GSE224548
GSE GEOA two-step in vivo CRISPR screen unveils pervasive RNA binding protein dependencies for leukemic stem cells and identifies ELAVL1 as a therapeutic target
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Summary
Acute myeloid leukemia (AML) progression and relapse is fueled by self-renewing leukemic stem cells (LSCs) whose molecular determinants have been difficult to discern from normal hematopoietic stem cells (HSCs) or to uncover in screening approaches focused on general AML cell properties. We have identified a unique set of RNA binding proteins (RBPs) that are enriched in human AML LSCs but repressed in HSCs. Using an in vivo two step CRISPR-Cas9-mediated screening approach to specifically score for cancer stem cell functionality, we found 32 RBPs essential for LSC propagation and self- renewal in MLL-AF9 translocated AML. Using knockdown or small molecule approaches we show that targeting key hit RBP ELAVL1 impaired LSC-driven in vivo leukemic reconstitution and selectively depleted primitive AML cells vs. normal hematopoietic stem and progenitors. Importantly, knockdown of Elavl1 spared HSCs while significantly reducing LSC numbers across genetically diverse leukemias. Integrative RNA-seq and eCLIP-seq profiling revealed hematopoietic differentiation, RNA splicing and mitochondrial metabolism as key features defining the leukemic ELAVL1-mRNA interactome with the mitochondrial import protein TOMM34 being a direct ELAVL1-stabilized target whose inhibition impairs AML propagation.
Overall Design
Human primary AML cells (AML #4) were infected in triplicate with shScramble or shELAVL1 lentivirus (pLKO.1) at an MOI of 50. At 48h, 100k 7AAD-EGFP+ transduced cells were FACS-sorted per sample and RNA was extracted using TRIzolLS.
Analysis (3 steps)
View Data Processing- Reads were quality checked using fastQC
- Sequencing reads were aligned to the hg38 reference genome using STAR (v2.7.2c)
- Gene-level quantification was performed using RSEM (v1.3.1)
Supplementary Files (1)
GEO Samples (6)
Dataset Citations (1)
SRA Experiments (6) and Runs (12)
Total: 19497 MBSample attributes
Original files (1)
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Linked Publications (2)
Data Files (12)
| Accession | File Name | Stored Type | Output Type | Mapping Assembly | Size | Download | |
|---|---|---|---|---|---|---|---|
| — | 1-090191-shSCR-Rep1_S1_L001_R1_001.fastq.gz | RNA-Seq | 2.3 GB | link | |||
| — | 1-090191-shSCR-Rep1_S1_L002_R1_001.fastq.gz | RNA-Seq | 2.3 GB | link | |||
| — | 2-090191-shSCR-Rep2_S2_L001_R1_001.fastq.gz | RNA-Seq | 1.6 GB | link | |||
| — | 2-090191-shSCR-Rep2_S2_L002_R1_001.fastq.gz | RNA-Seq | 1.6 GB | link | |||
| — | 3-090191-shSCR-Rep3_S3_L001_R1_001.fastq.gz | RNA-Seq | 1.5 GB | link | |||
| — | 3-090191-shSCR-Rep3_S3_L002_R1_001.fastq.gz | RNA-Seq | 1.5 GB | link | |||
| — | 4-090191-shELAVL1-7-Rep1_S4_L001_R1_001.fastq.gz | RNA-Seq | 1.5 GB | link | |||
| — | 4-090191-shELAVL1-7-Rep1_S4_L002_R1_001.fastq.gz | RNA-Seq | 1.5 GB | link | |||
| — | 5-090191-shELAVL1-7-Rep2_S5_L001_R1_001.fastq.gz | RNA-Seq | 1.2 GB | link | |||
| — | 5-090191-shELAVL1-7-Rep2_S5_L002_R1_001.fastq.gz | RNA-Seq | 1.2 GB | link | |||
| — | 6-090191-shELAVL1-7-Rep3_S6_L001_R1_001.fastq.gz | RNA-Seq | 1.4 GB | link | |||
| — | 6-090191-shELAVL1-7-Rep3_S6_L002_R1_001.fastq.gz | RNA-Seq | 1.4 GB | link |