scholarly journals STAU2 Protein Level is Controlled by Caspases and the CHK1 Pathway and Regulates Cell Cycle Progression in the Non-transformed hTERT-RPE1 Cells

2020 ◽  
Author(s):  
Lionel Conde ◽  
Remy Beaujois ◽  
Luc DesGroseillers
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cell Proliferation ◽  
Dna Damage ◽  
Steady State ◽  
Posttranscriptional Regulation ◽  
Cell Cycle Progression ◽  
Rna Binding ◽  
Regulation Of Gene Expression ◽  
Cycle Progression

Abstract Background: Staufen2 (STAU2) is an RNA binding protein involved in the posttranscriptional regulation of gene expression. In neurons, STAU2 is required to maintain the balance between differentiation and proliferation of neural stem cells through asymmetric cell division. However, the importance of controlling STAU2 expression for cell cycle progression is not clear in non-neuronal dividing cells. We recently showed that STAU2 transcription is inhibited in response to DNA-damage due to E2F1 displacement from the STAU2 gene promoter. We now study the regulation of STAU2 steady-state levels in unstressed cells and its consequence for cell proliferation. Results: CRISPR/Cas9-mediated and RNAi-dependent STAU2 depletion in the non-transformed hTERT-RPE1 cells both facilitate cell proliferation suggesting that STAU2 expression influences pathway(s) linked to cell cycle controls. Such effects are not observed in the CRISPR STAU2-KO cancer HCT116 cells nor in the STAU2-RNAi-depleted HeLa cells. Interestingly, a physiological decrease in the steady-state level of STAU2 is controlled by caspases. This effect of peptidases is counterbalanced by the activity of the CHK1 pathway suggesting that STAU2 partial degradation/stabilization fines tune cell cycle progression in unstressed cells. A large-scale proteomic analysis using STAU2/biotinylase fusion protein identifies known STAU2 interactors involved in RNA translation, localization, splicing, or decay confirming the role of STAU2 in the posttranscriptional regulation of gene expression. In addition, several proteins found in the nucleolus, including proteins of the ribosome biogenesis pathway and of the DNA damage response, are found in close proximity to STAU2. Strikingly, many of these proteins are linked to the kinase CHK1 pathway, reinforcing the link between STAU2 functions and the CHK1 pathway. Indeed, inhibition of the CHK1 pathway dissociates STAU1 from proteins involved in translation and RNA metabolism. Conclusions: These results indicate that STAU2 is involved in pathway(s) that control(s) cell proliferation, likely via mechanisms of posttranscriptional regulation, ribonucleoprotein complex assembly, genome integrity and/or checkpoint controls. This novel function of STAU2 is regulated by caspases and by the kinase CHK1 pathway.

scholarly journals STAU2 protein level is controlled by caspases and the CHK1 pathway and regulates cell cycle progression in the non-transformed hTERT-RPE1 cells

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Lionel Condé ◽  
Yulemi Gonzalez Quesada ◽  
Florence Bonnet-Magnaval ◽  
Rémy Beaujois ◽  
Luc DesGroseillers
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cell Proliferation ◽  
Dna Damage ◽  
Steady State ◽  
Posttranscriptional Regulation ◽  
Cell Cycle Progression ◽  
Rna Binding ◽  
Regulation Of Gene Expression ◽  
Cycle Progression

