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Purification of Ubiquitinated p53 Proteins from Mammalian Cells
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Context-Dependent Function of Long Noncoding RNA PURPL in Transcriptome Regulation during p53 Activation.

Corrine Corrina R Hartford1, Roshan L Shrestha2, Lorinc Pongor3

  • 1Regulatory RNAs and Cancer Section, Genetics Branch, Center for Cancer Research (CCR), National Cancer Institutegrid.48336.3a (NCI), National Institutes of Healthgrid.94365.3d (NIH), Bethesda, Maryland, USA.

Molecular and Cellular Biology
|November 7, 2022
PubMed
Summary

The long non-coding RNA PURPL fine-tunes mitotic gene expression upon p53 activation in liver cancer. Depleting PURPL significantly alters gene expression and reduces mitotic cells, revealing its context-dependent role in cell cycle arrest.

Keywords:
CRISPR/Cas9CRISPRiIso-SeqLINC01021PURPLPacBioalternative splicingcell cycle checkpointscontext-dependentintron retentionliver cancerlncRNAmitosisp53

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Area of Science:

  • Molecular Biology
  • Cancer Research
  • Genomics

Background:

  • The p53-induced long non-coding RNA (lncRNA) PURPL is known to suppress basal p53 levels.
  • PURPL exhibits significantly higher expression in liver cancer cells compared to other cell types.

Purpose of the Study:

  • To investigate the function of PURPL upon p53 activation in liver cancer.
  • To identify novel PURPL transcripts and elucidate its role in regulating gene expression and cell cycle.

Main Methods:

  • Isoform sequencing to discover novel PURPL transcripts.
  • CRISPR interference (CRISPRi) to deplete PURPL.
  • RNA-sequencing (RNA-Seq) and Nutlin treatment to induce p53.
  • Pathway analysis and cell cycle analysis.

Main Results:

  • Discovery of novel PURPL transcripts with retained introns and/or unannotated exons.
  • Loss of PURPL altered only 7 genes in untreated cells but ~800 genes upon p53 activation, indicating context-dependent function.
  • PURPL depletion significantly decreased the percentage of mitotic cells and pathway analysis implicated PURPL in fine-tuning mitotic gene expression.

Conclusions:

  • Novel PURPL transcripts have been identified.
  • PURPL plays a critical role in moderating the expression of mitotic genes during p53 activation.
  • PURPL is essential for controlling cell cycle arrest in response to p53 signaling in liver cancer.