Transcriptional Regulation

doi:10.1007/978-1-61779-376-9

Cited by 3 publications

(1 citation statement)

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“…For analysing the binding of C/EBPβ to the promoters of adipogenic genes, ChIP assay was performed as described earlier with minor modifications [23]. Briefly, cells transfected with gene and protein along with untreated cells were cross-linked with 1% formaldehyde for 15min at RT.…”

Section: Chromatin Immunoprecipitation Analysismentioning

confidence: 99%

Transient delivery of A-C/EBP protein perturbs differentiation of 3T3-L1 cells and induces preadipocyte marker genes

Sharma

Kaur

Yadav

et al. 2020

Preprint

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Transformation of committed 3T3-L1 preadipocytes to lipid-laden adipocytes involves timely appearance of numerous transcription factors (TFs), foremost among them is C/EBPβ, is expressed during early phases of differentiation. Here we describe liposome-mediated protein transfection approach to rapidly downregulate C/EBPβ by A-C/EBP protein inhibitor. Signals from tagged A-C/EBP were observed in 3T3-L1 cells within 2hrs of protein inhibitor transfections whereas for gene transfection signals appeared in 48hrs. Following transient transfections, expression profiles of 24 marker genes belonging to pro-and anti-adipogenic, cell cycle, and preadipocytes pathways was analyzed. mRNA and protein expression profiles of adipocyte-marker genes showed lower expression in both A-C/EBP protein and gene transfected samples. Interestingly, for preadipocytes and cell fate determinant genes, striking differences were observed between protein and gene transfected samples. Preadipocyte differentiation factors Stat5a and Creb were downregulated in A-C/EBP protein samples. Five preadipocyte markers, namely, Pdgfrα, Pdgfrβ, Ly6A, CD34 Itgb1 showed high expression in protein samples whereas only Ly6A and CD34 were expressed in gene transfected samples. Pdgfrα and Pdgfrβ, two known cell fate markers were expressed in protein transfected samples 5-days post-differentiation suggesting a possible reversal of differentiation. Our study provides evidences for the robust and efficient knockdown of C/EBPβ protein to understand time-dependent gene regulation during adipogenesis.

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Section: Chromatin Immunoprecipitation Analysismentioning

confidence: 99%

Transient delivery of A-C/EBP protein perturbs differentiation of 3T3-L1 cells and induces preadipocyte marker genes

Sharma

Kaur

Yadav

et al. 2020

Preprint

View full text Add to dashboard Cite

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RNA-Seq Data Analysis in Prokaryotes: A Review for Non-experts

Cubillos¹,

Perlaza-Jiménez²,

Giraldo³

2014

Acta biol. Colom.

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Methyltransferase 3A-mediated promoter methylation represses retinoic acid receptor responder 3 expression in basal-like breast cancer

Tuo¹,

LIU²

2023

Biocell

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Retinoic acid receptor responder 3 (RARRES3) has been characterized as a tumor suppressor in multiple types of cancer. This study aimed to examine the expression profile of RARRES3 across the PAM50 subtypes of breast cancer. The DNA methylation status of RARRES3 was checked in the basal-like subtype, and the underlying mechanisms of its dysregulation were explored. RNA-sequencing (seq) and methylation data from The Cancer Genome Atlas were used for in-silico analysis. Basal-like representative SUM149 and MDA-MB-468 cell lines were used for in vitro and in vivo studies.Compared to tumor-adjacent normal tissues, only the basal-like tumor tissues had significantly downregulated RARRES3 expression. The methylation level of four CpG sites in the promoter region showed a strong negative correlation with RARRES3 expression. The gene coding for DNA methyltransferase 3A (DNMT3A) had consistent positive correlations with the methylation of the CpG sites. Chromatin Immunoprecipitation-quantitative polymerase chain-reaction and bisulfite sequencing PCR showed that DNMT3A could bind to the promoter region of RARRES3 and promote methylation of the CpG sites within the region. DNMT3A knockdown significantly restored RARRES3 expression at the mRNA and protein level in the two cell lines. CCK-8, colony formation, and flow cytometric analysis showed that RARRES3 overexpression attenuated the growth-promoting effects of DNMT3A overexpression and also weakened the DNMT3A overexpression-induced activation of ERK1/2 and PI3K/AKT signaling. In summary, this study revealed that DNMT3A enhances promoter methylation of the RARRES3 gene and suppresses its transcription in basal-like breast cancer. The DNMT3A-RARRES3 signaling pathway might be a potential target for the treatment of this tumor subtype.

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Transcriptional Regulation

Cited by 3 publications

References 0 publications

Transient delivery of A-C/EBP protein perturbs differentiation of 3T3-L1 cells and induces preadipocyte marker genes

Transient delivery of A-C/EBP protein perturbs differentiation of 3T3-L1 cells and induces preadipocyte marker genes

RNA-Seq Data Analysis in Prokaryotes: A Review for Non-experts

Methyltransferase 3A-mediated promoter methylation represses retinoic acid receptor responder 3 expression in basal-like breast cancer

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