scholarly journals Mass Spectrometric Analyses of Purified Rhesus Monkey Rhadinovirus Reveal 33 Virion-Associated Proteins

2006 ◽  
Vol 80 (3) ◽  
pp. 1574-1583 ◽  
Author(s):  
Christine M. O'Connor ◽  
Dean H. Kedes
Keyword(s):  
Rhesus Monkey ◽  
High Titer ◽  
Mass Spectrometric ◽  
Structural Proteins ◽  
Human Pathogen ◽  
Tegument Proteins ◽  
New Findings ◽  
Viral Structure ◽  
Detergent Extraction ◽  
Associated Proteins

ABSTRACT The repertoire of proteins that comprise intact gammaherpesviruses, including the human pathogen Kaposi's sarcoma-associated herpesvirus (KSHV), is likely to have critical functions not only in viral structure and assembly but also in the early stages of infection and evasion of the host's rapidly deployed antiviral defenses. To develop a better understanding of these proteins, we analyzed the composition of rhesus monkey rhadinovirus (RRV), a close phylogenetic relative of KSHV. Unlike KSHV, RRV replicates to high titer in cell culture and thus serves as an effective model for studying primate gammaherpesvirus structure and virion proteomics. We employed two complementary mass spectrometric approaches and found that RRV contains at least 33 distinct virally encoded proteins. We have assigned 7 of these proteins to the capsid, 17 to the tegument, and 9 to the envelope. Of the five gammaherpesvirus-specific tegument proteins, three have no known function. We also found three proteins not previously associated with a purified herpesvirus and an additional seven that represent new findings for a member of the gamma-2 herpesviruses. Detergent extraction resulted in particles that contained six distinct tegument proteins in addition to the expected capsid structural proteins, suggesting that this subset of tegument components may interact more directly with or with higher affinity for the underlying capsid and, in turn, may play a role in assembly or transport of viral or subviral particles during entry or egress.

scholarly journals The Annexin A2/S100A10 System in Health and Disease: Emerging Paradigms

2012 ◽  
Vol 2012 ◽  
pp. 1-13 ◽  
Author(s):  
Nadia Hedhli ◽  
Domenick J. Falcone ◽  
Bihui Huang ◽  
Gabriela Cesarman-Maus ◽  
Rosemary Kraemer ◽  
...  
Keyword(s):  
Serotonin Receptor ◽  
Human Cancer ◽  
High Titer ◽  
Critical Role ◽  
Annexin A2 ◽  
Avascular Osteonecrosis ◽  
Kinase Substrate ◽  
New Findings ◽  
Human Disorders ◽  
Associated Proteins

Since its discovery as a src kinase substrate more than three decades ago, appreciation for the physiologic functions of annexin A2 and its associated proteins has increased dramatically. With its binding partner S100A10 (p11), A2 forms a cell surface complex that regulates generation of the primary fibrinolytic protease, plasmin, and is dynamically regulated in settings of hemostasis and thrombosis. In addition, the complex is transcriptionally upregulated in hypoxia and promotes pathologic neoangiogenesis in the tissues such as the retina. Dysregulation of both A2 and p11 has been reported in examples of rodent and human cancer. Intracellularly, A2 plays a critical role in endosomal repair in postarthroplastic osteolysis, and intracellular p11 regulates serotonin receptor activity in psychiatric mood disorders. In human studies, the A2 system contributes to the coagulopathy of acute promyelocytic leukemia, and is a target of high-titer autoantibodies in patients with antiphospholipid syndrome, cerebral thrombosis, and possibly preeclampsia. Polymorphisms in the humanANXA2gene have been associated with stroke and avascular osteonecrosis of bone, two severe complications of sickle cell disease. Together, these new findings suggest that manipulation of the annexin A2/S100A10 system may offer promising new avenues for treatment of a spectrum of human disorders.

scholarly journals DeNovo Infection with Rhesus Monkey Rhadinovirus Leads to theAccumulation of Multiple Intranuclear Capsid Species during LyticReplication but Favors the Release of Genome-ContainingVirions

2003 ◽  
Vol 77 (24) ◽  
pp. 13439-13447 ◽  
Author(s):  
Christine M. O'Connor ◽  
Blossom Damania ◽  
Dean H. Kedes
Keyword(s):  
Rhesus Monkey ◽  
B Cell ◽  
Cell Lines ◽  
De Novo ◽  
Lytic Replication ◽  
Lytic Infection ◽  
Structural Proteins ◽  
Human Pathogen ◽  
Genes Encoding ◽  
Latently Infected

