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Series GSE275118 Query DataSets for GSE275118
Status Public on Nov 20, 2024
Title An E2 ubiquitin conjugating enzyme links diubiquitinated H2B to H3K27M oncohistone function
Organism Caenorhabditis elegans
Experiment type Genome binding/occupancy profiling by high throughput sequencing
Summary The H3K27M oncogenic histone (oncohistone) mutation drives ~80% of incurable childhood brain tumors known as diffuse midline gliomas (DMGs). The major molecular feature of H3K27M mutant DMGs is a global loss of H3K27 tri-methylation (H3K27me3), a phenotype conserved in Caenorhabditis elegans (C. elegans). Here we perform unbiased genome-wide suppressor screens in C. elegans expressing H3K27M, and isolate 20 suppressors, all of which at least partially restore H3K27me3. 19/20 suppressor mutations map to the same histone H3.3 gene in which the K27M mutation was originally introduced. Most of these create single amino acid substitutions between residues R26-Y54, which do not disrupt oncohistone expression. Rather, they are predicted to impair interactions with the Polycomb Repressive Complex 2 (PRC2) and are functionally conserved in human cells. Further, we mapped a single extragenic H3K27M suppressor to ubc-20, an E2 ubiquitin conjugating enzyme, whose loss rescued H3K27me3 to nearly 50% wild-type levels despite continued oncohistone expression and chromatin incorporation. We demonstrate that ubc-20 is the major enzyme responsible for generating di-ubiquitinated histone H2B. Our study provides in vivo support for existing models of PRC2 inhibition via direct oncohistone contact, and suggests that the effects of H3K27M may be modulated by H2B ubiquitination.
 
Overall design To investigate how ubc-20 mutants suppress K27M phenotypes, we performed CUT&RUN for H3K27me3 and H3K27M in wild-type (N2), K27M, and K27M; ubc-20(G155R) mutants.
Wild-type (N2), K27M, and K27M;ubc-20(G155R) C. elegans populations were grown to starvation on NA22-seeded agar plates at 20C, and L2 larvae starved for approximately 24h were collected for CUT&RUN.
Web link https://pubmed.ncbi.nlm.nih.gov/39560642/
 
Contributor(s) Jiao A, Shi Y
Citation(s) 39560642
Submission date Aug 18, 2024
Last update date Nov 21, 2024
Contact name Alan Jiao
E-mail(s) alan.jiao@ludwig.ox.ac.uk
Organization name Ludwig Institute for Cancer Research, Oxford University
Street address Old Road Campus Research Building
City Oxford
State/province MA
ZIP/Postal code OX37DQ
Country United Kingdom
 
Platforms (1)
GPL32326 NextSeq 2000 (Caenorhabditis elegans)
Samples (18)
GSM8465855 CUTandRUN_H3K27me3_in_K27M_L2_Rep1
GSM8465856 CUTandRUN_H3K27me3_in_K27M_L2_Rep2
GSM8465857 CUTandRUN_H3K27me3_in_K27M_L2_Rep3
Relations
BioProject PRJNA1149477

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Supplementary file Size Download File type/resource
GSE275118_RAW.tar 482.0 Mb (http)(custom) TAR (of BW)
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Raw data are available in SRA

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