AbstractBackgroundStaufen2 (STAU2) is an RNA binding protein involved in the posttranscriptional regulation of gene expression. In neurons, STAU2 is required to maintain the balance between differentiation and proliferation of neural stem cells through asymmetric cell division. However, the importance of controlling STAU2 expression for cell cycle progression is not clear in non-neuronal dividing cells. We recently showed that STAU2 transcription is inhibited in response to DNA-damage due to E2F1 displacement from theSTAU2gene promoter. We now study the regulation of STAU2 steady-state levels in unstressed cells and its consequence for cell proliferation.ResultsCRISPR/Cas9-mediated and RNAi-dependent STAU2 depletion in the non-transformed hTERT-RPE1 cells both facilitate cell proliferation suggesting that STAU2 expression influences pathway(s) linked to cell cycle controls. Such effects are not observed in the CRISPR STAU2-KO cancer HCT116 cells nor in the STAU2-RNAi-depleted HeLa cells. Interestingly, a physiological decrease in the steady-state level of STAU2 is controlled by caspases. This effect of peptidases is counterbalanced by the activity of the CHK1 pathway suggesting that STAU2 partial degradation/stabilization fines tune cell cycle progression in unstressed cells. A large-scale proteomic analysis using STAU2/biotinylase fusion protein identifies known STAU2 interactors involved in RNA translation, localization, splicing, or decay confirming the role of STAU2 in the posttranscriptional regulation of gene expression. In addition, several proteins found in the nucleolus, including proteins of the ribosome biogenesis pathway and of the DNA damage response, are found in close proximity to STAU2. Strikingly, many of these proteins are linked to the kinase CHK1 pathway, reinforcing the link between STAU2 functions and the CHK1 pathway. Indeed, inhibition of the CHK1 pathway for 4 h dissociates STAU2 from proteins involved in translation and RNA metabolism.ConclusionsThese results indicate that STAU2 is involved in pathway(s) that control(s) cell proliferation, likely via mechanisms of posttranscriptional regulation, ribonucleoprotein complex assembly, genome integrity and/or checkpoint controls. The mechanism by which STAU2 regulates cell growth likely involves caspases and the kinase CHK1 pathway.

scholarly journals RB, p130 and p107 differentially repress G1/S and G2/M genes after p53 activation

2019 ◽  
Vol 47 (21) ◽  
pp. 11197-11208 ◽  
Author(s):  
Amy E Schade ◽  
Martin Fischer ◽  
James A DeCaprio
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Dna Damage ◽  
Cell Cycle Progression ◽  
Human Fibroblasts ◽  
Cell Cycle Gene ◽  
Cycle Progression ◽  
Cell Cycle Genes ◽  
P53 Activation ◽  
S Genes

Abstract Cell cycle gene expression occurs in two waves. The G1/S genes encode factors required for DNA synthesis and the G2/M genes contribute to mitosis. The Retinoblastoma protein (RB) and DREAM complex (DP, RB-like, E2F4 and MuvB) cooperate to repress all cell cycle genes during G1 and inhibit entry into the cell cycle. DNA damage activates p53 leading to increased levels of p21 and inhibition of cell cycle progression. Whether the G1/S and G2/M genes are differentially repressed by RB and the RB-like proteins p130 and p107 in response to DNA damage is not known. We performed gene expression profiling of primary human fibroblasts upon DNA damage and assessed the effects on G1/S and G2/M genes. Upon p53 activation, p130 and RB cooperated to repress the G1/S genes. In addition, in the absence of RB and p130, p107 contributed to repression of G1/S genes. In contrast, G2/M genes were repressed by p130 and p107 after p53 activation. Furthermore, repression of G2/M genes by p107 and p130 led to reduced entry into mitosis. Our data demonstrates specific roles for RB, p130-DREAM, and p107-DREAM in p53 and p21 mediated repression of cell cycle genes.

scholarly journals Vaccinia Virus Arrests and Shifts the Cell Cycle

2021 ◽  
Author(s):  
Caroline K. Martin ◽  
Jerzy Samolej ◽  
Annabel T. Olson ◽  
Cosetta Bertoli ◽  
Matthew S. Wiebe ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Dna Damage ◽  
Vaccinia Virus ◽  
Host Cell ◽  
Cell Cycle Progression ◽  
Effector Proteins ◽  
Viral Gene ◽  
Genome Replication ◽  
Cycle Progression

Modulation of the host cell cycle is a common strategy used by viruses to create a pro-replicative environment. To facilitate viral genome replication, vaccinia virus (VACV) has been reported to alter cell cycle regulation and trigger the host cell DNA damage response. However, the cellular factors and viral effectors that mediate these changes remain unknown. Here, we set out to investigate the effect of VACV infection on cell proliferation and host cell cycle progression. Using a subset of VACV mutants we characterize the stage of infection required for inhibition of cell proliferation and define the viral effectors required to dysregulate the host cell cycle. Consistent with previous studies, we show that VACV inhibits, and subsequently shifts the host cell cycle. We demonstrate that these two phenomena are independent of one another, with viral early genes being responsible for cell cycle inhibition, and post-replicative viral gene(s) responsible for the cell cycle shift. Extending previous findings, we show that the viral kinase F10 is required to activate the DNA damage checkpoint and that the viral B1/B12 (pseudo) kinases mediate degradation of checkpoint effectors p53 and p21 during infection. We conclude that VACV modulates host cell proliferation and host cell cycle progression through temporal expression of multiple VACV effector proteins.

