not absence of an unidentified component in cell extracts. Interestingly, importin 7, importin 8, and importin α bound GR even in the absence of hormone; thus, hormonal control of localization is exerted at a step downstream of import receptor binding.

scholarly journals Two nuclear localization signals regulate intracellular localization of the Duck enteritis virus UL13 protein

2020 ◽  
Author(s):  
Linjiang Yang ◽  
Xixia Hu ◽  
Anchun Cheng ◽  
Mingshu Wang ◽  
Renyong Jia ◽  
...  
Keyword(s):  
Nuclear Localization ◽  
Nuclear Import ◽  
Intracellular Localization ◽  
Duck Enteritis Virus ◽  
Significant Information ◽  
Nuclear Localization Signals ◽  
Dependent Process ◽  
Importin Α ◽  
Localization Signal ◽  
Enteritis Virus

Abstract Background UL13 multifunctional tegument protein duck enteritis virus (DEV) is predicted as conserved herpesvirus protein kinase (CHPK); however, little is known about its subcellular localization signal. Results In this study, by transfection with two predicted nuclear signals of DEV UL13 fused to enhanced green fluorescent protein (EGFP), two bipartite nuclear localization signals (NLS) were identified. We found that the NLSs block its nuclear import using ivermectin and proved that nuclear localization signal of DEV UL13 is a classical importin α/β-dependent process. And we constructed the DEV UL13 mutant strain, with the NLSs of DEV UL13 deleted, to explore whether it can affect the virus replication Conclusions The DEV pUL13 amino acids 4 to 7 and 90 to 96 was predicted, and proved that this nuclear import occurs via the classical importin α/β-dependent process. We also found NLSs of pUL13 have no effect on DEV replication in cell culture. Our study enhances the understanding of DEV pUL13. Taken together, these results would provide significant information for the biological function of pUL13 during DEV infection.

scholarly journals Vaccinia Virus Protein F12 Associates with Intracellular Enveloped Virions through an Interaction with A36

2008 ◽  
Vol 83 (4) ◽  
pp. 1708-1717 ◽  
Author(s):  
Sara C. Johnston ◽  
Brian M. Ward
Keyword(s):  
Amino Acids ◽  
Vaccinia Virus ◽  
Protein A ◽  
Integral Membrane Protein ◽  
Recombinant Vaccinia Virus ◽  
Virus Protein ◽  
Viral Egress ◽  
Plaque Phenotype ◽  
Infected Cells

ABSTRACT Vaccinia virus is the prototypical member of the family Poxviridae. Three morphologically distinct forms are produced during infection: intracellular mature virions (IMV), intracellular enveloped virions (IEV), and extracellular enveloped virions (EEV). Two viral proteins, F12 and A36, are found exclusively on IEV but not on IMV and EEV. Analysis of membranes from infected cells showed that F12 was only associated with membranes and is not an integral membrane protein. A yeast two-hybrid assay revealed an interaction between amino acids 351 to 458 of F12 and amino acids 91 to 111 of A36. We generated a recombinant vaccinia virus that expresses an F12, which lacks residues 351 to 458. Characterization of this recombinant revealed a small-plaque phenotype and a subsequent defect in virus release similar to a recombinant virus that had F12L deleted. In addition, F12 lacking residues 351 to 458 was unable to associate with membranes in infected cells. These results suggest that F12 associates with IEV through an interaction with A36 and that this interaction is critical for the function of F12 during viral egress.

Localization of the adenovirus E1Aa protein, a positive-acting transcriptional factor, in infected cells infected cells

1983 ◽  
Vol 3 (5) ◽  
pp. 829-838
Author(s):  
L T Feldman ◽  
J R Nevins
Keyword(s):  
Amino Acids ◽  
Mechanism Of Action ◽  
Hela Cells ◽  
Protein A ◽  
Keyhole Limpet Hemocyanin ◽  
Protein Product ◽  
Transcriptional Factor ◽  
Cellular Structures ◽  
Infected Cells

The function of the adenovirus E1Aa protein (the product of the 13S E1A mRNA) during a productive viral infection is to activate transcription of the six early viral transcription units. To study the mechanism of action of this protein, a peptide which was 13 amino acids long and had a sequence unique to the protein product of the adenovirus 13S E1A mRNA (pE1Aa) was coupled to keyhole limpet hemocyanin and used to raise an antibody in rabbits. The resulting antiserum was specific to this protein and did not react with the protein product of the 12S E1A mRNA, which shares considerable sequence with the E1Aa protein. This antiserum was used to probe for the E1Aa protein in situ by indirect immunofluorescence and in extracts of infected HeLa cells. We found that the protein was associated with large cellular structures both in the nucleus and in the cytoplasm. The nuclear form of the protein was analyzed further and was found to purify with the nuclear matrix.

