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| Status |
Public on Apr 20, 2023 |
| Title |
RNA binding protein PRRC2B mediates translation of specific proteins and regulates cell cycle progression [Polysome-seq] |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Accumulating evidence suggests that posttranscriptional regulation of gene expression, including regulation of RNA splicing, transport, modification, translation, and degradation, primarily relies on RNA binding proteins (RBPs). However, the functions of many RBPs remain understudied. Here, we characterized the function of a novel RBP, Proline-Rich Coiled-coil 2B (PRRC2B). Through photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) and deep sequencing, we identified transcriptome-wide CU- or GA-rich PRRC2B binding sites around the translation initiation codon on a specific cohort of mRNAs in HEK293T cells. These PRRC2B target mRNAs, including oncogenes and cell cycle regulators such as CCND2 (cyclin D2), exhibited decreased translation upon PRRC2B knockdown as revealed by sequencing polysome-associated RNA, resulting in decreased G1/S phase transition and cell proliferation. Antisense oligonucleotides (ASOs) blocking PRRC2B-CCND2 mRNA interaction decreased CCND2 translation, thus inhibited G1/S transition and cell proliferation. Mechanistically, PRRC2B interactome capture analysis revealed RNA-independent interactions with eukaryotic initiation factors eIF4G2, eIF3, and FXR1. The interaction with eIF4G2 is essential for PRRC2B function since unlike wildtype PRRC2B, eIF4G2-interacting defective mutants failed to rescue the translation deficiency caused by PRRC2B knockdown. Taken together, our findings reveal that PRRC2B, by interacting with eIF4G2, is essential for efficient translation of specific proteins required for cell cycle progression and cell proliferation.
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| Overall design |
We performed polysome-seq on HEK293T cells and its derivatives (shPRRC2B). Total RNA and polysome-associated RNA extracted from HEK293T cells and its knockdown derivatives (shPRRC2B) were subjected to RNA-seq. Biological triplicates were performed. Samples were subjected to DNA removal by DNase I and polyA enrichment before library construction by NGS Library Prep following the manufacturer's protocols. Paired-end sequencing was conducted at Novogene using NovaSeq6000 S4 with a depth of 20M reads/sample.
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| Web link |
https://pubmed.ncbi.nlm.nih.gov/37125639/
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| Contributor(s) |
Jiang F, Hedaya OM, Khor E, Wu J, Auguste M |
| Citation(s) |
37125639 |
| |
| Submission date |
Dec 05, 2022 |
| Last update date |
Jul 20, 2023 |
| Contact name |
feng jiang |
| E-mail(s) |
[email protected]
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| Phone |
5855002752
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| Organization name |
University of Rochester
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| Department |
Biochemistry and Biophysics
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| Lab |
Peng Yao lab
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| Street address |
601 Elmwood Ave box CVRI
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| City |
Rochester |
| State/province |
New York |
| ZIP/Postal code |
14620 |
| Country |
USA |
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| Platforms (1) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
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| Samples (12)
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| This SubSeries is part of SuperSeries: |
| GSE220059 |
RNA binding protein PRRC2B mediates translation of specific proteins and regulates cell cycle progression |
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| Relations |
| BioProject |
PRJNA908733 |
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