scholarly journals Microtubule plus end–associated CLIP-170 initiates HSV-1 retrograde transport in primary human cells

2015 ◽  
Vol 211 (2) ◽  
pp. 323-337 ◽  
Author(s):  
Vladimir Jovasevic ◽  
Mojgan H. Naghavi ◽  
Derek Walsh
Keyword(s):  
Retrograde Transport ◽  
Human Cells ◽  
Dominant Negative ◽  
Therapeutic Implications ◽  
Virus Particles ◽  
Range Transport ◽  
Post Entry ◽  
Simplex Virus ◽  
Transferrin Uptake ◽  
Primary Human Cells

Dynamic microtubules (MTs) continuously explore the intracellular environment and, through specialized plus end–tracking proteins (+TIPs), engage a variety of targets. However, the nature of cargoes that require +TIP-mediated capture for their movement on MTs remains poorly understood. Using RNA interference and dominant-negative approaches, combined with live cell imaging, we show that herpes simplex virus particles that have entered primary human cells exploit a +TIP complex comprising end-binding protein 1 (EB1), cytoplasmic linker protein 170 (CLIP-170), and dynactin-1 (DCTN1) to initiate retrograde transport. Depletion of these +TIPs completely blocked post-entry long-range transport of virus particles and suppressed infection ∼5,000-fold, whereas transferrin uptake, early endosome organization, and dynein-dependent movement of lysosomes and mitochondria remained unaffected. These findings provide the first insights into the earliest stages of viral engagement of MTs through specific +TIPs, akin to receptors, with therapeutic implications, and identify herpesvirus particles as one of a very limited number of cargoes absolutely dependent on CLIP-170–mediated capture to initiate transport in primary human cells.

scholarly journals Characterization and Intracellular Trafficking of Epstein-Barr Virus BBLF1, a Protein Involved in Virion Maturation

2012 ◽  
Vol 86 (18) ◽  
pp. 9647-9655 ◽  
Author(s):  
Ya-Fang Chiu ◽  
Bill Sugden ◽  
Pey-Jium Chang ◽  
Lee-Wen Chen ◽  
Ying-Ju Lin ◽  
...  
Keyword(s):  
Epstein Barr Virus ◽  
Retrograde Transport ◽  
Lytic Replication ◽  
Herpes Simplex Virus 1 ◽  
Virus Particles ◽  
Tegument Proteins ◽  
Barr Virus ◽  
Viral Maturation ◽  
Epstein Barr ◽  
Simplex Virus

Epstein-Barr virus (EBV) BBLF1 shares 13 to 15% amino acid sequence identities with the herpes simplex virus 1 UL11 and cytomegalovirus UL99 tegument proteins, which are involved in the final envelopment during viral maturation. This study demonstrates that BBLF1 is a myristoylated and palmitoylated protein, as are UL11 and UL99. Myristoylation of BBLF1 both facilitates its membrane anchoring and stabilizes it. BBLF1 is shown to localize to thetrans-Golgi network (TGN) along with gp350/220, a site where final envelopment of EBV particles takes place. The localization of BBLF1 at the TGN requires myristoylation and two acidic clusters, which interact with PACS-1, a cytosolic protein, to mediate retrograde transport from the endosomes to the TGN. Knockdown of the expression of BBLF1 during EBV lytic replication reduces the production of virus particles, demonstrating the requirement of BBLF1 to achieve optimal production of virus particles. BBLF1 is hypothesized to facilitate the budding of tegumented capsid into glycoprotein-embedded membrane during viral maturation.

scholarly journals Full Resistance of Herpes Simplex Virus Type 1-Infected Primary Human Cells to Alpha Interferon Requires both the Us11 and γ134.5 Gene Products

2004 ◽  
Vol 78 (18) ◽  
pp. 10193-10196 ◽  
Author(s):  
Matthew Mulvey ◽  
Vladimir Camarena ◽  
Ian Mohr
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Gene Product ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Human Cells ◽  
Alpha Interferon ◽  
Simplex Virus ◽  
Primary Human Cells

ABSTRACT The γ134.5 gene product is important for the resistance of herpes simplex virus type 1 (HSV-1) to interferon. However, since the inhibition of protein synthesis observed in cells infected with a γ134.5 mutant virus results from the combined loss of the γ134.5 gene product and the failure to translate the late Us11 mRNA, we sought to characterize the relative interferon sensitivity of mutants unable to produce either the Us11 or the γ134.5 polypeptide. We now demonstrate that primary human cells infected with a Us11 mutant virus are hypersensitive to alpha interferon, arresting translation upon entry into the late phase of the viral life cycle. Furthermore, immediate-early expression of Us11 by a γ134.5 deletion mutant is sufficient to render translation resistant to alpha interferon. Finally, we establish that the Us11 gene product is required for wild-type levels of replication in alpha interferon-treated cells and, along with the γ134.5 gene, is an HSV-1-encoded interferon resistance determinant.

