FIP1L1 Regulates Alternative Polyadenylation of Leukemia-Associated Genes in Acute Myeloid Leukemia

Alternative polyadenylation (APA) is a prevalent mechanism of post-transcriptional gene regulation. APA can impact the protein coding portion of a gene or the length of the 3' untranslated region (UTR), a crucial mRNA region mediating stability, localization, and translation efficiency. Global shifts in APA usage have been reported during development and in cancer transformation; however, the impact of APA in acute myeloid leukemia (AML) is currently unexplored. Therefore, we sought to address whether global shifts in APA patterns can be seen in leukemic blasts and if APA changes contribute to leukemogenesis. Here we report that leukemic blasts exhibit global transcript shortening that contributes to the upregulation of leukemia-associated genes: NRAS, BAALC, and MAPKAPK3. We also implicate FIP1L1 as an important mediator of APA alterations in AML.

To identify whether global trends of APA are seen in leukemia patients, we performed a 3'RNA sequencing experiment comparing healthy CD34+ hematopoietic stem and progenitor cells (HSPCs) with CD34+ enriched leukemic blasts from primary patient samples. We observed a trend in whole transcript shortening, with an emphasis on 3'UTR shortening of affected genes. Importantly, genes with altered APA were among pathways associated with oncogenic transformation and specifically AML. We selected three genes for further functional validation (NRAS, BAALC, and MAPKAPK3) that exhibited a correlation of shortened 3'UTRs with higher mRNA expression. Using mRNA stability assays and luciferase assays, we confirmed that 3'UTR shortening contributes to the upregulation of these leukemia-associated genes in blood cells.

To probe upstream regulators of APA alterations in leukemia, we performed overexpression and knockdown studies of APA machinery members that have altered expression in leukemia patients. We identified FIP1L1 as a novel regulator of APA in AML. Overexpression of FIP1L1 resulted in 3'UTR lengthening and downregulation of our three selected genes. Conversely, knockdown led to 3' UTR shortening and upregulation. Interestingly, altering FIP1L1 expression (both up and downregulation) is detrimental to leukemia cell lines, suggesting that modest changes in gene expression can have a dramatic impact on hematopoietic cell fitness. We conclude that APA changes represent another dysregulated layer of gene expression that can contribute to AML development. Current studies aimed at identifying the role of FIP1L1 in healthy HSPCs will elucidate the feasibility of FIP1L1 as a novel therapeutic target in AML.