CRISPRi proliferation and differentiation screening to identify functional long noncoding RNAs in pediatric AML

Long non-coding RNAs (lncRNAs) have been shown to be stage- and cell type-specific regulators of gene expression. We previously discovered stem cell- and AML subtype-specific lncRNA expression signatures. Using a CRISPRi approach we screened 619 genes in 7 AML cell lines, selecting for proliferation, as well as differentiation phenotype. ShRNA Knockdown of the lncRNA here termed LNCER transferred proliferation disadvantage in the erythroleukemic CMK cell line. LNCER is not expressed in human hematopoietic stem cells (HSC), yet highly expressed in patient samples of transient myeloproliferative disease and myeloid leukemia of Down syndrome. We performed RNA-sequencing upon shRNA knockdown in CMK, revealing down-regulation of megakaryocytic and up-regulation of erythroid signature genes. Next, we induced LNCER knockdown in HSC in vitro. Methocult and in vitro differentiation assays showed increased erythroid differentiation.

Our results implicate significance of lncRNA in malignant hematopoesis. CRISPRi screenings revealed candidate regulators of differentiation and proliferation in pediatric AML. LNCER stood out as a potential regulator of erythroleukemia and erythroid cell fate.

Publication History

Article published online:
17 May 2022