Coordinate Changes In RNA Expression, Transcript Splicing, and DNA Methylation In Pediatric AML

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3761-3761
Author(s):  
Jason Farrar ◽  
Michael Ochs ◽  
David W Lee ◽  
C.C. Talbot ◽  
Jonathan Buckley ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Dna Damage ◽  
Alternative Splicing ◽  
Dna Damage Response ◽  
Cell Cycle Progression ◽  
Expression Patterns ◽  
Cycle Progression ◽  
Damage Response ◽  
Pediatric Aml

Abstract While mutations of splicing and epigenetic factors have been reported in adult AML and related myeloid disorders, relatively few such changes have been identified in pediatric AML. We previously identified a chromatin remodeling helicase, PASG (SMARCA6, HELLS, LSH), by down-regulation in AML cell lines following cytokine withdrawal and identified an alternatively spliced variant lacking a highly conserved (STRAGGLG) domain. To assess the prevalence of this splicing variant (PASGΔ75) in pediatric AML, we tested 167 diagnostic specimens from the TARGET-AML cohort for fractional PASGΔ75 expression (PASGΔ75/PASG) using a discriminatory RT-qPCR assay. These studies demonstrated a broad, continuous distribution of PASGΔ75 with right skew (mean PASGΔ75: 26%, interquartile range: 9% – 41%) that was not significantly associated with cytogenetic class (inv16, t(8;21), MLL, normal) or FAB subtype. For further comparison, specimens were quantized by PASGΔ75 quartile. Given reported associations between loss of PASG function and abnormalities of genomic methylation, we tested 48 AML specimens at the extremes of the PASGΔ75 distribution for total 5-methylcytosine (5-mC) content by liquid chromatography/tandem mass spectrometry. The mean total methylation was significantly lower in the high compared to the low PASGΔ75 groups (mean 5-mC 3.95% vs. 4.22% of cytosine, p=0.015 Mann-Whitney). To identify specific regions of altered methylation, we used high-throughput sequencing of DNA enriched by pull-down with the methyl binding domain fragment of MBD2 (MBD-Seq). Comparison of summed methylation signal across regions flanking RefSeq transcriptional start sites (TSS) showed the expected decrease in methylation just upstream of the TSS in both groups. However, methylation more distal to the TSS was proportionally lower in PASGΔ75 high than PASGΔ75 low samples (Fig 1a). To evaluate methylation at CpG islands (CGI), UCSC CGI were scaled to 500 bp and MBD-Seq data were summed across 20Kb flanking CGI. While both groups showed the anticipated increase in methylation signal on CGI, methylation in the shore regions immediately flanking CGI was proportionally lower in high PASGΔ75 compared to low PASGΔ75 samples (Fig 1b), further suggesting epigenetic differences between these sample groups. Because we were unable to identify sequence variants in PASG intron 18 or flanking exons that might explain alternative splicing, we asked whether expression of PASGΔ75 was associated with global changes in transcript splicing. We evaluated gene expression patterns on the Affymetrix HuGene array, with assessment of alternative splicing using Partek software alternative splicing (altsplice) algorithm for quartile-grouped samples. In contrast to comparison of adjacent quartile groups, which showed modest changes in expression and relatively few transcripts with significant altsplice scores, comparison of the highest and lowest quartile samples showed marked changes in gene expression and a large number of alternatively spiced transcripts as assessed by significance of the altsplice score (Fig 2). In addition to splicing changes, these analyses suggested marked differences in gene expression patterns of AML specimens grouped by PASGΔ75 quartile, with clear separation of Q1 and Q4 samples by principal components analysis. Using a conservative Wilcoxon gene sets test and limiting ourselves to small, curated Biocarta pathways, we found expression patterns associated with high deletion variant expression strongly linked to overlapping pathways involving DNA repair, replication, and cell cycle progression. Table Pathways (Biocarta) Pathway Class Benjamini Hochberg Corrected p-Value ATR/BRCA1/BRCA2 DNA Damage Response 1.3E-6 RB/DNA Damage DNA Damage Response 3.5E-3 p27 Phosphorylation Cell Cycle Progression 3.5E-3 G2/M Checkpoint Cell Cycle Progression 3.5E-3 G1/S Checkpoint Cell Cycle Progression 4.1E-3 PLK3 Cell Cycle Progression 4.1E-3 MEF2D Apoptosis 0.01 SRC/PTPa Cell Cycle Progression 0.02 Mitochondrial Acetyl-Co Shuttle Metabolism 0.02 p53 Signaling DNA Damage Response 0.04 E2F-1 Cell Cycle Progression 0.04 ATM Signaling DNA Damage Response 0.04 These data suggest the existence of a previously unrecognized AML subclass characterized by widespread and coordinated changes in RNA expression, alternative transcript splicing, and epigenetic modifications. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Cell cycle progression inCaulobacterrequires a nucleoid-associated protein with high AT sequence recognition

