Different regulation of chromatin modification guide cellular reprogramming into a rate-limited step after initiation phase

Background. In the early and late stages of cell reprogramming to induced pluripotent stem cells (iPSCs) ectopic expression of Oct4, Sox2, Klf4 and Myc (OSKM) aroused two peaks of transcriptional and epigenetic change respectively. However, it was relatively quiet in the intermediate stage. In this paper our aim is to gain insight into the molecular events that occur after the initiation phase of pluripotency induction. Methods. GSE42379 containing 28 gene expression profiles and GSE42477 containing 10 genome binding/occupancy profiles of mouse embryonic fibroblasts (MEF) were downloaded from GEO. These datasets included untreated MEFs, OSKM-induced MEFs progressing and refractory to reprogram at 3, 6, 9, 12 day post-transduction, and iPS cell lines. Differentially expressed genes (DEGs) were identified between different cell lines. The Chip-seq peaks and putative target gene were obtained from GeneProf website. Gene ontology analysis was performed on the Ensemble website. Result. Compared with the progressing cells, the refractory cells obtained more than two times DEGs at 6 day post-transduction, in particular, down-regulated DEGs related to cell cycle, cell adhesion and development were over 4 times of that in progressing cells. The expression of the DEGs which could only be detected in refractory cells at 6 day were traced back to day 3, and we found the expression of the up-regulated DEGs at 3 day were higher in refractory cells, whereas the expression of the down-regulated DEGs at 3 day were lower in refractory cells. The analysis of histone modification in genome-wide and in DEGs showed that during the reprogramming process the increase of bivalent sites in genome were mainly attributed to gaining of H3K27me3 and losing of H3K4me3. Different regulation of H3K27me3 in DEGs was the key to regulate the expression differently in progressing and refractory cells. The expression of chromatin modifiers in the two cell populations were checked and found to be differential regulated at different time point during reprogramming process. Conclusion. Genes related to immune response, cell adhesion, DNA replication and cell cycle in the refractory cells responded to the induction factor earlier than in the progressing cells, which led to excessive conversion rate. We supposed that after initiation phase cellular reprogramming required to undergo a rate-limited step guided by different regulation of chromatin modification.

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Different regulation of chromatin modification guide cellular reprogramming into a rate-limited step after initiation phase

10.7287/peerj.preprints.1991v1 ◽

2016 ◽

Author(s):

Rong Hu ◽

Xianhua Dai ◽

Zhiming Dai ◽

Qian Xiang

Keyword(s):

Cell Cycle ◽

Cell Adhesion ◽

Cell Lines ◽

Expression Profiles ◽

Chromatin Modification ◽

Ectopic Expression ◽

Gene Expression Profiles ◽

Cellular Reprogramming ◽

Ips Cell ◽

Initiation Phase

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Gene Expression-Based Chemical Genomics Identifies Valproic Acid to Revert the Oncogenic Effect of GATA1s In Down Syndrome Megakaryoblastic Leukemia.

Blood ◽

10.1182/blood.v116.21.3646.3646 ◽

2010 ◽

Vol 116 (21) ◽

pp. 3646-3646

Author(s):

Kirsten Heitmann ◽

Zhe Li ◽

Jan-Henning Klusmann ◽

Basant Kumar Thakur ◽

Jennifer Schöning ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Valproic Acid ◽

Down Syndrome ◽

Progenitor Cells ◽

Cell Lines ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Megakaryoblastic Leukemia ◽