Microtubule plus end–associated CLIP-170 initiates HSV-1 retrograde transport in primary human cells

2015 ◽  
Vol 212 (12) ◽  
pp. 21212OIA100
Author(s):  
Vladimir Jovasevic ◽  
Mojgan H. Naghavi ◽  
Derek Walsh
Keyword(s):  
Retrograde Transport ◽  
Human Cells ◽  
Primary Human Cells ◽  
Hsv 1

scholarly journals SIGIRR/TIR-8 is an inhibitor of toll-like receptor signaling in primary human cells and regulates inflammation in models of rheumatoid arthritis

2010 ◽  
Vol 62 (8) ◽  
pp. 2249-2261 ◽  
Author(s):  
Stefan K. Drexler ◽  
Philip Kong ◽  
Julia Inglis ◽  
Richard O. Williams ◽  
Cecilia Garlanda ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Human Cells ◽  
Receptor Signaling ◽  
Toll Like Receptor ◽  
Primary Human Cells

Transcriptional transactivation functions localized to the glucocorticoid receptor N terminus are necessary for steroid induction of lymphocyte apoptosis

1992 ◽  
Vol 12 (2) ◽  
pp. 589-597
Author(s):  
E S Dieken ◽  
R L Miesfeld
Keyword(s):  
Transcriptional Regulation ◽  
Glucocorticoid Receptor ◽  
Target Genes ◽  
Dominant Negative ◽  
Receptor Expression ◽  
Transactivation Domain ◽  
Lymphocyte Apoptosis ◽  
Murine Cell Line ◽  
N Terminus ◽  
Simplex Virus

Genetic studies have suggested that transcriptional regulation of specific target genes (by either induction or repression) is the molecular basis of glucocorticoid-mediated lymphocyte apoptosis. To examine the role of transcriptional regulation more directly, we developed a complementation assay utilizing stable transfection of wild-type (wt) and mutant (nti) glucocorticoid receptor (GR) cDNA constructs into a GR-deficient S49 murine cell line (7r). Our data confirm that the level of functional GR is rate limiting for S49 apoptosis and moreover that the GR amino terminus (N terminus), which as been deleted from the nti GR, is absolutely required for complementation in this system. Surprisingly, we found that at physiological levels of receptor, expression of the nti GR in cells containing wt GR results in enhanced dexamethasone sensitivity rather than a dominant negative phenotype. One interpretation of these data is that DNA binding by wt-nti heterodimers may be functionally similar to that of wt-wt homodimers, indicating that GRE occupancy by at least one transactivation domain may be sufficient to induce the hormonal response. To determine whether acidic activating sequences such as those localized to the GR N terminus are important in the induction of lymphocyte apoptosis, we tested the activity of a chimeric receptor in which we replaced the entire GR N terminus with sequences from the herpes simplex virus VP16 protein. Our results demonstrate that 7r cells expressing VP-GR fusions are indeed steroid sensitive, strongly supporting the idea that S49 apoptosis is dependent on transcriptional regulation of specific genes which respond to acidic activating domains, implying that induction, rather than repression, may be the critical initiating event.

Multiscale porous ceramic scaffolds forin vitroculturing of primary human cells

2012 ◽  
Vol 111 (5-6) ◽  
pp. 262-268 ◽  
Author(s):  
A Finoli ◽  
N Ostrowski ◽  
E Schmelzer ◽  
I Nettleship ◽  
J Gerlach
Keyword(s):  
Porous Ceramic ◽  
Human Cells ◽  
Primary Human Cells

scholarly journals Tumour susceptibility gene 101 and the vacuolar protein sorting pathway are required for the release of hepatitis E virions

2011 ◽  
Vol 92 (12) ◽  
pp. 2838-2848 ◽  
Author(s):  
Shigeo Nagashima ◽  
Masaharu Takahashi ◽  
Suljid Jirintai ◽  
Toshinori Tanaka ◽  
Tsutomu Nishizawa ◽  
...  
Keyword(s):  
Susceptibility Gene ◽  
Multivesicular Body ◽  
Hepatitis E ◽  
Negative Control ◽  
Dominant Negative ◽  
Wild Type ◽  
Orf3 Protein ◽  
Virus Particles ◽  
Virion Release ◽  
Vacuolar Protein

We have previously demonstrated that an intact PSAP motif in the ORF3 protein is required for the formation and release of membrane-associated hepatitis E virus (HEV) particles with ORF3 proteins on their surface. In this study, we investigated the direct interaction between the ORF3 protein and tumour susceptibility gene 101 (Tsg101), a cellular factor involved in the budding of viruses containing the P(T/S)AP late-domain, in PLC/PRF/5 cells expressing the wild-type or PSAP-mutated ORF3 protein and Tsg101 by co-immunoprecipitation. Tsg101 bound to wild-type ORF3 protein, but not to the PSAP-inactive ORF3 protein. To examine whether HEV utilizes the multivesicular body (MVB) pathway to release the virus particles, we analysed the efficiency of virion release from cells upon introduction of small interfering RNA (siRNA) against Tsg101 or dominant-negative (DN) mutants of Vps4 (Vps4A and Vps4B). The relative levels of virus particles released from cells depleted of Tsg101 decreased to 6.4 % of those transfected with negative control siRNA. Similarly, virion egress was significantly reduced by the overexpression of DN forms (Vps4AEQ or Vps4BEQ). The relative levels of virus particles released from cells expressing Vps4AEQ and Vps4BEQ were 19.2 and 15.6 %, respectively, while the overexpression of wild-type Vps4A and Vps4B did not alter the levels of virus release. These results indicate that the ORF3 protein interacts with Tsg101 through the PSAP motifs in infected cells, and that Tsg101 and the enzymic activities of Vps4A and Vps4B are involved in HEV release, thus suggesting that HEV requires the MVB pathway for egress of virus particles.

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