An MXD1-derived repressor peptide identifies noncoding mediators of MYC-driven cell proliferation

Proceedings of the National Academy of Sciences of the United States of America
Philipp RaffeinerPeter K Vogt

Abstract

MYC controls the transcription of large numbers of long noncoding RNAs (lncRNAs). Since MYC is a ubiquitous oncoprotein, some of these lncRNAs probably play a significant role in cancer. We applied CRISPR interference (CRISPRi) to the identification of MYC-regulated lncRNAs that are required for MYC-driven cell proliferation in the P493-6 and RAMOS human lymphoid cell lines. We identified 320 noncoding loci that play positive roles in cell growth. Transcriptional repression of any one of these lncRNAs reduces the proliferative capacity of the cells. Selected hits were validated by RT-qPCR and in CRISPRi competition assays with individual GFP-expressing sgRNA constructs. We also showed binding of MYC to the promoter of two candidate genes by chromatin immunoprecipitation. In the course of our studies, we discovered that the repressor domain SID (SIN3-interacting domain) derived from the MXD1 protein is highly effective in P493-6 and RAMOS cells in terms of the number of guides depleted in library screening and the extent of the induced transcriptional repression. In the cell lines used, SID is superior to the KRAB repressor domain, which serves routinely as a transcriptional repressor domain in CRISPRi. The SID transcriptional r...Continue Reading

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Citations

Mar 12, 2020·Proceedings of the National Academy of Sciences of the United States of America·Philipp RaffeinerPeter K Vogt
Apr 5, 2020·Proceedings of the National Academy of Sciences of the United States of America·James D Phelan, Louis M Staudt
Sep 8, 2020·Expert Opinion on Therapeutic Targets·Vivian Weiwen XueWilliam Chi Shing Cho
Jan 10, 2021·Nature Cell Biology·Muneaki NakamuraLei S Qi
Mar 10, 2021·RNA Biology·Meira S ZibittAshish Lal
Nov 8, 2020·Seminars in Cancer Biology·Rongfu TuGuoliang Qing
Jul 3, 2021·International Journal of Molecular Sciences·Katarzyna Horodecka, Markus Düchler

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Methods Mentioned

BETA
co-IP
immunoprecipitation
flow cytometry
ChIP
ChIP-seq
RNA-seq
PCR
transfection
electrophoresis

Software Mentioned

edgeR
R
count
RSEM
ggplot2
GNU
CASSAVA
RAMOS

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