Phosphorylation of MCM3 Protein by Cyclin E/Cyclin-dependent Kinase 2 (Cdk2) Regulates Its Function in Cell Cycle

Li, Junhui; Deng, Min; Qian, Wei; Liu, Ting; Tong, Xiaomei; Ye, Xin

doi:10.1074/jbc.m111.226464

Cited by 40 publications

(37 citation statements)

References 38 publications

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“…For gene knockdown analysis, cells were harvested 48 h posttransfection and specific protein levels were analyzed by immunoblotting. For cell cycle analysis, 24 h after transfection, cells were treated with bleomycin (4 g/ml) for an additional 24 h. For polyploidy analysis, cells were blocked in G 1 phase with thymidine for 16 h after transfection with 20 nM siRNA for 24 h, and then cells were treated with bleomycin (4 g/ml) for an additional 48 h. The siRNA duplexes were as follows: for the small interfering WDHD1-1 (si-WDHD1-1) sense strand, 5=-GCAUGUACCCUAAGAAUAA-3=; for the si-WDHD1-2 sense strand, 5=-GCAAAGUUAUGGAAAGUAU-3=; for the si-MCM3 sense strand, 5=-GCATTGTCACTAAATGTTCTCTAGT-3= (27); and for the negative-control (NC) siRNA sense strand, 5=-UUCUCCGAA CGUGUCACGU-3=.…”

Section: Methodsmentioning

confidence: 99%

Role of WDHD1 in Human Papillomavirus-Mediated Oncogenesis Identified by Transcriptional Profiling of E7-Expressing Cells

Yang

Zhang

Gao

et al. 2016

J Virol

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The E7 oncoprotein of the high-risk human papillomavirus (HPV) plays a major role in HPV-induced carcinogenesis. E7 abrogates the G 1 cell cycle checkpoint and induces genomic instability, but the mechanism is not fully understood. In this study, we performed RNA sequencing (RNA-seq) to characterize the transcriptional profile of keratinocytes expressing HPV 16 (HPV-16) E7. At the transcriptome level, 236 genes were differentially expressed between E7 and vector control cells. A subset of the differentially expressed genes, most of them novel to E7-expressing cells, was further confirmed by real-time PCR. Of interest, the activities of multiple transcription factors were altered in E7-expressing cells. Through bioinformatics analysis, pathways altered in E7-expressing cells were investigated. The upregulated genes were enriched in cell cycle and DNA replication, as well as in the DNA metabolic process, transcription, DNA damage, DNA repair, and nucleotide metabolism. Specifically, we focused our studies on the gene encoding WDHD1 (WD repeat and high mobility group [HMG]-box DNA-binding protein), one of the genes that was upregulated in E7-expressing cells. WDHD1 is a component of the replisome that regulates DNA replication. Recent studies suggest that WDHD1 may also function as a DNA replication initiation factor as well as a G 1 checkpoint regulator. We found that in E7-expressing cells, the steady-state level of WDHD1 protein was increased along with the half-life. Moreover, downregulation of WDHD1 reduced E7-induced G 1 checkpoint abrogation and rereplication, demonstrating a novel function for WDHD1. These studies shed light on mechanisms by which HPV induces genomic instability and have therapeutic implications. IMPORTANCEThe high-risk HPV types induce cervical cancer and encode an E7 oncoprotein that plays a major role in HPV-induced carcinogenesis. However, the mechanism by which E7 induces carcinogenesis is not fully understood; specific anti-HPV agents are not available. In this study, we performed RNA-seq to characterize transcriptional profiling of keratinocytes expressing HPV-16 E7 and identified more than 200 genes that were differentially expressed between E7 and vector control cells. Through bioinformatics analysis, pathways altered in E7-expressing cells were identified. Significantly, the WDHD1 gene, one of the genes that is upregulated in E7-expressing cells, was found to play an important role in E7-induced G 1 checkpoint abrogation and rereplication. These studies shed light on mechanisms by which HPV induces genomic instability and have therapeutic implications. H uman papillomaviruses (HPVs) are small DNA viruses that replicate in squamous epithelia. Specific types of HPV (highrisk HPVs) are the causative agents for cervical and several other cancers (1). The transforming properties of high-risk HPVs such as HPV 16 (HPV-16) primarily depend on E7 as well as E6 oncogenes (1, 2). HPV E6 and E7 proteins promote the degradation of p53 and pRb, respectively (3, 4). E7 from the high-risk...