Chemical Genomic

Abstract Abstract 3646 Children with Down syndrome (DS) are at high risk to develop acute megakaryoblastic leukemia (DS-AMKL) and the antecedent transient leukemia (DS-TL). Acquired mutations in the hematopoietic transcription factor GATA1, leading to expression of a shorter GATA1 variant (referred to as GATA1s) truncated at its N-terminus, are consistently present in the affected cells of children with DS-AMKL and DS-TL. Mechanistically, we recently found that in fetal megakaryocytic progenitor cells, GATA1 coordinates proliferation and differentiation by repressing E2F target genes through a direct interaction with E2F activators. Failure of this GATA1-E2F interaction in mutated GATA1s likely converges with overactive IGF signaling to promote cellular transformation. The treatment of DS-AMKL is hampered by their sensitivity against current cytostatic agents, resulting in treatment-related mortality as the main cause of death. To develop novel targeted and less toxic treatment options for DS-AMKL and DS-TL, we conducted a gene expression-based chemical genomic screen. We connected a DS-AMKL gene expression signature (compared to non-DS-AMKL, i.e. GATA1s vs. GATA1) to a reference collection of gene-expression profiles from cultured human cells treated with bioactive small molecules (Connectivity Map). We discovered the histone deacetylase (HDAC) inhibitor valproic acid (VPA) reverses the DS-AMKL gene expression program. Cell viability assays, cell cycle analyses, growths curves and colony-forming assays revealed exceptional sensitivity of DS-AMKL cell lines (CMK, CMY; IC50 1mM) and primary DS-AMKL and DS-TL blasts to VPA treatment compared to control cell lines K562 (IC50 4.75mM), M07 (IC50 6.75mM) and CD34+ hematopoietic stem and progenitor cells (IC50 4.75mM). VPA induces apoptosis (26.8% 7-AAD-)Annexin V+ and 38.8% 7-AAD+ CMK cells after 48h at 2mM VPA) and cell cycle arrest (50% reduction of CMK cells in S-phase at 2mM) via activation of the cell cycle inhibitor p21 and the proapoptotic genes BAX and BAK. Gene expression profiles indicated that VPA interferes with the oncogenic effects of GATA1s by globally repressing the deregulated E2F targets. The effects of VPA on leukemic growth in DS-AMKL could be attributed to its HDAC inhibitory function, as the global HDAC inhibitors SAHA and TSA induced a similar response. Thus, by using a gene expression-based chemical genomic approach, we identified VPA as an efficient and well-tolerated treatment option for DS-AMKL and DS-TL by targeting GATA1s-mediated deregulation of the E2F transcription network. Disclosures: No relevant conflicts of interest to declare.

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X-Reactivation Impacts Human iPSC Differentiation Potential Towards Blood

Blood ◽

10.1182/blood.v122.21.4838.4838 ◽

2013 ◽

Vol 122 (21) ◽

pp. 4838-4838

Author(s):

Roksana Moraghebi ◽

Agnete Kirkeby ◽

Marcus Larsson ◽

Andreas Herbst ◽

Malin Parmar ◽

...

Keyword(s):

Gene Expression ◽

Cell Differentiation ◽

Amniotic Fluid ◽

Cell Lines ◽

X Chromosome ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Differentiation Potential ◽

Ips Cell ◽

Differentiation Capacity

Abstract To determine novel key regulators that direct ES/iPS cell differentiation to hematopoietic lineages, we compared the gene expression profiles of multiple iPS cell lines with defferential blood forming capacity. We generated multiple iPS cell lines from amniotic fluid derived mesenchymal stromal cells (AF-iPS) which differentiated towards hematopoietic lineages using our standardized and highly reproducible differentiation protocol. Of the 9 AF-iPS cell lines derived from an individual female patient, the average efficiency of CD45+ hematopoietic cells was 14.2 +/- 9% (range 1.6 to 26.3%). To elucidate the possible reasons for this diversity in efficiency, we grouped the AF-iPS cell lines on the basis of lowest and highest blood differentiation capacity and compared their gene expression profiles by microarray. We found very few changes above 1.5-fold, but interestingly, among the 11 genes that were over-expressed in the AF-iPSC lines with poor blood differentiation efficiency 10 were located on X chromosome, and the remaining one reported to be involved in Notch signalling. A combination of cumulative sum analysis and the location of differentially expressed genes on the X chromosome identified putative regions of reactivation at multiple, but distinct locations. The possibility of X-reactivation in these female lines was reinforced further where lower levels of XIST were seen in AF-iPSC lines shown to have low blood forming potential, however only half of the iPS cell lines with high blood differentiation capacity showed normal XIST expression when compared to the amniotic fluid mesenchymal starting cell material. To determine whether the block in differentiation was tissue specific we tested the differentiation capacity of the AF-iPSC lines towards neuronal lineages. Intriguingly we found neural cell differentiation was not hampered within all lines with poor blood potential suggesting that the over-expression of genes as a consequence of X-reactivation can impart a specific negative effect on differentiation towards the blood lineages from pluripotency stage, while not having an effect on neuronal cell development. To further define the source of this block, we have begun working knocking down the overexpressed genes on X chromosome in lines with poor blood differentiation potential to determine whether the efficiency can be increased (or fully rescued) with one, or a combination of these 11 candidate genes. These results have implications for the identification and selection of female iPS lines suitable for therapeutic purposes. Disclosures: No relevant conflicts of interest to declare.

