scholarly journals Three Regions of the pRB Pocket Domain Affect Its Inactivation by Human Papillomavirus E7 Proteins

2002 ◽  
Vol 76 (12) ◽  
pp. 6224-6234 ◽  
Author(s):  
Frederick A. Dick ◽  
Nicholas J. Dyson
Keyword(s):  
Amino Acids ◽  
Human Papillomavirus ◽  
Hela Cells ◽  
Critical Event ◽  
Interaction Point ◽  
Discrete Interaction ◽  
Binding Cleft ◽  
Human Papillomavirus 18 ◽  
E7 Proteins ◽  
Pocket Domain

ABSTRACT A critical event in papillomavirus transformation of human cells is the inactivation of pRB by the E7 protein. E7, like many other viral oncoproteins, possesses a well-characterized LXCXE peptide motif that interacts with the pocket domain of pRB. Disruption of the LXCXE-binding cleft on pRB renders it resistant to E7 binding and inactivation. Such binding cleft mutants of pRB are capable of inducing a G1 arrest in the human papillomavirus 18-transformed HeLa cell line. We show here that the efficient inactivation of pRB in HeLa cells does not simply depend on the integrity of the LXCXE-binding cleft. Multiple site-directed mutants that alter conserved surfaces of the pRB pocket domain cause HeLa cells to accumulate in G1. We divide these mutants into two classes: those that can be bound by E7 and those that cannot. The E7 interacting mutants include changes in conserved residues that lie in a groove between the A and B halves of the pocket. Surprisingly, none of these mutants show a clear defect in any of the known mechanisms for pRB inactivation by E7. Analysis of mutants that are compromised for E7 binding reveals that this interaction depends on both the LXCXE-binding cleft and on a conserved group of lysines adjacent to the cleft. These basic amino acids on pRB define a discrete interaction point with E7. These residues most likely form ionic interactions with conserved acidic amino acids on E7 since a stable pRB/E7 interaction was restored when the lysine residues on pRB and the acidic residues on E7 were interchanged.

scholarly journals Human papillomavirus 18 E6 inhibits phosphorylation of p53 expressed in HeLa cells

2012 ◽  
Vol 2 (1) ◽  
pp. 2 ◽  
Author(s):  
Amrendra K Ajay ◽  
Avtar S Meena ◽  
Manoj K Bhat
Keyword(s):  
Human Papillomavirus ◽  
Hela Cells ◽  
Human Papillomavirus 18

scholarly journals Zombies in TCGA

2015 ◽  
Vol 89 (8) ◽  
pp. 4044-4046 ◽  
Author(s):  
Daniel DiMaio
Keyword(s):  
Human Papillomavirus ◽  
Next Generation Sequencing ◽  
Hela Cells ◽  
Somatic Mutations ◽  
Hpv Infection ◽  
Next Generation ◽  
Human Cancers ◽  
Human Papillomavirus 18 ◽  
Tumor Types ◽  
Generation Sequencing

Next-generation sequencing results obtained to detect somatic mutations in human cancers can also be searched for viruses that contribute to cancer. Recently, human papillomavirus 18 RNA was detected in tumor types not typically associated with HPV infection. Analyses reported in this issue ofJournal of Virologydemonstrate that the apparent presence of HPV18 RNA in these atypical tumors is due in at least some cases to contamination of samples with HeLa cells, which harbor HPV18.

scholarly journals HeLa Nucleic Acid Contamination in The Cancer Genome Atlas Leads to the Misidentification of Human Papillomavirus 18

2015 ◽  
Vol 89 (8) ◽  
pp. 4051-4057 ◽  
Author(s):  
Paul G. Cantalupo ◽  
Joshua P. Katz ◽  
James M. Pipas
Keyword(s):  
Cervical Cancer ◽  
Human Papillomavirus ◽  
Nucleic Acid ◽  
Hela Cells ◽  
Cancer Genome ◽  
The Cancer Genome Atlas ◽  
Cancer Genome Atlas ◽  
Sequence Databases ◽  
Human Papillomavirus 18 ◽  
Genome Atlas

ABSTRACTWe searched The Cancer Genome Atlas (TCGA) database for viruses by comparing non-human reads present in transcriptome sequencing (RNA-Seq) and whole-exome sequencing (WXS) data to viral sequence databases. Human papillomavirus 18 (HPV18) is an etiologic agent of cervical cancer, and as expected, we found robust expression of HPV18 genes in cervical cancer samples. In agreement with previous studies, we also found HPV18 transcripts in non-cervical cancer samples, including those from the colon, rectum, and normal kidney. However, in each of these cases, HPV18 gene expression was low, and single-nucleotide variants and positions of genomic alignments matched the integrated portion of HPV18 present in HeLa cells. Chimeric reads that match a known virus-cell junction of HPV18 integrated in HeLa cells were also present in some samples. We hypothesize that HPV18 sequences in these non-cervical samples are due to nucleic acid contamination from HeLa cells. This finding highlights the problems that contamination presents in computational virus detection pipelines.IMPORTANCEViruses associated with cancer can be detected by searching tumor sequence databases. Several studies involving searches of the TCGA database have reported the presence of HPV18, a known cause of cervical cancer, in a small number of additional cancers, including those of the rectum, kidney, and colon. We have determined that the sequences related to HPV18 in non-cervical samples are due to nucleic acid contamination from HeLa cells. To our knowledge, this is the first report of the misidentification of viruses in next-generation sequencing data of tumors due to contamination with a cancer cell line. These results raise awareness of the difficulty of accurately identifying viruses in human sequence databases.

