Live-cell monitoring of periodic gene expression in synchronous human cells identifies Forkhead genes involved in cell cycle control

Grant, Gavin D.; Gamsby, Joshua J.; Martyanov, Viktor; Brooks, Lionel; George, Lacy K.; Mahoney, James; Loros, Jennifer J.; Dunlap, Jay C.; Whitfield, Michael L.

doi:10.1091/mbc.e11-02-0170

Cited by 30 publications

(28 citation statements)

References 62 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…At this time, the partial dissociation of VprBP from DDB1 and Cul4 (Fig. 5) would enhance its ability to activate FoxM1 (14). Consistent with this prediction, VprBP partially dissociates from CRL4 in mitosis and a mutant version of VprBP that is impaired for DDB1 binding can still activate FoxM1.…”

Section: Discussionsupporting

confidence: 56%

See 1 more Smart Citation

VprBP/DCAF1 Regulates the Degradation and Nonproteolytic Activation of the Cell Cycle Transcription Factor FoxM1

Wang

Arceci

Bird

et al. 2017

Molecular and Cellular Biology

View full text Add to dashboard Cite

The oncogenic transcription factor FoxM1 plays a vital role in cell cycle progression, is activated in numerous human malignancies, and is linked to chromosome instability. We characterize here a cullin 4-based E3 ubiquitin ligase and its substrate receptor, VprBP/DCAF1 (CRL4 VprBP ), which we show regulate FoxM1 ubiquitylation and degradation. Paradoxically, we also found that the substrate receptor VprBP is a potent FoxM1 activator. VprBP depletion reduces expression of FoxM1 target genes and impairs mitotic entry, whereas ectopic VprBP expression strongly activates a FoxM1 transcriptional reporter. VprBP binding to CRL4 is reduced during mitosis, and our data suggest that VprBP activation of FoxM1 is ligase independent. This implies a nonproteolytic activation mechanism that is reminiscent of, yet distinct from, the ubiquitin-dependent transactivation of the oncoprotein Myc by other E3s. Significantly, VprBP protein levels were upregulated in high-grade serous ovarian patient tumors, where the FoxM1 signature is amplified. These data suggest that FoxM1 abundance and activity are controlled by VprBP and highlight the functional repurposing of E3 ligase substrate receptors independent of the ubiquitin system. KEYWORDS cell cycle, cullin ring ligase, FoxM1, transcriptional regulation, ubiquitination C hanges in gene expression combined with targeted protein degradation dynamically shape the protein landscape. Gene expression is coordinated by transcription factors that specify genes for activation and cofactors that modulate transcription factor activity or alter the local chromatin environment. Posttranslational modifications (PTMs) play a crucial role in transcriptional dynamics. Phosphorylation, acetylation, methylation, and ubiquitylation of histone proteins are well studied and contribute significantly to gene expression dynamics (1). Similarly, posttranslational modification of transcription factors plays an important role in regulating genome output.FoxM1 is an oncogenic, cell cycle-regulated transcription factor that was discovered as both a marker and a key mediator of cell proliferation (2-4). Subsequent work clarified the importance of FoxM1 in proliferation through its role in cell cycle progression (reviewed in reference 5). FoxM1 controls the mitotic transcriptional program, and its depletion significantly impairs normal mitotic entry and progression (6-9). In addition, FoxM1 and its transcriptional network have been associated with numerous cancers (5, 10). Notably, FoxM1 is the key regulator of a proliferative gene expression signature found in high-grade serous ovarian cancer (HGSOC), basal-like breast cancers,

show abstract

Section: Discussionsupporting

confidence: 56%

“…FoxM1 activity peaks in G 2 /M phase, consistent with its role in dictating mitotic gene expression, and several kinases have been implicated in its activation (6,(14)(15)(16)(17)(18). FoxM1 is repressed by an intramolecular interaction with its amino-terminal domain, and this inhibition is relieved by cyclin-dependent kinase (CDK) phosphorylation (19).…”

mentioning

confidence: 87%

VprBP/DCAF1 Regulates the Degradation and Nonproteolytic Activation of the Cell Cycle Transcription Factor FoxM1

Wang

Arceci

Bird

et al. 2017

Molecular and Cellular Biology

View full text Add to dashboard Cite

show abstract

“…Classical regulators include the transcription factors E2F1 and FOXM1 that are required for the temporal control of gene expression in G1-S and G2-M stages of cell cycle, respectively [4,[19][20][21]. Given that many of the 1 182 genes identified in this study have not been previously observed as being periodic, we sought to discover novel regulatory mechanisms underlying periodic gene expression.…”

