The lncRNA LAMP5-AS1 drives leukemia cell stemness by directly modulating DOT1L methyltransferase activity in MLL leukemia

Abstract Background Mixed-lineage leukemia ( MLL ) gene rearrangements trigger aberrant epigenetic modification and gene expression in hematopoietic stem and progenitor cells, which generates one of the most aggressive subtypes of leukemia with an apex self-renewal. It remains a challenge to directly inhibit rearranged MLL itself because of its multiple fusion partners and the poorly annotated downstream genes of MLL fusion proteins; therefore, novel therapeutic targets are urgently needed. Methods qRT-PCR, receiver operating characteristic (ROC), and leukemia-free survival analysis were used to validate LAMP5-AS1 ( LAMP5 antisense 1) expression and evaluate its clinical value. We performed in vitro and in vivo experiments to investigate the functional relevance of LAMP5-AS1 in MLL leukemia progression and leukemia cell stemness. RNA electrophoretic mobility shift assays (EMSA), histone methyltransferase assay, RNA pull-down assay, and RNA fluorescence in situ hybridization (FISH) were used to validate the relationship between LAMP5-AS1 and the methyltransferase activity of DOT1L. The downstream ectopic target genes of LAMP5-AS1/DOT1L were validated by the chromatin immunoprecipitation (ChIP) and western blot. Results We discovered that a long noncoding RNA (lncRNA) LAMP5-AS1 can promote higher degrees of H3K79 methylation, followed by upregulated expression of the self-renewal genes in the HOXA cluster, which are responsible for leukemia stemness in context of MLL rearrangements. We found that LAMP5-AS1 is specifically overexpressed in MLL leukemia patients ( n = 58) than that in the MLL -wt leukemia ( n = 163) ( p < 0.001), and the patients with a higher expression level of LAMP5-AS1 exhibited a reduced 5-year leukemia-free survival ( p < 0.01). LAMP5-AS1 suppression significantly reduced colony formation and increased differentiation of primary MLL leukemia CD34+ cells. Mechanistically, LAMP5-AS1 facilitated the methyltransferase activity of DOT1L by directly binding its Lys-rich region of catalytic domain, thus promoting the global patterns of H3K79 dimethylation and trimethylation in cells. These observations supported that LAMP5-AS1 upregulated H3K79me2/me3 and the transcription of DOT1L ectopic target genes. Conclusions This is the first study that a lncRNA regulates the self-renewal program and differentiation block in MLL leukemia cells by facilitating the methyltransferase activity of DOT1L and global H3K79 methylation, showing its potential as a therapeutic target for MLL leukemia.