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Gene Expression Analysis in NCI-H520 human cancer cells upon deadenylase silencing

ABSTRACT: The first and rate-limiting step in eukaryotic mRNA decay is the shortening of the poly(A) tail catalyzed by a family of enzymes known as deadenylases. In humans, several deadenylases have been recognized so far, yet it is not clear what the advantage is to have many enzymes catalyzing the same reaction. It is hypothesized that specific deadenylases may target unique subsets of mRNAs, or multiple deadenylases can act on the same mRNA, with discrete but overlapping functions. To understand the biological significance of the diversity of these enzymes we silenced the expression of several deadenylases, including PARN, NOC, CNOT6, CNOT6L, and CNOT7, in human cells of cancer origin (NCI-H520; squamous lung cancer), and analyze the impact on gene expression with microarrays.

ORGANISM(S): Homo sapiens

PROVIDER: GSE67598 | GEO | 2015/10/01

SECONDARY ACCESSION(S): PRJNA280404

REPOSITORIES: GEO

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Gene Expression Analysis in Hep2 human cancer cells upon deadenylase silencing

Project description:The first and rate-limiting step in eukaryotic mRNA decay is the shortening of the poly(A) tail catalyzed by a family of enzymes known as deadenylases. In humans, several deadenylases have been recognized so far, yet it is not clear what the advantage is to have many enzymes catalyzing the same reaction. It is hypothesized that specific deadenylases may target unique subsets of mRNAs, or multiple deadenylases can act on the same mRNA, with discrete but overlapping functions. To understand the biological significance of the diversity of these enzymes we silenced the expression of several deadenylases, including PARN, NOC, CNOT6, CNOT7 and CNOT8, in human cells of cancer origin (Hep2; epithelial carcinoma cells), and analyze the impact on gene expression with microarrays.

2015-10-01 | GSE67536 | GEO

RNAi profiling by array of human MCF7 cells with siRNA-mediated knockdown of CNOT6 and/or CNOT6L, or CNOT7 and CNOT8

Project description:siRNA-mediated knockdown of CNOT6 and/or CNOT6L, or CNOT7 and CNOT8 compared to control to study role in the regulation of gene expression by these factors.

2011-09-14 | E-MEXP-2926 | biostudies-arrayexpress

CNOT6/6L-mediated mRNA Degradation in Ovarian Granulosa Cells is a Key Mechanism of Gonadotropin-triggered Follicle Development

Project description:CCR4-NOT deadenylase is a major regulator of mRNA turnover in eukaryotes. It contains 2 heterogeneous catalytic subunits encoded by Cnot7/8 and Cnot6/6l genes in vertebrates, respectively. The physiological function of each catalytic subunit was unclear due to potential gene redundancy in vivo. In this study Cnot6/6l double knockout mice were generated. These mice were viable and generally healthy, but females were infertile with irregular estrus cycle and poor ovarian responses to gonadotropins. Follicle-stimulating hormone (FSH) stimulated transcription and translation of mRNAs encoding CNOT6 and CNOT6L in ovarian granulosa cells through the activity of phosphoinositide 3-kinase signaling pathway. Results of transcriptome analyses indicated that FSH downregulated a large number of transcripts in granulosa cells during the transition from pre-antral to antral follicles in a CNOT6/6L-dependent manner. These two deadenylases functioned as key effectors of FSH in altering the developmental programs in granulosa cells and triggered clearance of specific transcripts in growing follicles, particularly those encoding repressing factors of granulosa cell proliferation and steroidogenesis. These results demonstrated that FSH modulates granulosa cells function by stimulating selective translational activation and degradation of existed mRNAs, in additional to inducing de novo gene transcription. Meanwhile, this study provides in vivo evidence that CCR4-NOTCNOT6/6L-mediated mRNA deadenylation is dispensable in most somatic cell types, but is essential for the female reproductive endocrine regulation.

2021-11-04 | GSE158773 | GEO

The PARN deadenylase targets a discrete set of mRNAs for decay and regulates cell motility in mouse myoblasts

Project description:Almost all cellular mRNAs terminate in a 3M-bM-^@M-^Y poly(A) tail, the removal of which can induce both translational silencing and mRNA decay. Mammalian cells encode many poly(A)-specific exoribonucleases but their individual roles are poorly understood. Here, we undertook an analysis of the role of PARN deadenylase in mouse myoblasts using global measurements of mRNA decay rates. Our results reveal that a discrete set of mRNAs exhibit altered mRNA decay as a result of PARN depletion and that stabilization is associated with increased poly(A) tail length and translation. We determined that stabilization of mRNAs does not generally result in their increased abundance supporting the idea that mRNA decay is coupled to transcription. Importantly, PARN knockdown has wide ranging effects on gene expression that specifically impact the extracellular matrix and cell migration. Finally, although PARN has its own unique target transcripts it also influences some genes whose expression is modulated by other deadenylases. In order to investigate the role of PARN deadenyl on mRNA decay in mouse muscle cells , C2C12 cells were transfected by shRNA knockdown of PARN and treated with actinomycin D to inhibit transcription. Total RNA was isolated from three replicates at 0, 15, 30, 60, 120 and 240 minutes. The CTRL cell line stably transfected with LKO1 vector was described previously (GSE21233).

