scholarly journals Human Papillomavirus Type 16 E6 Promotes Retinoblastoma Protein Phosphorylation and Cell Cycle Progression

2004 ◽  
Vol 78 (24) ◽  
pp. 13769-13778 ◽  
Author(s):  
Ilaria Malanchi ◽  
Rosita Accardi ◽  
Frank Diehl ◽  
Anouk Smet ◽  
Elliot Androphy ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Human Papillomavirus ◽  
Cell Cycle Progression ◽  
Cellular Proliferation ◽  
Human Fibroblasts ◽  
Cyclin A ◽  
Hpv16 E6 ◽  
Protein Levels ◽  
Human Papillomavirus Type 16 ◽  
P53 Inactivation

ABSTRACT We show that E6 proteins from benign human papillomavirus type 1 (HPV1) and oncogenic HPV16 have the ability to alter the regulation of the G1/S transition of the cell cycle in primary human fibroblasts. Overexpression of both viral proteins induces cellular proliferation, retinoblastoma (pRb) phosphorylation, and accumulation of products of genes that are negatively regulated by pRb, such as p16INK4a, CDC2, E2F-1, and cyclin A. Hyperphosphorylated forms of pRb are present in E6-expressing cells even in the presence of ectopic levels of p16INK4a. The E6 proteins strongly increased the cyclin A/cyclin-dependent kinase 2 (CDK2) activity, which is involved in pRb phosphorylation. In addition, mRNA and protein levels of the CDK2 inhibitor p21WAF1/CIP1 were strongly down-regulated in cells expressing E6 proteins. The down-regulation of the p21WAF1/CIP1 gene appears to be independent of p53 inactivation, since HPV1 E6 and an HPV16 E6 mutant unable to target p53 were fully competent in decreasing p21WAF1/CIP1 levels. E6 from HPV1 and HPV16 also enabled cells to overcome the G1 arrest imposed by oncogenic ras. Immunofluorescence staining of cells coexpressing ras and E6 from either HPV16 or HPV1 revealed that antiproliferative (p16INK4a) and proliferative (Ki67) markers were coexpressed in the same cells. Together, these data underline a novel activity of E6 that is not mediated by inactivation of p53.

scholarly journals Human Papillomavirus Type 16 E7 Maintains Elevated Levels of the cdc25A Tyrosine Phosphatase during Deregulation of Cell Cycle Arrest

2002 ◽  
Vol 76 (2) ◽  
pp. 619-632 ◽  
Author(s):  
Don X. Nguyen ◽  
Thomas F. Westbrook ◽  
Dennis J. McCance
Keyword(s):  
Cell Cycle ◽  
Human Papillomavirus ◽  
Cell Cycle Arrest ◽  
Cell Cycle Progression ◽  
Tyrosine Phosphatase ◽  
Early Gene ◽  
Cycle Progression ◽  
Hpv 16 ◽  
Human Papillomavirus Type 16 ◽  
Cycle Arrest

ABSTRACT Essential to the oncogenic properties of human papillomavirus type 16 (HPV-16) are the activities encoded by the early gene product E7. HPV-16 E7 (E7.16) binds to cellular factors involved in cell cycle regulation and differentiation. These include the retinoblastoma tumor suppressor protein (Rb) and histone deacetylase (HDAC) complexes. While the biological significance of these interactions remains unclear, E7 is believed to help maintain cells in a proliferative state, thus establishing an environment that is conducive to viral replication. Most pathways that govern cell growth converge on downstream effectors. Among these is the cdc25A tyrosine phosphatase. cdc25A is required for G1/S transition, and its deregulation is associated with carcinogenesis. Considering the importance of cdc25A in cell cycle progression, it represents a relevant target for viral oncoproteins. Accordingly, the present study focuses on the putative deregulation of cdc25A by E7.16. Our results indicate that E7.16 can impede growth arrest induced during serum starvation and keratinocyte differentiation. Importantly, these E7-specific phenotypes correlate with elevated cdc25A steady-state levels. Reporter assays performed with NIH 3T3 cell lines and human keratinocytes indicate that E7 can transactivate the cdc25A promoter. In addition, transcriptional activation by E7.16 requires the distal E2F site within the cdc25A promoter. We further demonstrate that the ability of E7 to abrogate cell cycle arrest, activate cdc25A transcription, and increase cdc25A protein levels requires intact Rb and HDAC-1 binding domains. Finally, by using the cdk inhibitor roscovitine, we reveal that E7 activates the cdc25A promoter independently of cell cycle progression and cdk activity. Consequently, we propose that E7.16 can directly target cdc25A transcription and maintains cdc25A gene expression by disrupting Rb/E2F/HDAC-1 repressor complexes.