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Genome-Wide Role of HSF1 in Transcriptional Regulation of Desiccation Tolerance in the Anhydrobiotic Cell Line, Pv11

International Journal of Molecular Sciences ◽

10.3390/ijms22115798 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5798

Author(s):

Shoko Tokumoto ◽

Yugo Miyata ◽

Ruslan Deviatiiarov ◽

Takahiro G. Yamada ◽

Yusuke Hiki ◽

...

Keyword(s):

Cell Line ◽

Cell Lines ◽

Desiccation Tolerance ◽

Expression Profiles ◽

Insect Cell Line ◽

Gene Expression Profiles ◽

Late Embryogenesis Abundant ◽

Heat Shock Factor 1 ◽

Genome Wide ◽

A Genome

The Pv11, an insect cell line established from the midge Polypedilum vanderplanki, is capable of extreme hypometabolic desiccation tolerance, so-called anhydrobiosis. We previously discovered that heat shock factor 1 (HSF1) contributes to the acquisition of desiccation tolerance by Pv11 cells, but the mechanistic details have yet to be elucidated. Here, by analyzing the gene expression profiles of newly established HSF1-knockout and -rescue cell lines, we show that HSF1 has a genome-wide effect on gene regulation in Pv11. The HSF1-knockout cells exhibit a reduced desiccation survival rate, but this is completely restored in HSF1-rescue cells. By comparing mRNA profiles of the two cell lines, we reveal that HSF1 induces anhydrobiosis-related genes, especially genes encoding late embryogenesis abundant proteins and thioredoxins, but represses a group of genes involved in basal cellular processes, thus promoting an extreme hypometabolism state in the cell. In addition, HSF1 binding motifs are enriched in the promoters of anhydrobiosis-related genes and we demonstrate binding of HSF1 to these promoters by ChIP-qPCR. Thus, HSF1 directly regulates the transcription of anhydrobiosis-related genes and consequently plays a pivotal role in the induction of anhydrobiotic ability in Pv11 cells.

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The cell type-specific effect of TAp73 isoforms on the cell cycle and apoptosis

Cellular & Molecular Biology Letters ◽

10.2478/s11658-008-0011-z ◽

2008 ◽

Vol 13 (3) ◽

Cited By ~ 9

Author(s):

Jitka Holcakova ◽

Pavla Ceskova ◽

Roman Hrstka ◽

Petr Muller ◽

Lenka Dubska ◽

...

Keyword(s):

Cell Cycle ◽

Cancer Patients ◽

Transcriptional Activity ◽

Human Cancer ◽

Binding Capacity ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Specific Effect ◽

P53 Family ◽

Null Cell

Abstractp73, a member of the p53 family, exhibits activities similar to those of p53, including the ability to induce growth arrest and apoptosis. p73 influences chemotherapeutic responses in human cancer patients, in association with p53. Alternative splicing of the TP73 gene produces many p73 C- and N-terminal isoforms, which vary in their transcriptional activity towards p53-responsive promoters. In this paper, we show that the C-terminal spliced isoforms of the p73 protein differ in their DNA-binding capacity, but this is not an accurate predictor of transcriptional activity. In different p53-null cell lines, p73β induces either mitochondrial-associated or death receptor-mediated apoptosis, and these differences are reflected in different gene expression profiles. In addition, p73 induces cell cycle arrest and p21WAF1 expression in H1299 cells, but not in Saos-2. This data shows that TAp73 isoforms act differently depending on the tumour cell background, and have important implications for p73-mediated therapeutic responses in individual human cancer patients.

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Comprehensive analysis of the gene expression profiles in human gastric cancer cell lines

Oncogene ◽

10.1038/sj.onc.1205829 ◽

2002 ◽

Vol 21 (42) ◽

pp. 6549-6556 ◽

Cited By ~ 39

Author(s):

Jiafu Ji ◽

Xin Chen ◽

Suet Yi Leung ◽

Jen-Tsan A Chi ◽

Kent Man Chu ◽

...

Keyword(s):

Gene Expression ◽

Gastric Cancer ◽

Cell Lines ◽

Cancer Cell ◽

Gastric Cancer Cell ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Comprehensive Analysis ◽

Human Gastric Cancer ◽

Gastric Cancer Cell Lines

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Baicalein, a Natural Anti-Cancer Compound, Alters MicroRNA Expression Profiles in Bel-7402 Human Hepatocellular Carcinoma Cells

Cellular Physiology and Biochemistry ◽

10.1159/000470815 ◽

2017 ◽

Vol 41 (4) ◽

pp. 1519-1531 ◽

Cited By ~ 22

Author(s):