ABSTRACT Rhesus monkey rhadinovirus (RRV) is one of the closest phylogenetic relatives to the human pathogen Kaposi's sarcoma-associated herpesvirus (KSHV), yet it has the distinct experimental advantage of entering efficiently into lytic replication and growing to high titers in culture. RRV therefore holds promise as a potentially attractive model with which to study gammaherpesvirus structure and assembly. We have isolated RRV capsids, determined their molecular composition, and identified the genes encoding five of the main capsid structural proteins. Our data indicate that, as with other herpesviruses, lytic infection with RRV leads to the synthesis of three distinct intranuclear capsid species. However, in contrast to the inefficiency of KSHV maturation following reactivation from latently infected B-cell lines (K. Nealon, W. W. Newcomb, T. R. Pray, C. S. Craik, J. C. Brown, and D. H. Kedes, J. Virol. 75:2866-2878, 2001), de novo infection of immortalized rhesus fibroblasts with RRV results in the release of high levels of infectious virions with genome-containing C capsids at their center. Together, our findings argue for the use of RRV as a powerful model with which to study the structure and assembly of gammaherpesviruses and, specifically, the human rhadinovirus,KSHV.

scholarly journals Non-Proteasomal UbL-UbA Family of Proteins in Neurodegeneration

2019 ◽  
Vol 20 (8) ◽  
pp. 1893 ◽  
Author(s):  
Salinee Jantrapirom ◽  
Luca Lo Piccolo ◽  
Masamitsu Yamaguchi
Keyword(s):  
Neurodegenerative Diseases ◽  
Family Members ◽  
Proteasomal Activity ◽  
New Findings ◽  
Evolutionarily Conserved ◽  
Associated Proteins ◽  
Ubiquitin Receptors ◽  
Drosophila Models

Ubiquitin-like/ubiquitin-associated proteins (UbL-UbA) are a well-studied family of non-proteasomal ubiquitin receptors that are evolutionarily conserved across species. Members of this non-homogenous family facilitate and support proteasomal activity by promoting different effects on proteostasis but exhibit diverse extra-proteasomal activities. Dysfunctional UbL-UbA proteins render cells, particularly neurons, more susceptible to stressors or aging and may cause earlier neurodegeneration. In this review, we summarized the properties and functions of UbL-UbA family members identified to date, with an emphasis on new findings obtained using Drosophila models showing a direct or indirect role in some neurodegenerative diseases.

scholarly journals Identification of the Nucleocapsid, Tegument, and Envelope Proteins of the Shrimp White Spot Syndrome Virus Virion

2006 ◽  
Vol 80 (6) ◽  
pp. 3021-3029 ◽  
Author(s):  
Jyh-Ming Tsai ◽  
Han-Ching Wang ◽  
Jiann-Horng Leu ◽  
Andrew H.-J. Wang ◽  
Ying Zhuang ◽  
...  
Keyword(s):  
White Spot Syndrome Virus ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Envelope Proteins ◽  
White Spot ◽  
Structural Proteins ◽  
Nucleocapsid Proteins ◽  
Tegument Proteins ◽  
White Spot Syndrome ◽  
Protein Components

ABSTRACT The protein components of the white spot syndrome virus (WSSV) virion have been well established by proteomic methods, and at least 39 structural proteins are currently known. However, several details of the virus structure and assembly remain controversial, including the role of one of the major structural proteins, VP26. In this study, Triton X-100 was used in combination with various concentrations of NaCl to separate intact WSSV virions into distinct fractions such that each fraction contained envelope and tegument proteins, tegument and nucleocapsid proteins, or nucleocapsid proteins only. From the protein profiles and Western blotting results, VP26, VP36A, VP39A, and VP95 were all identified as tegument proteins distinct from the envelope proteins (VP19, VP28, VP31, VP36B, VP38A, VP51B, VP53A) and nucleocapsid proteins (VP664, VP51C, VP60B, VP15). We also found that VP15 dissociated from the nucleocapsid at high salt concentrations, even though DNA was still present. These results were confirmed by CsCl isopycnic centrifugation followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and liquid chromatography-nanoelectrospray ionization-tandem mass spectrometry, by a trypsin sensitivity assay, and by an immunogold assay. Finally, we propose an assembly process for the WSSV virion.

scholarly journals Cross-Dressing the Virion: the Transcapsidation of Adeno-Associated Virus Serotypes Functionally Defines Subgroups