scholarly journals c-Myc Is Necessary for DNA Damage-Induced Apoptosis in the G2 Phase of the Cell Cycle

2001 ◽  
Vol 21 (15) ◽  
pp. 4929-4937 ◽  
Author(s):  
Susumu Adachi ◽  
Alvaro J. Obaya ◽  
Zhiyong Han ◽  
Noemi Ramos-Desimone ◽  
James H. Wyche ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Cell Cycle Progression ◽  
Cellular Proliferation ◽  
Ectopic Expression ◽  
Cyclin A ◽  
Cycle Progression ◽  
Downstream Effector ◽  
Induced Apoptosis ◽  
Kinetics Of

ABSTRACT The c-myc proto-oncogene encodes a transcription factor that participates in the regulation of cellular proliferation, differentiation, and apoptosis. Ectopic overexpression of c-Myc has been shown to sensitize cells to apoptosis. We report here that cells lacking c-Myc activity due to disruption of the c-myc gene by targeted homologous recombination are defective in DNA damage-initiated apoptosis in the G2 phase of the cell cycle. The downstream effector of c-Myc is cyclin A, whose ectopic expression in c-myc −/− cells rescues the apoptosis defect. The kinetics of the G2 response indicate that the induction of cyclin A and the concomitant activation of Cdk2 represent an early step during commitment to apoptosis. In contrast, expression of cyclins E and D1 does not rescue the apoptosis defect, and apoptotic processes in G1 phase are not affected in c-myc −/− cells. These observations link DNA damage-induced apoptosis with cell cycle progression and implicate c-Myc in the functioning of a subset of these pathways.

COX-2 expression and cell cycle progression in human fibroblasts

2001 ◽  
Vol 281 (1) ◽  
pp. C188-C194 ◽  
Author(s):  
Derek W. Gilroy ◽  
Michael A. Saunders ◽  
Kenneth K. Wu
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Cellular Proliferation ◽  
Human Fibroblasts ◽  
S Phase ◽  
Prostaglandin E ◽  
Cycle Progression ◽  
Proliferation And Apoptosis ◽  
Serum Inhibition ◽  
Cox 2

Cyclooxygenase-2 (COX-2) is continuously expressed in most cancerous cells where it appears to modulate cellular proliferation and apoptosis. However, little is known about the contribution of transient COX-2 induction to cell cycle progression or programmed cell death in primary cells. In this study we determined whether COX-2 regulates proliferation or apoptosis in human fibroblasts. COX-2 mRNA, protein, and prostaglandin E2(PGE2) were not detected in quiescent cells but were expressed during the G0/G1 phase of the cell cycle induced by serum. Inhibition of COX-2 did not alter G0/G1 to S phase transition or induce apoptosis at concentrations that diminished PGE2. Addition of interleukin-1β to serum enhanced COX-2 expression and PGE2 synthesis over that by serum alone but had no effect on the progression of these cells into S phase. Furthermore, platelet-derived growth factor drove the G0 fibroblasts into the cell cycle without inducing detectable levels of COX-2 or PGE2. Collectively, these data show that transient COX-2 expression in primary human fibroblasts does not influence cell cycle progression.