Section: Systematic Analysis Of Periodic Gene Promotersmentioning

confidence: 99%

A high-resolution transcriptome map of cell cycle reveals novel connections between periodic genes and cancer

et al. 2016

View full text Add to dashboard Cite

Progression through the cell cycle is largely dependent on waves of periodic gene expression, and the regulatory networks for these transcriptome dynamics have emerged as critical points of vulnerability in various aspects of tumor biology. Through RNA-sequencing of human cells during two continuous cell cycles (>2.3 billion paired reads), we identified over 1 000 mRNAs, non-coding RNAs and pseudogenes with periodic expression. Periodic transcripts are enriched in functions related to DNA metabolism, mitosis, and DNA damage response, indicating these genes likely represent putative cell cycle regulators. Using our set of periodic genes, we developed a new approach termed "mitotic trait" that can classify primary tumors and normal tissues by their transcriptome similarity to different cell cycle stages. By analyzing >4 000 tumor samples in The Cancer Genome Atlas (TCGA) and other expression data sets, we found that mitotic trait significantly correlates with genetic alterations, tumor subtype and, notably, patient survival. We further defined a core set of 67 genes with robust periodic expression in multiple cell types. Proteins encoded by these genes function as major hubs of protein-protein interaction and are mostly required for cell cycle progression. The core genes also have unique chromatin features including increased levels of CTCF/RAD21 binding and H3K36me3. Loss of these features in uterine and kidney cancers is associated with altered expression of the core 67 genes. Our study suggests new chromatin-associated mechanisms for periodic gene regulation and offers a predictor of cancer patient outcomes.

show abstract

“…Flow cytometry has been broadly utilized for the analysis of biological processes at the single cell level, such as gene expression, cell cycle, apoptosis, and signaling through phosphorylation [37][38][39][40][41][42][43] .The stable expression of fluorescent protein genes in mammalian cells has further enhanced the utility of flow cytometry for cell analysis 38,44 and ligand-receptor interactions 45 . Enhanced capabilities have allowed flow cytometry to become a widely utilized methodology for high-throughput and high-content screening 46 .…”

Section: Introductionmentioning

confidence: 99%

Genetic Barcoding with Fluorescent Proteins for Multiplexed Applications

Smurthwaite

Williams

Fetsko

et al. 2015

JoVE

View full text Add to dashboard Cite

Fluorescent proteins, fluorescent dyes and fluorophores in general have revolutionized the field of molecular cell biology. In particular, the discovery of fluorescent proteins and their genes have enabled the engineering of protein fusions for localization, the analysis of transcriptional activation and translation of proteins of interest, or the general tracking of individual cells and cell populations. The use of fluorescent protein genes in combination with retroviral technology has further allowed the expression of these proteins in mammalian cells in a stable and reliable manner. Shown here is how one can utilize these genes to give cells within a population of cells their own biosignature. As the biosignature is achieved with retroviral technology, cells are barcoded ´indefinitely´. As such, they can be individually tracked within a mixture of barcoded cells and utilized in more complex biological applications. The tracking of distinct populations in a mixture of cells is ideal for multiplexed applications such as discovery of drugs against a multitude of targets or the activation profile of different promoters. The protocol describes how to elegantly develop and amplify barcoded mammalian cells with distinct genetic fluorescent markers, and how to use several markers at once or one marker at different intensities. Finally, the protocol describes how the cells can be further utilized in combination with cell-based assays to increase the power of analysis through multiplexing. Video LinkThe video component of this article can be found at

show abstract

Live-cell monitoring of periodic gene expression in synchronous human cells identifies Forkhead genes involved in cell cycle control

Cited by 30 publications

References 62 publications

VprBP/DCAF1 Regulates the Degradation and Nonproteolytic Activation of the Cell Cycle Transcription Factor FoxM1

VprBP/DCAF1 Regulates the Degradation and Nonproteolytic Activation of the Cell Cycle Transcription Factor FoxM1

A high-resolution transcriptome map of cell cycle reveals novel connections between periodic genes and cancer

Genetic Barcoding with Fluorescent Proteins for Multiplexed Applications

Contact Info

Product

Resources

About