2016 ◽  
Vol 113 (40) ◽  
pp. E5952-E5961 ◽  
Author(s):  
Dante P. Ricci ◽  
Michael D. Melfi ◽  
Keren Lasker ◽  
David L. Dill ◽  
Harley H. McAdams ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Binding Sites ◽  
Cell Cycle Progression ◽  
Target Genes ◽  
Caulobacter Crescentus ◽  
Regulation Of Gene Expression ◽  
Cycle Progression ◽  
Swarmer Cell ◽  
Sequence Recognition

Faithful cell cycle progression in the dimorphic bacteriumCaulobacter crescentusrequires spatiotemporal regulation of gene expression and cell pole differentiation. We discovered an essential DNA-associated protein, GapR, that is required forCaulobactergrowth and asymmetric division. GapR interacts with adenine and thymine (AT)-rich chromosomal loci, associates with the promoter regions of cell cycle-regulated genes, and shares hundreds of recognition sites in common with known master regulators of cell cycle-dependent gene expression. GapR target loci are especially enriched in binding sites for the transcription factors GcrA and CtrA and overlap with nearly all of the binding sites for MucR1, a regulator that controls the establishment of swarmer cell fate. Despite constitutive synthesis, GapR accumulates preferentially in the swarmer compartment of the predivisional cell. Homologs of GapR, which are ubiquitous among the α-proteobacteria and are encoded on multiple bacteriophage genomes, also accumulate in the predivisional cell swarmer compartment when expressed inCaulobacter. TheEscherichia colinucleoid-associated protein H-NS, like GapR, selectively associates with AT-rich DNA, yet it does not localize preferentially to the swarmer compartment when expressed exogenously inCaulobacter, suggesting that recognition of AT-rich DNA is not sufficient for the asymmetric accumulation of GapR. Further, GapR does not silence the expression of H-NS target genes when expressed inE. coli, suggesting that GapR and H-NS have distinct functions. We propose thatCaulobacterhas co-opted a nucleoid-associated protein with high AT recognition to serve as a mediator of cell cycle progression.

scholarly journals RNA-binding protein CELF6 is cell cycle regulated and controls cancer cell proliferation by stabilizing p21

2019 ◽  
Vol 10 (10) ◽  
Author(s):  
Gang Liu ◽  
Qianwen Zhang ◽  
Li Xia ◽  
Mengjuan Shi ◽  
Jin Cai ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Cancer Cell ◽  
Cell Cycle Progression ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Cancer Cell Proliferation ◽  
Cycle Progression ◽  
Specific Alternative ◽  
Cycle Dependent

Abstract CELF6, a member of the CELF family of RNA-binding proteins, regulates muscle-specific alternative splicing and contributes to the pathogenesis of myotonic dystrophy (DM), however the role of CELF6 in cancer cell proliferation is less appreciated. Here, we show that the expression of CELF6 is cell cycle regulated. The cell cycle-dependent expression of CELF6 is mediated through the ubiquitin-proteasome pathway, SCF-β-TrCP recognizes a nonphospho motif in CELF6 and regulates its proteasomal degradation. Overexpression or depletion of CELF6 modulates p21 gene expression. CELF6 binds to the 3′UTR of p21 transcript and increases its mRNA stability. Depletion of CELF6 promotes cell cycle progression, cell proliferation and colony formation whereas overexpression of CELF6 induces G1 phase arrest. The effect of CELF6 on cell proliferation is p53 and/or p21 dependent. Collectively, these data demonstrate that CELF6 might be a potential tumor suppressor, CELF6 regulates cell proliferation and cell cycle progression via modulating p21 stability.

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