The oncoprotein NPM-ALK of anaplastic large-cell lymphoma induces JUNB transcription via ERK1/2 and JunB translation via mTOR signaling

Blood ◽  
2007 ◽  
Vol 110 (9) ◽  
pp. 3374-3383 ◽  
Author(s):  
Philipp B. Staber ◽  
Paul Vesely ◽  
Naznin Haq ◽  
Rene G. Ott ◽  
Kotaro Funato ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Translational Control ◽  
Cell Cycle Progression ◽  
Cellular Proliferation ◽  
Mrna Translation ◽  
Large Cell ◽  
Protein Levels ◽  
M Phase ◽  
Alk Positive ◽  
Interfering Rna

Abstract Anaplastic large cell lymphomas (ALCLs) are highly proliferating tumors that commonly express the AP-1 transcription factor JunB. ALK fusions occur in approximately 50% of ALCLs, and among these, 80% have the t(2;5) translocation with NPM-ALK expression. We report greater activity of JunB in NPM-ALK–positive than in NPM-ALK–negative ALCLs. Specific knockdown of JUNB mRNA using small interfering RNA and small hairpin RNA in NPM-ALK–expressing cells decreases cellular proliferation as evidenced by a reduced cell count in the G2/M phase of the cell cycle. Expression of NPM-ALK results in ERK1/2 activation and transcriptional up-regulation of JUNB. Both NPM-ALK–positive and –negative ALCL tumors demonstrate active ERK1/2 signaling. In contrast to NPM-ALK–negative ALCL, the mTOR pathway is active in NPM-ALK–positive lymphomas. Pharmacological inhibition of mTOR in NPM-ALK–positive cells down-regulates JunB protein levels by shifting JUNB mRNA translation from large polysomes to monosomes and ribonucleic particles (RNPs), and decreases cellular proliferation. Thus, JunB is a critical target of mTOR and is translationally regulated in NPM-ALK–positive lymphomas. This is the first study demonstrating translational control of AP-1 transcription factors in human neoplasia. In conjunction with NPM-ALK, JunB enhances cell cycle progression and may therefore represent a therapeutic target.

scholarly journals The Ubiquitin-Specific Peptidase USP15 Regulates Human Papillomavirus Type 16 E6 Protein Stability

2009 ◽  
Vol 83 (17) ◽  
pp. 8885-8892 ◽  
Author(s):  
Robin M. Vos ◽  
Jennifer Altreuter ◽  
Elizabeth A. White ◽  
Peter M. Howley
Keyword(s):  
Human Papillomavirus ◽  
Protein Stability ◽  
Affinity Purification ◽  
Peptidase Activity ◽  
Hpv16 E6 ◽  
Protein Levels ◽  
Human Papillomavirus Type 16 ◽  
Associated Proteins ◽  
Interfering Rna ◽  
E6 Protein

ABSTRACT Proteomic identification of human papillomavirus type 16 (HPV16) E6-interacting proteins revealed several proteins involved in ubiquitin-mediated proteolysis. In addition to the well-characterized E6AP ubiquitin-protein ligase, a second HECT domain protein (HERC2) and a deubiquitylating enzyme (USP15) were identified by tandem affinity purification of HPV16 E6-associated proteins. This study focuses on the functional consequences of the interaction of E6 with USP15. Overexpression of USP15 resulted in increased levels of the E6 protein, and the small interfering RNA-mediated knockdown of USP15 decreased E6 protein levels. These results implicate USP15 directly in the regulation of E6 protein stability and suggest that ubiquitylated E6 could be a substrate for USP15 ubiquitin peptidase activity. It remains possible that E6 could affect the activity of USP15 on specific cellular substrates, a hypothesis that can be tested as more is learned about the substrates and pathways controlled by USP15.

scholarly journals Functional cdc25C Dual-Specificity Phosphatase Is Required for S-Phase Entry in Human Cells

