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GSE195493

GSE GEO
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Cytoplasmic and nuclear fractionation from HEK293T cells

Organism: Homo sapiens
Platform: GPL24676
Samples: 4
Experiment Types:
Other
Submitted: Jan 26 2022
Last Updated: May 11 2022
Status: Public on Mar 30 2022
Contact: Amy,,Pasquinelli (University of California, San Diego)

Relations

BioProject: https://www.ncbi.nlm.nih.gov/bioproject/PRJNA800803

Summary

The poly(A) tail appended to the 3’ end of many eukaryotic transcripts plays a key role in the stability, nuclear transport, and translation of the RNA. These roles are largely mediated by the Poly(A) Binding Proteins (PABPs) that coat the poly(A) tails and interact with various proteins involved in the biogenesis and function of the RNA. While it is well established that the nuclear PABP (PABPN) binds newly synthesized poly(A) tails and is apparently replaced by the cytoplasmic PABP (PABPC) on transcripts exported to the cytoplasm, the distribution of transcripts for different genes or isoforms of the same gene on these distinct PABPs has not been investigated on a genome-wide scale. Here, we analyzed the identity, splicing status, poly(A) tail size, and translation status of RNAs co-immunoprecipitated with endogenous PABPN or PABPC in human cells. At steady state, many protein coding and non-coding RNAs exhibit strong bias for association with PABPN or PABPC. While PABPN-enriched transcripts more often were incompletely spliced and harbored longer poly(A) tails and PABPC-enriched RNAs tended to have longer half-lives and higher translation efficiency, there are curious outliers. Overall, our study reveals the landscape of RNAs bound by PABPN and PABPC, providing new details that support and advance the current understanding of the roles these proteins play in poly(A) tail synthesis, maintenance and function.

Overall Design

2 biological replicates were crosslinked and collected; subcellular fractionation was performed to separate the cytoplasm from the nuclear fraction

Analysis (5 steps)

View Data Processing
Processing steps for GSE195493
  1. All reads were aligned to the human genome hg38 primary assembly using STAR.
  2. featureCounts version 2.0.2 was used to annotate reads, using the flag --minOverlap 20 and a custom GTF file derived from gencode v34
  3. Differential expression was calculated using DESeq2.
  4. Cut-offs used to determine significant differential expression were baseMean > 50 and padj <=0.01.
  5. In this comparison, a negative log2FoldChange indicates that a transcript was more nuclear, while a positive log2FoldChange indicates that a transcript was more cytoplasmic.

Supplementary Files (1)

GSE195493_CYTOvNUCLEAR_fractionation.csv.gz Download
GEO Samples (4)

Dataset Citations (1)

Nuclear and cytoplasmic poly(A) binding proteins (PABPs) favor distinct transcripts and isoforms.
PMID 35438785 · 2022 · Nucleic acids research
Angela L Nicholson-Shaw, Eric R Kofman, Gene W Yeo, Amy E Pasquinelli

SRA Experiments (4) and Runs (4)

Total: 7107 MB
SRX13933608 SRP356888 RIP-Seq PAIRED
GSM5839349: HEK293T- Nuclear fraction RNA Rep1; Homo sapiens; RIP-Seq
Sample: SRS11777513
BioProject: PRJNA800803
BioSample: SAMN25284384
Platform: ILLUMINA
Instrument: Illumina NovaSeq 6000
Organism: Homo sapiens
Sample attributes
source_name: HEK293T Total cell
cell line: HEK293T
genotype: wildtype
treatment: formaldehyde crosslinked
fraction: Nuclear
Original files (1)
HEK293T Total cell
Runs (1)
Run Spots Bases Size (MB) Files Link
SRR17771273 27857815 5627278630 1700.32 Nuclear_frac_Rep1_R1.fastq.gz, Nuclear_frac_Rep1_R2.fastq.gz, SRR1777… SRA
SRX13933609 SRP356888 RIP-Seq PAIRED
GSM5839350: HEK293T- Nuclear fraction RNA Rep2; Homo sapiens; RIP-Seq
Sample: SRS11777514
BioProject: PRJNA800803
BioSample: SAMN25284383
Platform: ILLUMINA
Instrument: Illumina NovaSeq 6000
Organism: Homo sapiens
Sample attributes
source_name: HEK293T Total cell
cell line: HEK293T
genotype: wildtype
treatment: formaldehyde crosslinked
fraction: Nuclear
Original files (1)
HEK293T Total cell
Runs (1)
Run Spots Bases Size (MB) Files Link
SRR17771272 30911634 6244150068 1893.04 Nuclear_frac_Rep2_R1.fastq.gz, Nuclear_frac_Rep2_R2.fastq.gz, SRR1777… SRA
SRX13933610 SRP356888 RIP-Seq PAIRED
GSM5839351: HEK293T- Cytoplasm fraction RNA Rep1; Homo sapiens; RIP-Seq
Sample: SRS11777515
BioProject: PRJNA800803
BioSample: SAMN25284382
Platform: ILLUMINA
Instrument: Illumina NovaSeq 6000
Organism: Homo sapiens
Sample attributes
source_name: HEK293T Total cell
cell line: HEK293T
genotype: wildtype
treatment: formaldehyde crosslinked
fraction: Cytoplasmic
Original files (1)
HEK293T Total cell
Runs (1)
Run Spots Bases Size (MB) Files Link
SRR17771271 29793114 6018209028 1787.78 Cyto_frac_Rep1_R1.fastq.gz, Cyto_frac_Rep1_R2.fastq.gz, SRR17771271, … SRA
SRX13933611 SRP356888 RIP-Seq PAIRED
GSM5839352: HEK293T- Cytoplasm fraction RNA Rep2; Homo sapiens; RIP-Seq
Sample: SRS11777516
BioProject: PRJNA800803
BioSample: SAMN25284381
Platform: ILLUMINA
Instrument: Illumina NovaSeq 6000
Organism: Homo sapiens
Sample attributes
source_name: HEK293T Total cell
cell line: HEK293T
genotype: wildtype
treatment: formaldehyde crosslinked
fraction: Cytoplasmic
Original files (1)
HEK293T Total cell
Runs (1)
Run Spots Bases Size (MB) Files Link
SRR17771270 28775151 5812580502 1726.35 Cyto_frac_Rep2_R1.fastq.gz, Cyto_frac_Rep2_R2.fastq.gz, SRR17771270, … SRA

Linked Publications (2)

Data Files (6)

Accession File Name Stored Type Output Type Mapping Assembly Size Download
Cyto_frac_Rep1_R1.fastq.gz RIP-Seq 1.7 GB link
Cyto_frac_Rep2_R1.fastq.gz RIP-Seq 1.7 GB link
Nuclear_frac_Rep1_R1.fastq.gz RIP-Seq 1.7 GB link
Nuclear_frac_Rep2_R1.fastq.gz RIP-Seq 1.8 GB link
SRR17771272.lite RIP-Seq 1.8 GB link
SRR17771273.lite RIP-Seq 1.7 GB link