Beibei Bie ◽

Jin Sun ◽

Jun Li ◽

Ying Guo ◽

Wei Jiang ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Cell Cycle ◽

Cell Proliferation ◽

Cell Lines ◽

Mirna Expression ◽

Expression Profiles ◽

Akt Pathway ◽

Qrt Pcr ◽

Cycle Arrest ◽

Mirna Expression Profiles

Background/Aims: Baicalein has been shown to possess significant anti-hepatoma activity by inhibiting cell proliferation. Whether the anti-proliferative effect of baicalein is related to its modulation of miRNA expression in hepatocellular carcinoma (HCC) is still unknown. Methods: The anti-proliferative effects of baicalein on HCC cell line Bel-7402 was assessed by detecting the proliferation activity, cell cycle distribution, expression changes of p21/CDKN1A, P27/CDKN1B, total Akt and phosphoryted AKT. Microarray analysis was conducted to determine the miRNA expression profiles in baicalein-treated or untreated Bel-7402 cells and then validated by qRT-PCR in two HCC cell lines (Bel-7402 and Hep3B). The gain-of-function of miR-3127-5p was performed by detecting anti-proliferative effects after transfecting miRNA mimics in cells. Finally, the expression level of miR-3127-5p in different HCC cell lines was determined by qRT-PCR. Results: Baicalein was able to inhibit the proliferation of Bel-7402 cells by inducing cell cycle arrest at the S and G2/M phase via up-regulating the expression of p21/CDKN1A and P27/CDKN1B and suppressing the PI3K/Akt pathway. Baicalein could alter the miRNA expression profiles in Bel-7402 cells. Putative target genes for differentially expressed miRNAs could be enriched in terms of cell proliferation regulation, cell cycle arrest and were mainly involved in MAPK, PI3K-Akt, Wnt, Hippo and mTOR signaling pathways. MiR- 3127-5p, one of up-regulated miRNAs, exhibits low expression level in several HCC cell lines and its overexpression could inhibit cell growth of Bel-7402 and Hep3B cell lines by inducing S phase arrest by up-regulating the expression of p21and P27 and repressing the PI3K/Akt pathway. Conclusions: Modulation of miRNA expression may be an important mechanism underlying the anti-hepatoma effects of baicalein.

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Elucidating the expression and function of Numbl during cell adhesion-mediated drug resistance (CAM-DR) in multiple myeloma (MM)

BMC Cancer ◽

10.1186/s12885-019-6446-y ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 1

Author(s):

Yuejiao Huang ◽

Xianting Huang ◽

Chun Cheng ◽

Xiaohong Xu ◽

Hong Liu ◽

...

Keyword(s):

Cell Cycle ◽

Multiple Myeloma ◽

Drug Resistance ◽

Cell Adhesion ◽

Cell Lines ◽

Molecular Mechanisms ◽

Clinical Problem ◽

Integrin Β1 ◽

Cell Viability Assay ◽

Adhesion Model

Abstract Background Cell adhesion-mediated drug resistance (CAM-DR) is a major clinical problem that prevents successful treatment of multiple myeloma (MM). In particular, the expression levels of integrin β1 and its sub-cellular distribution (internalization and trafficking) are strongly associated with CAM-DR development. Methods Development of an adhesion model of established MM cell lines and detection of Numbl and Integrinβ1 expression by Western Blot analysis. The interaction between Numbl and Integrinβ1 was assessed by a co-immunoprecipitation (CO-IP) method. Calcein AM assay was performed to investigate the levels of cell adhesion. Finally, the extent of CAM-DR in myeloma cells was measured using cell viability assay and flow cytometry analysis. Results Our preliminary date suggest that Numbl is differentially expressed in a cell adhesion model of MM cell lines. In addition to binding to the phosphotyrosine-binding (PTB) domain, the carboxyl terminal of Numbl can also interact with integrin β1 to regulate the cell cycle by activating the pro-survival PI3K/AKT signaling pathway. This study intends to verify and elucidate the interaction between Numbl and integrin β1 and its functional outcome on CAM-DR. We have designed and developed a CAM-DR model using MM cells coated with either fibronectin or bone marrow stromal cells. We assessed whether Numbl influences cell-cycle progression and whether it, in turn, contributes to activation of PI3K/AKT signal pathway through the adjustment of its carboxyl end. Finally, we showed that the interaction of Numbl with integrin β1 promotes the formation of CAM-DR in MM cells. Conclusions Our findings elucidated the specific molecular mechanisms of CAM-DR induction and confirmed that Numbl is crucial for the development of CAM-DR in MM cells.

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