2004 ◽  
Vol 78 (9) ◽  
pp. 4421-4432 ◽  
Author(s):  
Joseph E. Rabinowitz ◽  
Dawn E. Bowles ◽  
Susan M. Faust ◽  
Julie G. Ledford ◽  
Scott E. Cunningham ◽  
...  
Keyword(s):  
Viral Entry ◽  
Fluorescent Protein ◽  
High Titer ◽  
Threshold Effect ◽  
Structural Proteins ◽  
Adeno Associated Virus ◽  
Serotype 1 ◽  
Cross Dressing ◽  
Helper Plasmids ◽  
Green Fluorescent

ABSTRACT For all adeno-associated virus (AAV) serotypes, 60 monomers of the Vp1, Vp2, and Vp3 structural proteins assemble via an unknown mechanism to form an intact capsid. In an effort to better understand the properties of the capsid monomers and their role in viral entry and infection, we evaluated whether monomers from distinct serotypes can be mixed to form infectious particles with unique phenotypes. This transcapsidation approach consisted of the transfection of pairwise combinations of AAV serotype 1 to 5 helper plasmids to produce mosaic capsid recombinant AAV (rAAV). All ratios (19:1, 3:1, 1:1, 1:3, and 1:19) of these mixtures were able to replicate the green fluorescent protein transgene and to produce capsid proteins. A high-titer rAAV was obtained with mixtures that included either serotype 1, 2, or 3, whereas an rAAV of intermediate titer was obtained from serotype 5 mixtures. Only mixtures containing the AAV4 capsid exhibited reduced packaging capacity. The binding profiles of the mixed-virus preparations to either heparin sulfate (HS) or mucin agarose revealed that only AAV3-AAV5 mixtures at the 3:1 ratio exhibited duality in binding. All other mixtures displayed either an abrupt shift or a gradual alteration in the binding profile to the respective ligand upon increase of a capsid component that conferred either HS or mucin binding. The transduction of cell lines was used to further evaluate the phenotypes of these transcapsidated virions. Three transduction profiles were observed: (i) small to no change regardless of ratio, (ii) a gradual increase in transduction consistent with titration of a second capsid component, or (iii) an abrupt increase in transduction (threshold effect) dependent on the specific ratios used. Interestingly, an unexpected synergistic effect in transduction was observed when AAV1 helper constructs were combined with type 2 or type 3 recipient helpers. Further studies determined that at least two components contributed to this observed synergy: (i) heparin-mediated binding from AAV2 and (ii) an unidentified enhancement activity from AAV1 structural proteins. Using this procedure of mixing different AAV helper plasmids to generate “cross-dressed” AAV virions, we propose an additional means of classifying new AAV serotypes into subgroups based on functional approaches to analyze AAV capsid assembly, receptor-mediated binding, and virus trafficking. Exploitation of this approach in generating custom-designed AAV vectors should be of significant value to the field of gene therapy.

New Approaches And Concepts in The Study of Differentiation of Oral Epithelia

1990 ◽  
Vol 1 (3) ◽  
pp. 167-190 ◽  
Author(s):  
Beverly A. Dale ◽  
Jukka Salonen ◽  
Alma H. Jones
Keyword(s):  
Protein Expression ◽  
Structural Proteins ◽  
Basal Cells ◽  
Critical Factors ◽  
Promoter Regions ◽  
Receptor Proteins ◽  
Specific Manner ◽  
Associated Proteins ◽  
Extracellular Factor ◽  
Oral Epithelia

Epithelial structural proteins, the keratins and keratin-associated proteins, are useful as markers of differentiation because their expression is both region-specific and differentiation-specific. In general, basal cells in all stratified oral epithelia express similar keratins, while the suprabasal cells express a specific set of markers indicating commitment to a distinct program of differentiation. Critical factors in the regulation of epithelial protein expression are now under investigation. The promoter regions of keratin genes are being characterized to determine what sequences within the genes are responsible for differential expression. One important extracellular factor that influences epithelial protein expression is retinol (vitamin A), which exerts its effects via a group of nuclear receptor proteins that may also be expressed in a region-specific manner. These molecular biological approaches enhance our understanding of the mechanisms regulating differentiation of oral epithelia and its regional complexity.

scholarly journals Illuminating structural proteins in viral “dark matter” with metaproteomics

