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Global analysis of differential polyadenylation of mRNAs in CPEB1 knockdown and CPEB4 knockdown at G2/M.

ABSTRACT: We compared the poly(A) tail length status of mRNAs of HeLa cells expressing a CPEB1 shRNA (CPEB1 knockdown) versus a control shRNA, and expressing a CPEB4 shRNA (CPEB4 knockdown) versus a control shRNA. Results provide insight into the extent of gene regulation mediated by CPEB1 and CPEB4 activity during mitotic cell cycle progression. The different shRNA expressing cells were synchronized with double thymidine blockade (12 hours with 2 mM thymidine, 12 hours release, and 12 hours with 2 mM thymidine), and samples were taken after 8 hours release (G2/M phase). For each shRNA expressing HeLa cell line total RNA was purified by two different procedures: poly(U) chromatography and oligo(dT)-chromatography. Poly(U)-chromatography (Jacobson, 1987): 100 μg of total RNA were bound to poly(U)-sepharose (Sigma) and eluted at 35ºC to isolate mRNAs with short poly(A) tail (<30As, SHORT fraction). Oligo(dT) chromatography: mRNAs were purified independently of their poly(A) tail length with Ambion Poly(A)Purist kit from 20 μg total RNA (ALL fraction). Jacobson, A. Purification and fractionation of poly(A)+ RNA. Methods in Enzymology (1987) 152: 254-261. Keywords: knock-down experiment

ORGANISM(S): Homo sapiens

PROVIDER: GSE15549 | GEO | 2010/04/01

SECONDARY ACCESSION(S): PRJNA115989

REPOSITORIES: GEO

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Global analysis of differential polyadenylation of mRNAs during mitotic cell cycle: S phase versus G2/M phase.

Project description:We compared the poly(A) tail length status of mRNAs of HeLa cells between two phases of the mitotic cell cycle: S and G2/M phases. Hundreds of mRNAs were found to be regulated by changes in their poly(A) tail length during mitotic cell cycle in a phase specific manner. Many of these differentially polyadenylated mRNAs encode proteins related to cell death, cell cycle and cellular growth and proliferation. HeLa cells were synchronized with double thymidine blockade (12 hours with 2 mM thymidine, 12 hours release, and 12 hours with 2 mM thymidine), and samples were taken after 2 hours release (S phase) and 8 hours release (G2/M phase). For each condition total RNA was purified by two different procedures: poly(U) chromatography and oligo(dT)-chromatography. Poly(U)-chromatography (Jacobson, 1987): 100 μg of total RNA were bound to poly(U)-sepharose (Sigma) and eluted at 35ºC to isolate mRNAs with short poly(A) tail (<30As, SHORT fraction). Oligo(dT) chromatography: mRNAs were purified independently of their poly(A) tail length with Ambion Poly(A)Purist kit from 20 μg total RNA (ALL fraction). Jacobson, A. Purification and fractionation of poly(A)+ RNA. Methods in Enzymology (1987) 152: 254-261. Keywords: time course

2010-04-01 | GSE15399 | GEO

Global analysis of differential polyadenylation of mRNAs during mitotic cell cycle: S phase versus G2/M phase.

Project description:We compared the poly(A) tail length status of mRNAs of HeLa cells between two phases of the mitotic cell cycle: S and G2/M phases. Hundreds of mRNAs were found to be regulated by changes in their poly(A) tail length during mitotic cell cycle in a phase specific manner. Many of these differentially polyadenylated mRNAs encode proteins related to cell death, cell cycle and cellular growth and proliferation. HeLa cells were synchronized with double thymidine blockade (12 hours with 2 mM thymidine, 12 hours release, and 12 hours with 2 mM thymidine), and samples were taken after 2 hours release (S phase) and 8 hours release (G2/M phase). For each condition total RNA was purified by two different procedures: poly(U) chromatography and oligo(dT)-chromatography. Poly(U)-chromatography (Jacobson, 1987): 100 Î¼g of total RNA were bound to poly(U)-sepharose (Sigma) and eluted at 35ÂºC to isolate mRNAs with short poly(A) tail (<30As, SHORT fraction). Oligo(dT) chromatography: mRNAs were purified independently of their poly(A) tail length with Ambion Poly(A)Purist kit from 20 Î¼g total RNA (ALL fraction). Jacobson, A. Purification and fractionation of poly(A)+ RNA. Methods in Enzymology (1987) 152: 254-261. Keywords: time course Comparison of ALL fraction mRNAs and SHORT fraction mRNAs measured after 2 hours (S phase) and 8 hours release (G2/M) from double thymidine blockade; 3 biological replicates at each of the two time points; two technical replicates with dye swapping per comparison.

2010-05-07 | E-GEOD-15399 | biostudies-arrayexpress

Genome-wide analysis of mRNA polyadenylation in Cortex/Striatum of CPEB4 modified mice [I]

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Project description:To identify CPEB1 and CPEB4 regulated RNA we performed CPEB1 and CPEB4 RNA immunoprecipitation (RIP) followed by microarray hybridization analysis with striatal (St) RNA from wild-type (WT) and R6/1 mice (HD mice).

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Genome-wide analysis of mRNA polyadenylation in Cortex/Striatum of CPEB4 modified mice [II]

Project description:To test the mRNA polyadenylation status in brain of CPEB4 modified mice we performed poly(U)-chromatography on RNA purified from a pool of cortex and striatum tissues of control and TgCPEB4∆4 mice

2020-03-06 | GSE113840 | GEO

Oppositional Poly(A) Tail Length Regulation by FMRP and CPEB1

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2022-03-01 | GSE188840 | GEO

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2021-11-04 | PXD020822 | Pride

Oppositional Poly(A) Tail Length Regulation by FMRP and CPEB1

Project description:Oppositional Poly(A) Tail Length Regulation by FMRP and CPEB1

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Spindle-localized mRNA translation mediated by CPEB1 and CPEB4 controls mitotic progression

Project description:Sequencial functions of CPEB1 and CPEB4 in the localization of repressed mRNAs to the mitotic spindle and their subsequent phase-specific activation to promote cell phase transitions and correct chromosome segregation

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Project description:Here we show sequential functions of CPEB1 and CPEB4 in the localization of repressed mRNAs to the mitotic spindle and their subsequent phase -specific activation to promote cell phase transitions and correct chromosome segregation.

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