scholarly journals Quantitative Profiling of Adaptation to Cyclin E Overproduction

2021 ◽  
Author(s):  
Juanita C Limas ◽  
Aimee N. Littlejohn ◽  
Amy M. House ◽  
Katarzyna M Kedziora ◽  
Dalia Fleifel ◽  
...  

Cyclin E/CDK2 drives cell cycle progression from G1 to S phase. Cyclin E overproduction is toxic to mammalian cells, although the gene encoding cyclin E (CCNE1) is overexpressed in some cancers. It is not yet understood how cancer cells tolerate high levels of cyclin E. To address this question, we extensively characterized non-transformed epithelial cells subjected to chronic cyclin E overproduction. Cells overproducing human cyclin E briefly experienced truncated G1 phases, then consistently endured a transient period of DNA replication origin underlicensing, replication stress, and severely impaired proliferation. Individual cells displayed substantial intercellular heterogeneity in both cell cycle dynamics and CDK activity. Each of these phenotypes improved rapidly despite maintaining high cyclin E-associated activity. Transcriptome analysis revealed that adapted cells downregulated a cohort of G1-regulated genes. These cells also shared at least one unique change also found in breast tumors that overproduce cyclin E, expression of the cancer/testis antigen HORMAD1. Withdrawing cyclin E induction partially reversed the intermediate licensing phenotype of adapted cells indicating that adaptation is at least partly independent of genetic alterations. This study provides evidence that mammalian cyclin E/CDK inhibits origin licensing by an indirect mechanism through premature S phase onset. It serves as an example of specific oncogene adaptation that can identify key molecular changes during tumorigenesis.

1999 ◽  
Vol 19 (1) ◽  
pp. 612-622 ◽  
Author(s):  
Jean M. Gudas ◽  
Marc Payton ◽  
Sushil Thukral ◽  
Eddy Chen ◽  
Michael Bass ◽  
...  

ABSTRACT A novel cyclin gene was discovered by searching an expressed sequence tag database with a cyclin box profile. The human cyclin E2 gene encodes a 404-amino-acid protein that is most closely related to cyclin E. Cyclin E2 associates with Cdk2 in a functional kinase complex that is inhibited by both p27Kip1 and p21Cip1. The catalytic activity associated with cyclin E2 complexes is cell cycle regulated and peaks at the G1/S transition. Overexpression of cyclin E2 in mammalian cells accelerates G1, demonstrating that cyclin E2 may be rate limiting for G1 progression. Unlike cyclin E1, which is expressed in most proliferating normal and tumor cells, cyclin E2 levels were low to undetectable in nontransformed cells and increased significantly in tumor-derived cells. The discovery of a novel second cyclin E family member suggests that multiple unique cyclin E-CDK complexes regulate cell cycle progression.


2003 ◽  
Vol 23 (22) ◽  
pp. 8110-8123 ◽  
Author(s):  
Partha Mitra ◽  
Rong-Lin Xie ◽  
Ricardo Medina ◽  
Hayk Hovhannisyan ◽  
S. Kaleem Zaidi ◽  
...  

ABSTRACT At the G1/S phase cell cycle transition, multiple histone genes are expressed to ensure that newly synthesized DNA is immediately packaged as chromatin. Here we have purified and functionally characterized the critical transcription factor HiNF-P, which is required for E2F-independent activation of the histone H4 multigene family. Using chromatin immunoprecipitation analysis and ligation-mediated PCR-assisted genomic sequencing, we show that HiNF-P interacts with conserved H4 cell cycle regulatory sequences in vivo. Antisense inhibition of HiNF-P reduces endogenous histone H4 gene expression. Furthermore, we find that HiNF-P utilizes NPAT/p220, a substrate of the cyclin E/cyclin-dependent kinase 2 (CDK2) kinase complex, as a key coactivator to enhance histone H4 gene transcription. The biological role of HiNF-P is reflected by impeded cell cycle progression into S phase upon antisense-mediated reduction of HiNF-P levels. Our results establish that HiNF-P is the ultimate link in a linear signaling pathway that is initiated with the growth factor-dependent induction of cyclin E/CDK2 kinase activity at the restriction point and culminates in the activation of histone H4 genes through HiNF-P at the G1/S phase transition.


2010 ◽  
Vol 9 (1) ◽  
pp. 302 ◽  
Author(s):  
Hicham H Baydoun ◽  
Joanna Pancewicz ◽  
XueTao Bai ◽  
Christophe Nicot

2002 ◽  
Vol 76 (24) ◽  
pp. 12543-12552 ◽  
Author(s):  
Amy Mauser ◽  
Elizabeth Holley-Guthrie ◽  
Adam Zanation ◽  
Wendall Yarborough ◽  
William Kaufmann ◽  
...  