2003 ◽  
Vol 14 (7) ◽  
pp. 2984-2998 ◽  
Author(s):  
Patric Turowski ◽  
Celine Franckhauser ◽  
May C. Morris ◽  
Philippe Vaglio ◽  
Anne Fernandez ◽  
...  
Keyword(s):  
Hela Cells ◽  
Cell Cycle Progression ◽  
Human Fibroblasts ◽  
Cyclin A ◽  
S Phase ◽  
Protein Levels ◽  
Dual Specificity ◽  
Activity Measurements ◽  
Inhibition Of Dna Synthesis ◽  
Interfering Rna

In view of the common regulatory mechanism that induces transcription of the mitotic phosphatase cdc25C and cyclin A at the beginning of S-phase, we investigated whether cdc25C was required for S-phase transit. Here, we show that in both nontransformed human fibroblasts and HeLa cells, cdc25C protein levels significantly increased concomitant with S-phase onset and cyclin A synthesis. Activity measurements on immunoprecipitates from synchronized HeLa cells revealed a sharp rise in cdc25C-associated phosphatase activity that coincided with S-phase. Microinjection of various antisense-cdc25C molecules led to inhibition of DNA synthesis in both HeLa cells and human fibroblasts. Furthermore, transfection of small interfering RNA directed against cdc25C specifically depleted cdc25C in HeLa cells without affecting cdc25A or cdc25B levels. Cdc25C RNA interference was also accompanied by S-phase inhibition. In cells depleted of cdc25C by antisense or siRNA, normal cell cycle progression could be re-established through microinjection of wild-type cdc25C protein but not inactive C377S mutant protein. Taken together, these results show that cdc25C not only plays a role at the G2/M transition but also in the modulation of DNA replication where its function is distinct from that of cdc25A.

scholarly journals Disruption of the G1/S Transition in Human Papillomavirus Type 16 E7-Expressing Human Cells Is Associated with Altered Regulation of Cyclin E

1998 ◽  
Vol 72 (2) ◽  
pp. 975-985 ◽  
Author(s):  
Larry G. Martin ◽  
G. William Demers ◽  
Denise A. Galloway
Keyword(s):  
Cell Cycle ◽  
Human Papillomavirus ◽  
Cell Cycle Progression ◽  
Cyclin E ◽  
Cell Cycle Control ◽  
Cell Cycle Checkpoints ◽  
Hpv 16 ◽  
Human Papillomavirus Type 16 ◽  
E6 And E7 ◽  
Cell Cycle Machinery

ABSTRACT The development of neoplasia frequently involves inactivation of the p53 and retinoblastoma (Rb) tumor suppressor pathways and disruption of cell cycle checkpoints that monitor the integrity of replication and cell division. The human papillomavirus type 16 (HPV-16) oncoproteins, E6 and E7, have been shown to bind p53 and Rb, respectively. To further delineate the mechanisms by which E6 and E7 affect cell cycle control, we examined various aspects of the cell cycle machinery. The low-risk HPV-6 E6 and E7 proteins did not cause any significant change in the levels of cell cycle proteins analyzed. HPV-16 E6 resulted in very low levels of p53 and p21 and globally elevated cyclin-dependent kinase (CDK) activity. In contrast, HPV-16 E7 had a profound effect on several aspects of the cell cycle machinery. A number of cyclins and CDKs were elevated, and despite the elevation of the levels of at least two CDK inhibitors, p21 and p16, CDK activity was globally increased. Most strikingly, cyclin E expression was deregulated both transcriptionally and posttranscriptionally and persisted at high levels in S and G2/M. Transit through G1 was shortened by the premature activation of cyclin E-associated kinase activity. Elevation of cyclin E levels required both the CR1 and CR2 domains of E7. These data suggest that cyclin E may be a critical target of HPV-16 E7 in the disruption of G1/S cell cycle progression and that the ability of E7 to regulate cyclin E involves activities in addition to the release of E2F.