scholarly journals Identification and characterization of complex dual nuclear localization signals in human bocavirus NP1

2013 ◽  
Vol 94 (6) ◽  
pp. 1335-1342 ◽  
Author(s):  
Qian Li ◽  
Zhenfeng Zhang ◽  
Zhenhua Zheng ◽  
Xianliang Ke ◽  
Huanle Luo ◽  
...  
Keyword(s):  
Amino Acids ◽  
Nuclear Localization ◽  
Nuclear Import ◽  
Nuclear Localization Signal ◽  
Human Bocavirus ◽  
Structural Protein ◽  
Nuclear Localization Signals ◽  
Importin Α ◽  
Localization Signal ◽  
Bovine Parvovirus

Human bocavirus (HBoV), closely related to canine minute virus (MVC) and bovine parvovirus (BPV), is a new member of the Bocavirus genus within the Parvoviridae family. The non-structural protein NP1 of HBoV is a nuclear localized protein and plays an important role in DNA replication as well as in the evasion of host innate immunity. In the current study, we provide the first evidence that NP1 possesses a non-classical nuclear localization signal (ncNLS) (amino acids 7–50). Embedded within this ncNLS is a classical bipartite nuclear localization signal (cNLS) (amino acids 14–28), capable of transporting a heterologous cytoplasmic protein β-galactosidase fusion protein (β-gal-EGFP) to the nucleus via the classical importin α/β1-mediated pathway. Amino acids 7–50 containing the cNLS and the ncNLS of NP1 or full-length NP1 interact with importin α1, importin β1 and importin β1Δ, which lacks the importin α binding domain, indicating that the nuclear import of NP1 is through both conventional importin α/β1 heterodimer- and non-classical importinß1-mediated pathways. Given that the arrangement of a cNLS embedded within an ncNLS is unusual in viral proteins, our data together reveal a novel molecular mechanism underlying the nuclear import of HBoV NP1, providing a basis for further understanding its biological function.

scholarly journals Two nuclear localization signals regulate intracellular localization of the Duck enteritis virus UL13 protein

2020 ◽  
Author(s):  
Linjiang Yang ◽  
Xixia Hu ◽  
Anchun Cheng ◽  
Mingshu Wang ◽  
Renyong Jia ◽  
...  
Keyword(s):  
Nuclear Localization ◽  
Fluorescent Protein ◽  
Intracellular Localization ◽  
Duck Enteritis Virus ◽  
Significant Information ◽  
Nuclear Localization Signals ◽  
Dependent Process ◽  
Importin Α ◽  
Localization Signal ◽  
Enteritis Virus

Abstract Background UL13 multifunctional tegument protein of duck enteritis virus (DEV) is predicted as protein kinase (CHPK); however, little is known concerning its subcellular localization signal. Results In this study, by transfection with two predicted nuclear signals of DEV UL13 fused to enhanced green fluorescent protein (EGFP), two bipartite nuclear localization signal (NLS) was identified. We identified the nuclear localization signals (NLSs) that control its nuclear importing using fluorescence assay and proved that nuclear localization of DEV UL13 is a classical importin α/β-dependent process. And we constructed the mutant DEV, with the NLSs of UL13 deleted, to explore whether it will affect the replication of virus particles. Conclusions DEV UL13 protein is directed by amino acids 4 to 7 and 90 to 96, and proved that this nuclear import occurs via the classical importin α/β-dependent process. And Entry nucleus of UL13 protein has no effect on DEV replication in cell culture. Our study enhances the understanding of DEV pUL13. Taken together, these results would provide significant information for the stud y of the biological function of UL13 during DEV infection.

scholarly journals Conserved regions of bovine adenovirus-3 pVIII contain functional domains involved in nuclear localization and packaging in mature infectious virions

2014 ◽  
Vol 95 (8) ◽  
pp. 1743-1754 ◽  
Author(s):  
Lisanework E. Ayalew ◽  
Amit Gaba ◽  
Pankaj Kumar ◽  
Suresh K. Tikoo
Keyword(s):  
Capsid Protein ◽  
Nuclear Localization ◽  
Post Infection ◽  
Minor Capsid Protein ◽  
Bovine Adenovirus ◽  
The Core ◽  
C Terminus ◽  
Core Proteins ◽  
Importin Α ◽  
Infected Cells