2016 ◽  
Vol 113 (9) ◽  
pp. 2436-2441 ◽  
Author(s):  
Jennifer R. Brum ◽  
J. Cesar Ignacio-Espinoza ◽  
Eun-Hae Kim ◽  
Gareth Trubl ◽  
Robert M. Jones ◽  
...  
Keyword(s):  
Dark Matter ◽  
Structural Proteins ◽  
Protein Fold ◽  
Biological Structure ◽  
Genomic Context ◽  
Environmental Virology ◽  
Culture Independent ◽  
Associated Proteins ◽  
Domains Of Life ◽  
Viral Communities

Viruses are ecologically important, yet environmental virology is limited by dominance of unannotated genomic sequences representing taxonomic and functional “viral dark matter.” Although recent analytical advances are rapidly improving taxonomic annotations, identifying functional dark matter remains problematic. Here, we apply paired metaproteomics and dsDNA-targeted metagenomics to identify 1,875 virion-associated proteins from the ocean. Over one-half of these proteins were newly functionally annotated and represent abundant and widespread viral metagenome-derived protein clusters (PCs). One primarily unannotated PC dominated the dataset, but structural modeling and genomic context identified this PC as a previously unidentified capsid protein from multiple uncultivated tailed virus families. Furthermore, four of the five most abundant PCs in the metaproteome represent capsid proteins containing the HK97-like protein fold previously found in many viruses that infect all three domains of life. The dominance of these proteins within our dataset, as well as their global distribution throughout the world’s oceans and seas, supports prior hypotheses that this HK97-like protein fold is the most abundant biological structure on Earth. Together, these culture-independent analyses improve virion-associated protein annotations, facilitate the investigation of proteins within natural viral communities, and offer a high-throughput means of illuminating functional viral dark matter.

scholarly journals Cyclin B interaction with microtubule-associated protein 4 (MAP4) targets p34cdc2 kinase to microtubules and is a potential regulator of M-phase microtubule dynamics.

1995 ◽  
Vol 128 (5) ◽  
pp. 849-862 ◽  
Author(s):  
K Ookata ◽  
S Hisanaga ◽  
J C Bulinski ◽  
H Murofushi ◽  
H Aizawa ◽  
...  
Keyword(s):  
Microtubule Dynamics ◽  
Cyclin B ◽  
Microtubule Associated Proteins ◽  
Mitotic Kinase ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Detergent Extraction ◽  
M Phase ◽  
Associated Proteins ◽  
Free Microtubules

We previously demonstrated (Ookata et al., 1992, 1993) that the p34cdc2/cyclin B complex associates with microtubules in the mitotic spindle and premeiotic aster in starfish oocytes, and that microtubule-associated proteins (MAPs) might be responsible for this interaction. In this study, we have investigated the mechanism by which p34cdc2 kinase associates with the microtubule cytoskeleton in primate tissue culture cells whose major MAP is known to be MAP4. Double staining of primate cells with anti-cyclin B and anti-MAP4 antibodies demonstrated these two antigens were colocalized on microtubules and copartitioned following two treatments that altered MAP4 distribution. Detergent extraction before fixation removed cyclin B as well as MAP4 from the microtubules. Depolymerization of some of the cellular microtubules with nocodazole preferentially retained the microtubule localization of both cyclin B and MAP4. The association of p34cdc2/cyclin B kinase with microtubules was also shown biochemically to be mediated by MAP4. Cosedimentation of purified p34cdc2/cyclin B with purified microtubule proteins containing MAP4, but not with MAP-free microtubules, as well as binding of MAP4 to GST-cyclin B fusion proteins, demonstrated an interaction between cyclin B and MAP4. Using recombinant MAP4 fragments, we demonstrated that the Pro-rich C-terminal region of MAP4 is sufficient to mediate the cyclin B-MAP4 interaction. Since p34cdc2/cyclin B physically associated with MAP4, we examined the ability of the kinase complex to phosphorylate MAP4. Incubation of a ternary complex of p34cdc2, cyclin B, and the COOH-terminal domain of MAP4, PA4, with ATP resulted in intracomplex phosphorylation of PA4. Finally, we tested the effects of MAP4 phosphorylation on microtubule dynamics. Phosphorylation of MAP4 by p34cdc2 kinase did not prevent its binding to microtubules, but abolished its microtubule stabilizing activity. Thus, the cyclin B/MAP4 interaction we have described may be important in targeting the mitotic kinase to appropriate cytoskeletal substrates, for the regulation of spindle assembly and dynamics.

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