scholarly journals Productive Replication of Adeno-Associated Virus Can Occur in Human Papillomavirus Type 16 (HPV-16) Episome-Containing Keratinocytes and Is Augmented by the HPV-16 E2 Protein

2000 ◽  
Vol 74 (8) ◽  
pp. 3494-3504 ◽  
Author(s):  
Phyllis Ogston ◽  
Kenneth Raj ◽  
Peter Beard
Keyword(s):  
Cell Cycle ◽  
Human Papillomavirus ◽  
Dna Replication ◽  
Cell Cycle Progression ◽  
Cycle Progression ◽  
Hpv 16 ◽  
Adeno Associated Virus ◽  
E2 Protein ◽  
Human Papillomavirus Type 16 ◽  
Productive Replication

ABSTRACT We used a sensitive assay to test whether an adeno-associated virus (AAV) productive replication cycle can occur in immortalized human keratinocytes carrying episomal human papillomavirus type 16 (HPV-16) DNA. Following transfection with cloned AAV DNA, infectious AAV was produced, and the infectivity was blocked by anti-AAV antiserum. The HPV-16 E2 protein substantially increased the yield of AAV. Other HPV early proteins did not, in our experiments, show this ability. E2 has been shown to be able to affect p53 levels and to block cell cycle progression at mitosis. We tested the effect of changes in p53 expression on AAV replication and found that large differences in the level of p53 did not alter AAV DNA replication. In extension of this, we found that cellular help for AAV in response to stress was also independent of p53. To test if a mitotic block could trigger AAV DNA replication, we treated the cells with the mitotic inhibitor nocodazole. AAV DNA replication was stimulated by the presence of nocodazole in these and a number of other cell types tested. Yields of infectious virus, however, were not increased by this treatment. We conclude that the HPV-16 E2 protein stimulates AAV multiplication in these cells and propose that this occurs independently of the effects of E2 on p53 and cell cycle progression. Since the effect of E2 was not seen in keratinocytes lacking the HPV-16 episome, we suggest that E2 can help AAV by working in concert with other HPV-16 proteins.

scholarly journals Alu RNA Modulates the Expression of Cell Cycle Genes in Human Fibroblasts

2019 ◽  
Vol 20 (13) ◽  
pp. 3315 ◽  
Author(s):  
Simona Cantarella ◽  
Davide Carnevali ◽  
Marco Morselli ◽  
Anastasia Conti ◽  
Matteo Pellegrini ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Human Fibroblasts ◽  
Rna Polymerase Iii ◽  
Protein Coding ◽  
Non Coding Rna ◽  
Genome Wide ◽  
Hela Cell Lines ◽  
Significant Enrichment ◽  
Alu Rna

Alu retroelements, whose retrotransposition requires prior transcription by RNA polymerase III to generate Alu RNAs, represent the most numerous non-coding RNA (ncRNA) gene family in the human genome. Alu transcription is generally kept to extremely low levels by tight epigenetic silencing, but it has been reported to increase under different types of cell perturbation, such as viral infection and cancer. Alu RNAs, being able to act as gene expression modulators, may be directly involved in the mechanisms determining cellular behavior in such perturbed states. To directly address the regulatory potential of Alu RNAs, we generated IMR90 fibroblasts and HeLa cell lines stably overexpressing two slightly different Alu RNAs, and analyzed genome-wide the expression changes of protein-coding genes through RNA-sequencing. Among the genes that were upregulated or downregulated in response to Alu overexpression in IMR90, but not in HeLa cells, we found a highly significant enrichment of pathways involved in cell cycle progression and mitotic entry. Accordingly, Alu overexpression was found to promote transition from G1 to S phase, as revealed by flow cytometry. Therefore, increased Alu RNA may contribute to sustained cell proliferation, which is an important factor of cancer development and progression.

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