ABSTRACT The Epstein-Barr virus (EBV) immediate-early protein BZLF1 mediates the switch between the latent and lytic forms of EBV infection and has been previously shown to induce a G1/S block in cell cycle progression in some cell types. To examine the effect of BZLF1 on cellular gene expression, we performed microarray analysis on telomerase-immortalized human keratinocytes that were mock infected or infected with a control adenovirus vector (AdLacZ) or a vector expressing the EBV BZLF1 protein (AdBZLF1). Cellular genes activated by BZLF1 expression included E2F-1, cyclin E, Cdc25A, and a number of other genes involved in cell cycle progression. Immunoblot analysis confirmed that BZLF1 induced expression of E2F-1, cyclin E, Cdc25A, and stem loop binding protein (a protein known to be primarily expressed during S phase) in telomerase-immortalized keratinocytes. Similarly, BZLF1 increased expression of E2F-1, cyclin E, and stem loop binding protein (SLBP) in primary tonsil keratinocytes. In contrast, BZLF1 did not induce E2F-1 expression in normal human fibroblasts. Cell cycle analysis revealed that while BZLF1 dramatically blocked G1/S progression in normal human fibroblasts, it did not significantly affect cell cycle progression in primary human tonsil keratinocytes. Furthermore, in EBV-infected gastric carcinoma cells, the BZLF1-positive cells had an increased number of cells in S phase compared to the BZLF1-negative cells. Thus, in certain cell types (but not others), BZLF1 enhances expression of cellular proteins associated with cell cycle progression, which suggests that an S-phase-like environment may be advantageous for efficient lytic EBV replication in some cell types.


2013 ◽  
Vol 203 (2) ◽  
pp. 233-250 ◽  
Author(s):  
Albert Lu ◽  
Suzanne R. Pfeffer

Cyclin E regulates the cell cycle transition from G1 to S phase and is degraded before entry into G2 phase. Here we show that RhoBTB3, a Golgi-associated, Rho-related ATPase, regulates the S/G2 transition of the cell cycle by targeting Cyclin E for ubiquitylation. Depletion of RhoBTB3 arrested cells in S phase, triggered Golgi fragmentation, and elevated Cyclin E levels. On the Golgi, RhoBTB3 bound Cyclin E as part of a Cullin3 (CUL3)-dependent RING–E3 ubiquitin ligase complex comprised of RhoBTB3, CUL3, and RBX1. Golgi association of this complex was required for its ability to catalyze Cyclin E ubiquitylation and allow normal cell cycle progression. These experiments reveal a novel role for a Ras superfamily member in catalyzing Cyclin E turnover during S phase, as well as an unexpected, essential role for the Golgi as a ubiquitylation platform for cell cycle control.


2008 ◽  
Vol 415 (3) ◽  
pp. 439-448 ◽  
Author(s):  
Katherine A. Kaproth-Joslin ◽  
Xiangquan Li ◽  
Sarah E. Reks ◽  
Grant G. Kelley

In the present study, we examined the role of PLCδ1 (phospholipase C δ1) in the regulation of cellular proliferation. We demonstrate that RNAi (RNA interference)-mediated knockdown of endogenous PLCδ1, but not PLCβ3 or PLCϵ, induces a proliferation defect in Rat-1 and NIH 3T3 fibroblasts. The decreased proliferation was not due to an induction of apoptosis or senescence, but was associated with an approx. 60% inhibition of [3H]thymidine incorporation. Analysis of the cell cycle with BrdU (bromodeoxyuridine)/propidium iodide-labelled FACS (fluorescence-activated cell sorting) demonstrated an accumulation of cells in G0/G1-phase and a corresponding decrease in cells in S-phase. Further examination of the cell cycle after synchronization by serum-starvation demonstrated normal movement through G1-phase but delayed entry into S-phase. Consistent with these findings, G1 cyclin (D2 and D3) and CDK4 (cyclin-dependent kinase 4) levels and associated kinase activity were not affected. However, cyclin E-associated CDK2 activity, responsible for G1-to-S-phase progression, was inhibited. This decreased activity was accompanied by unchanged CDK2 protein levels and paradoxically elevated cyclin E and cyclin E-associated CDK2 levels, suggesting inhibition of the cyclin E–CDK2 complex. This inhibition was not due to altered stimulatory or inhibitory phosphorylation of CDK2. However, p27, a Cip/Kip family CKI (CDK inhibitor)-binding partner, was elevated and showed increased association with CDK2 in PLCδ1-knockdown cells. The result of the present study demonstrate a novel and critical role for PLCδ1 in cell-cycle progression from G1-to-S-phase through regulation of cyclin E–CDK2 activity and p27 levels.


2003 ◽  
Vol 23 (8) ◽  
pp. 2821-2833 ◽  
Author(s):  
Guang Gao ◽  
Adrian P. Bracken ◽  
Karina Burkard ◽  
Diego Pasini ◽  
Marie Classon ◽  
...  