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Section: Methodsmentioning

confidence: 99%

Role of WDHD1 in Human Papillomavirus-Mediated Oncogenesis Identified by Transcriptional Profiling of E7-Expressing Cells

Yang

Zhang

Gao

et al. 2016

J Virol

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“…E-4), ␤-actin catalog no. [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19], and lamin B (catalog no. M-20) antibodies were purchased from Santa Cruz Biotechnology.…”

Section: Methodsmentioning

confidence: 99%

“…NPAT is phosphorylated by the cyclin E-CDK2 complex to increase histone transcription required for S phase entry and progression (13). In addition to its function in promoting G 1 /S transition, cyclin E-CDK2 is also involved in DNA damage checkpoint control by phosphorylating the DNA helicase complex subunits MCM3 and MCM7 (14,15).…”

mentioning

confidence: 99%

Cyclin E-CDK2 Protein Phosphorylates Plant Homeodomain Finger Protein 8 (PHF8) and Regulates Its Function in the Cell Cycle

Sun

Huang

Qian

et al. 2015

Journal of Biological Chemistry

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Background: Cyclin E-CDK2 is a key protein kinase in the cell cycle. Results: Phosphorylation of PHF8 by cyclin E-CDK2 enhances its demethylase activity toward H3K9me2 and promotes rDNA transcription as well as S phase progression. Conclusion: Cyclin E-CDK2-dependent phosphorylation of PHF8 affects its function. Significance: We unveiled the mechanism of how cyclin E-CDK2 regulates the function of PHF8 in cell cycle regulation.

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“…As the degree of DNA replication regulation may be another marker of cell proliferation, MCM protein expression may present an extremely promising tool for the evaluation of cell cycle progression [20,21]. The MCM family may act as potential diagnostic and prognostic markers of various tumors [22].…”

Section: Discussionmentioning

confidence: 99%

Comparison between immunohistochemical expression of Ki-67 and MCM-3 in major salivary gland epithelial tumors in children and adolescents. Preliminary study

Zieliński¹,

Kobos²,

Zakrzewska³

2016

pjp

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While Ki-67 expression is frequently used as an indicator of tumor cell proliferation, alternative markers have also been proposed. Possible alternative indicators of proliferation are the minichromosome maintenance (MCM) proteins, whose levels are inversely associated with tumor cell differentiation. The aim of this preliminary study was to compare the levels of Ki-67 and MCM-3 expression in major salivary gland epithelial tumors in all children and adolescents who underwent surgery in our department in the years 2009-2014. The histopathological diagnosis of the subjects was reviewed, as well as the expression of Ki-67 and MCM-3 in post-op specimens of the tumors. The normality of data was checked with the Shapiro-Wilk test. The t test for independent variables or the U test was used as appropriate to determine statistically significant differences in the expression of Ki-67 and MCM-3. Five cases of pleomorphic adenoma, one of myoepithelioma, one of basal cell adenoma and one of mucoepidermoid carcinoma were identified. Significantly greater MCM-3 than Ki-67 expression was observed in every case. The results of our preliminary study emphasize the need for future research on MCM-3 as a sensitive proliferation marker, providing an alternative to Ki-67, in cases of various major salivary gland epithelial tumors in children and adolescents.

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Phosphorylation of MCM3 Protein by Cyclin E/Cyclin-dependent Kinase 2 (Cdk2) Regulates Its Function in Cell Cycle

Cited by 40 publications

References 38 publications

Role of WDHD1 in Human Papillomavirus-Mediated Oncogenesis Identified by Transcriptional Profiling of E7-Expressing Cells

Role of WDHD1 in Human Papillomavirus-Mediated Oncogenesis Identified by Transcriptional Profiling of E7-Expressing Cells

Cyclin E-CDK2 Protein Phosphorylates Plant Homeodomain Finger Protein 8 (PHF8) and Regulates Its Function in the Cell Cycle

Comparison between immunohistochemical expression of Ki-67 and MCM-3 in major salivary gland epithelial tumors in children and adolescents. Preliminary study

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