scholarly journals Enterovirus 70 Receptor Utilization Is Controlled by Capsid Residues That Also Regulate Host Range and Cytopathogenicity

2007 ◽  
Vol 81 (16) ◽  
pp. 8648-8655 ◽  
Author(s):  
Melissa Stewart Kim ◽  
Vincent R. Racaniello
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Rhesus Monkey ◽  
Host Range ◽  
Hela Cells ◽  
Lytic Replication ◽  
Monkey Kidney ◽  
Kidney Cells ◽  
Cytopathic Effects ◽  
Axis Of Symmetry

ABSTRACT Enterovirus type 70, an etiologic agent of acute hemorrhagic conjunctivitis, may bind different cellular receptors depending on cell type. To understand how EV70-receptor interaction is controlled, we studied two variants of the virus with distinct receptor utilization. EV70-Rmk, derived by passage in rhesus monkey kidney cells, replicates poorly in HeLa cells and does not cause cytopathic effects. Decay accelerating factor (DAF) is not a cell receptor for EV70-Rmk. Passage of EV70-Rmk in HeLa cells lead to isolation of EV70-Dne, which does not replicate in rhesus monkey kidney cells but grows to high titers in HeLa cells and causes cytopathic effects. DAF is sufficient for cell entry of EV70-Dne. EV70-Rmk replicates in human eye and brain-derived cell lines, whereas the Dne strain replicates only in HeLa cells and in conjunctiva-derived 15C4 cells. The two EV70 strains differ by five amino acid changes in the viral capsid. Single substitution of four of the five EV70-Rmk amino acids with the residue from EV70-Dne leads to lytic replication in HeLa cells. Conversely, substitution of any of the five EV70-Dne amino acids with the EV70-Rmk amino acid does not alter replication in HeLa cells. Three of these capsid amino acids are predicted to be located in the canyon encircling the fivefold axis of symmetry, one amino acid is found at the fivefold axis of symmetry, and one is located the interior of the capsid. The five EV70 residues define a region of the capsid that controls viral host range, DAF utilization, and cytopathogenicity.

scholarly journals Stability of the Human Papillomavirus Type 18 E2 Protein Is Regulated by a Proteasome Degradation Pathway through Its Amino-Terminal Transactivation Domain

2001 ◽  
Vol 75 (16) ◽  
pp. 7244-7251 ◽  
Author(s):  
Sophie Bellanger ◽  
Caroline Demeret ◽  
Sylvain Goyat ◽  
Françoise Thierry
Keyword(s):  
Human Papillomavirus ◽  
Hela Cells ◽  
Half Life ◽  
Fluorescent Protein ◽  
Growth Arrest ◽  
Transactivation Domain ◽  
Mcf7 Cells ◽  
E2 Protein ◽  
Amino Terminal

ABSTRACT The E2 proteins of papillomaviruses regulate both viral transcription and DNA replication. The human papillomavirus type 18 (HPV18) E2 protein has been shown to repress transcription of the oncogenic E6 and E7 genes, inducing growth arrest in HeLa cells. Using HPV18 E2 fused to the green fluorescent protein (GFP), we showed that this protein was short-lived in transfected HeLa cells. Real-time microscopy experiments indicated that the E2-dependent signal increased for roughly 24 h after transfection and then rapidly disappeared, indicating that E2 was unstable in HeLa cells and could confer instability to GFP. Similar studies done with a protein lacking the transactivation domain indicated that this truncation strongly stabilizes the E2 protein. In vitro, full-length E2 or the transactivation domain alone was efficiently ubiquitinated, whereas deletion of the transactivation domain strongly decreased the ubiquitination of the E2 protein. Proteasome inhibition in cells expressing E2 increased its half-life about sevenfold, which was comparable to the half-life of the amino-terminally truncated protein. These characteristics of E2 instability were independent of the E2-mediated G1 growth arrest in HeLa cells, as they were reproduced in MCF7 cells, where E2 does not affect the cell cycle. Altogether, these experiments showed that the HPV18 E2 protein was degraded by the ubiquitin-proteasome pathway through its amino-terminal transactivation domain. Tight regulation of the stability of the HPV 18 E2 protein may be essential to avoid accumulation of a potent transcriptional repressor and antiproliferative agent during the viral vegetative cycle.

scholarly journals Effect of Hormonal Steroids on the Uptake of Labeled Amino Acids by HeLa Cells

1971 ◽  
Vol 19 (1) ◽  
pp. 11-15 ◽  
Author(s):  
TETSURO MOHRI ◽  
CHIYOKO NAKAGAWA ◽  
HARUO KITAGAWA
Keyword(s):  
Amino Acids ◽  
Hela Cells ◽  
Hormonal Steroids
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