Adenoviruses are non-enveloped DNA viruses that replicate in the nucleus of infected cells. One of the core proteins, named pVIII, is a minor capsid protein connecting the core with the inner surface of the capsid. Here, we report the characterization of minor capsid protein pVIII encoded by the L6 region of bovine adenovirus (BAdV)-3. Anti-pVIII serum detected a 24 kDa protein at 12–48 h post-infection and an additional 8 kDa protein at 24–48 h post-infection. While the 24 kDa protein was detected in empty capsids, only the C-terminal-cleaved 8 kDa protein was detected in the mature virion, suggesting that amino acids147–216 of the conserved C-terminus of BAdV-3 pVIII are incorporated in mature virions. Detection of hexon protein associated with both precursor (24 kDa) and cleaved (8 kDa) forms of pVIII suggest that the C-terminus of pVIII interacts with the hexon. The pVIII protein predominantly localizes to the nucleus of BAdV-3-infected cells utilizing the classical importin α/β dependent nuclear import pathway. Analysis of mutant pVIII demonstrated that amino acids 52–72 of the conserved N-terminus bind to importin α-3 with high affinity and are required for the nuclear localization.

scholarly journals Importin α/β mediates nuclear import of individual SUMO E1 subunits and of the holo-enzyme

2011 ◽  
Vol 22 (5) ◽  
pp. 652-660 ◽  
Author(s):  
Marie Christine Moutty ◽  
Volkan Sakin ◽  
Frauke Melchior
Keyword(s):  
Amino Acids ◽  
Nuclear Localization ◽  
Nuclear Import ◽  
Enzyme Complex ◽  
Intact Cells ◽  
Nuclear Localization Signals ◽  
Importin Α ◽  
Essential Enzyme ◽  
Nuclear Targets

SUMOylation, reversible attachment of small ubiquitin-related modifier (SUMO), serves to regulate hundreds of proteins. Consistent with predominantly nuclear targets, enzymes required for attachment and removal of SUMO are highly enriched in this compartment. This is true also for the first enzyme of the SUMOylation cascade, the SUMO E1 enzyme heterodimer, Aos1/Uba2 (SAE1/SAE2). This essential enzyme serves to activate SUMO and to transfer it to the E2-conjugating enzyme Ubc9. Although the last 40 amino acids in yeast Uba2 have been implicated in its nuclear localization, little was known about the import pathways of Aos1, Uba2, and/or of the assembled E1 heterodimer. Here we show that the mammalian E1 subunits can be imported separately, identify nuclear localization signals (NLSs) in Aos1 and in Uba2, and demonstrate that their import is mediated by importin α/β in vitro and in intact cells. Once assembled into a stable heterodimer, the E1 enzyme can still be efficiently imported by importin α/β, due to the Uba2 NLS that is still accessible. These pathways may serve distinct purposes: import of nascent subunits prior to assembly and reimport of stable E1 enzyme complex after mitosis.

An immunocytochemical study of maternal immunoglobulin localization in the suckling pig intestine

1990 ◽  
Vol 48 (3) ◽  
pp. 922-923
Author(s):  
László G. Kömüves ◽  
Donna S. Turner ◽  
Kathy S. McKee ◽  
Buford L. Nichols ◽  
Julian P. Heath
Keyword(s):  
Sodium Ethoxide ◽  
Intracellular Localization ◽  
Protein A ◽  
Tween 20 ◽  
Intestinal Epithelial ◽  
Immunocytochemical Study ◽  
Fish Skin ◽  
Newborn Piglets ◽  
Rabbit Igg ◽  
Fish Skin Gelatin

In this study we used colloidal gold probes to detect the intracellular localization of colostral immunoglobulins in intestinal epithelial cells of newborn piglets.Tissues were obtained from non-suckled newborn and suckled piglets aged between 1 hour to 1 month. Samples were fixed in 2.5 % glutaraldehyde, osmicated and embedded into Spurr’s resin. Thin (80 nm) sections were etched with 5% sodium ethoxide for 5 min, washed and treated with 4 % sodium-m-periodate in distilled water for 30 min. The sections were then first incubated with blocking buffer (2 % BSA, 0.25 % fish skin gelatin, 0.5 % Tween 20 in 10 mM Trizma buffer, pH=7.4 containing 500 mM NaCl) for 30 min followed by the immunoreagents diluted in the same buffer, 1 hr each. For the detection of pig immunoglobulins a rabbit anti-pig IgG antiserum was used followed by goat anti-rabbit IgG-Au10 or protein A-Au15 probes.

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