2012-08-31 | E-GEOD-35944 | biostudies-arrayexpress

The PARN deadenylase targets a discrete set of mRNAs for decay and regulates cell motility in mouse myoblasts

Project description:Almost all cellular mRNAs terminate in a 3’ poly(A) tail, the removal of which can induce both translational silencing and mRNA decay. Mammalian cells encode many poly(A)-specific exoribonucleases but their individual roles are poorly understood. Here, we undertook an analysis of the role of PARN deadenylase in mouse myoblasts using global measurements of mRNA decay rates. Our results reveal that a discrete set of mRNAs exhibit altered mRNA decay as a result of PARN depletion and that stabilization is associated with increased poly(A) tail length and translation. We determined that stabilization of mRNAs does not generally result in their increased abundance supporting the idea that mRNA decay is coupled to transcription. Importantly, PARN knockdown has wide ranging effects on gene expression that specifically impact the extracellular matrix and cell migration. Finally, although PARN has its own unique target transcripts it also influences some genes whose expression is modulated by other deadenylases.

2012-08-31 | GSE35944 | GEO

The CNOT4 Subunit of the CCR4-NOT Complex is Involved in mRNA Degradation, Efficient DNA Damage Repair, and XY Chromosome Crossover during Male Germ Cell Meiosis

Project description:The CCR4-NOT complex is a major mRNA deadenylase in eukaryotes and comprise CNOT6/6L and CNOT7/8, both of which are two types of catalytic subunits, as well as CNOT4, which is a regulatory subunit with previously undetermined functions. These subunits are previously hypothesized to play synergistic biochemical functions; however, their in vivo functions have remained poorly investigated. In this study, we constructed viable Cnot6/6l double knockout mice and the males were fertile, whereas Cnot7–/– male mice have been reported to be infertile. These results are against canonical viewpoints and indicate that CNOT7 has CNOT6/6L-independent functions in vivo. We also demonstrate that the mouse testis has an endogenous complex that contains CNOT7 and CNOT4, but not CNOT6/6L. CNOT4 is required for post-implantation embryo development and meiosis progression during spermatogenesis. Selective knockout of Cnot4 in male germ cells leads to defective DNA damage repair and homologous crossover formation between X and Y chromosomes in a cell autonomous manner. CNOT4 functions as a previously unrecognized mRNA adaptor of CCR4-NOT by targeting mRNAs to CNOT7 for deadenylation of the poly(A) tails, thereby mediating the degradation of a subset of transcripts from the zygotene to pachytene stage. Therefore, the mRNA removal promoted by CNOT4-regulated CCR4-NOT complex during the zygotene to pachytene transition is crucial for the selection of DNA repair pathways, subsequent activation of meiosis-related genes, sex chromosome paring, and ultimately, male fertility.

2021-02-19 | GSE158471 | GEO

Expression data from control or CCR4b/CNOT6L-depleted cells

2011-08-03 | GSE31148 | GEO

PARN and TOE1 constitute a 3′ end maturation module for nuclear non-coding RNAs

Project description:Poly(A)-specific ribonuclease (PARN) and target of EGR1 protein 1 (TOE1) are nuclear granule-associated deadenylases, whose mutations are linked to multiple human diseases. Here, we applied mTAIL-seq and RNA sequencing (RNA-seq) to systematically identify the substrates of PARN and TOE1 and elucidate their molecular functions. We found that PARN and TOE1 do not modulate the length of mRNA poly(A) tails. Rather, they promote the maturation of nuclear small non-coding RNAs (ncRNAs). PARN and TOE1 act redundantly on some ncRNAs, most prominently small Cajal body-specific RNAs (scaRNAs). scaRNAs are strongly downregulated when PARN and TOE1 are compromised together, leading to defects in small nuclear RNA (snRNA) pseudouridylation. They also function redundantly in the biogenesis of telomerase RNA component (TERC), which shares sequence motifs found in H/ACA box scaRNAs. Our findings extend the knowledge of nuclear ncRNA biogenesis, and they provide insights into the pathology of PARN/TOE1-associated genetic disorders whose therapeutic treatments are currently unavailable.

2018-03-30 | GSE111511 | GEO

Expression data from control or CCR4b/CNOT6L-depleted cells

Project description:The stability of mRNA influences the abundance of cellular transcripts and proteins. Deadenylases play critical roles in mRNA turnover and thus are important for the regulation of arious biological events. Here, we report the identification and characterization of CCR4b/CNOT6L, which is homologous to yeast CCR4 mRNA deadenylase. We compared the expression profiles of the cellular genes between control and CCR4b/CNOT6L-depleted cells by microarray analysis Total RNAs were extracted from control or CCR4b-depleted cells. Following the isolation of RNA, all subsequent technical procedures, including quality control of RNA, labeling, hybridization, and scanning of the arrays, were performed in BIO MATRIX Research, Inc., cDNA, and biotinlabeled cRNA was synthesized according to protocols for Affymetrix array analysis. Biotin-labeled cRNAs (15 g) were hybridized to the GeneChip Mouse Genome 430 2.0 Array (Affymetrix, Santa Clara, CA). To determine the average differences for each probe set, changes were calculated by a global normalization method using Affymetrix GCOS (GeneChipOperating Software) software.

2011-08-02 | E-GEOD-31148 | biostudies-arrayexpress

Transcription profiling of human MCF7 cell line with siRNA mediated knockdown of CNOT7 and/or CNOT8 subunits of Ccr4-Not deadenylase compared to control

Project description:siRNA-mediated knockdown of CNOT7 and/or CNOT8 compared to control to study role in the regulation of gene expression by these factors.

2009-07-15 | E-MEXP-2218 | biostudies-arrayexpress

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