ABSTRACT NPAT is an in vivo substrate of cyclin E-Cdk2 kinase and is thought to play a critical role in coordinated transcriptional activation of histone genes during the G1/S-phase transition and in S-phase entry in mammalian cells. Here we show that NPAT transcription is up-regulated at the G1/S-phase boundary in growth-stimulated cells and that the NPAT promoter responds to activation by E2F proteins. We demonstrate that endogenous E2F proteins interact with the promoter of the NPAT gene in vivo and that induced expression of E2F1 stimulates NPAT mRNA expression, supporting the idea that the expression of NPAT is regulated by E2F. Consistently, we find that the E2F sites in the NPAT promoter are required for its activation during the G1/S-phase transition. Moreover, we show that the expression of NPAT accelerates S-phase entry in cells released from quiescence. The inhibition of NPAT expression by small interfering RNA duplexes impedes cell cycle progression and histone gene expression in tissue culture cells. Thus, NPAT is an important E2F target that is required for cell cycle progression in mammalian cells. As NPAT is involved in the regulation of S-phase-specific histone gene transcription, our findings indicate that NPAT links E2F to the activation of S-phase-specific histone gene transcription.


2005 ◽  
Vol 168 (5) ◽  
pp. 713-722 ◽  
Author(s):  
Sabrina La Terra ◽  
Christopher N. English ◽  
Polla Hergert ◽  
Bruce F. McEwen ◽  
Greenfield Sluder ◽  
...  

It has been reported that nontransformed mammalian cells become arrested during G1 in the absence of centrioles (Hinchcliffe, E., F. Miller, M. Cham, A. Khodjakov, and G. Sluder. 2001. Science. 291:1547–1550). Here, we show that removal of resident centrioles (by laser ablation or needle microsurgery) does not impede cell cycle progression in HeLa cells. HeLa cells born without centrosomes, later, assemble a variable number of centrioles de novo. Centriole assembly begins with the formation of small centrin aggregates that appear during the S phase. These, initially amorphous “precentrioles” become morphologically recognizable centrioles before mitosis. De novo–assembled centrioles mature (i.e., gain abilities to organize microtubules and replicate) in the next cell cycle. This maturation is not simply a time-dependent phenomenon, because de novo–formed centrioles do not mature if they are assembled in S phase–arrested cells. By selectively ablating only one centriole at a time, we find that the presence of a single centriole inhibits the assembly of additional centrioles, indicating that centrioles have an activity that suppresses the de novo pathway.


2005 ◽  
Vol 171 (3) ◽  
pp. 437-445 ◽  
Author(s):  
Chaozhong Zou ◽  
Jun Li ◽  
Yujie Bai ◽  
William T. Gunning ◽  
David E. Wazer ◽  
...  

In mammalian cells, the centrosome consists of a pair of centrioles and amorphous pericentriolar material. The pair of centrioles, which are the core components of the centrosome, duplicate once per cell cycle. Centrosomes play a pivotal role in orchestrating the formation of the bipolar spindle during mitosis. Recent studies have linked centrosomal activity on centrioles or centriole-associated structures to cytokinesis and cell cycle progression through G1 into the S phase. In this study, we have identified centrobin as a centriole-associated protein that asymmetrically localizes to the daughter centriole. The silencing of centrobin expression by small interfering RNA inhibited centriole duplication and resulted in centrosomes with one or no centriole, demonstrating that centrobin is required for centriole duplication. Furthermore, inhibition of centriole duplication by centrobin depletion led to impaired cytokinesis.


2015 ◽  
Vol 65 (4) ◽  
pp. 463-471 ◽  
Author(s):  
Zhiwei Huang ◽  
Lianqiu Wang ◽  
Lifeng Chen ◽  
Yifei Zhang ◽  
Ping Shi

Abstract Clioquinol has been shown to have anticancer activity in several carcinoma cells. In this study, we preliminarily examined the effect of clioquinol in human SMMC-7721 hepatoma and QSG-7701 normal hepatic cells. Our results indicated that clioquinol did not significantly affect survival of QSG-7701 cells, whereas it reduced cell viability in a concentration- and time-dependent manner in SMMC-7721 cells. Clioquinol did not trigger autophagy and apoptosis, while it induced cell cycle arrest in the S-phase in SMMC- 7721 cells. Additionally, down-regulation of cyclin D1, A2, E1, Cdk2 and up-regulation of p21, p27 were detected after the treatment with clioquinol. The results demonstrated for the first time that clioquinol suppressed cell cycle progression in the S-phase in SMMC-7721 cells via the p21, p27-cyclin E,A/Cdk2 pathway. This suggests that clioquinol may have a therapeutic potential as an anticancer drug for certain malignances.


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