U2AF1 mutations induce oncogenic IRAK4 isoforms and activate innate immune pathways in myeloid malignancies

Abstract

Spliceosome mutations are common in MDS and AML, yet the oncogenic changes due to these mutations have not been identified. A global analysis of exon usage in AML samples revealed distinct molecular subsets containing alternative spliced isoforms of inflammatory and immune genes. IRAK4 was the dominant alternatively spliced isoform in MDS/AML and is characterized by a longer isoform that retains exon 4, encoding a protein, IRAK4-Long (L) that assembles with the Myddosome, results in maximal activation of NF-κB, and is essential for leukemic cell function. Expression of IRAK4-L is mediated by mutant U2AF1 and is associated with oncogenic signaling in MDS/AML. Inhibition of IRAK4-L abrogates leukemic growth, particularly in AML cells with higher expression of the IRAK4-L isoform. Collectively, mutations in U2AF1 induce expression of therapeutically targetable “active” IRAK4 isoforms and provide a genetic link to activation of chronic innate immune signaling in MDS and AML.

Nanopore sequencing reveals U2AF1 S34F-associated full-length isoforms

“…In contrast to gene sets identified from our differential isoform usage analysis, we found that differentially expressed isoforms belonged to genes regulated by NF-kB via TNF signaling, most of which are downregulated (corrected p-value <0.05; Supplemental Table 6). This observation is particularly important given recent reports implicating U2AF1 S34F in altering immune-related genes (Palangat et al, 2019;Smith et al, 2019). We further examined FLAIR isoforms derived from genes in the NF-kB pathway to determine if any U2AF1 S34F-associated splicing alterations could explain expression changes.…”

“…In ADC, the most recurrent U2AF1 mutation occurs at amino acid residue 34, in which a C>T transition causes a change from serine to phenylalanine (S34F). The impact of U2AF1 S34F on pre-mRNA splicing has been widely studied (Przychodzen et al, 2013;Brooks et al, 2014;Coulon et al, 2014;Ilagan et al, 2015;Shirai et al, 2015;Park et al, 2016;Yip et al, 2017;Palangat et al, 2019;Smith et al, 2019), and previous work has shown that mutant U2AF1 has an altered binding affinity with its pre-mRNA substrate (Okeyo-Owuor et al, 2015;Fei et al, 2016). In ADC, mutant U2AF1 has been shown to alter pre-mRNA splicing and other posttranscriptional processes (Brooks et al, 2014;Fei et al, 2016;Park et al, 2016;Palangat et al, 2019).…”

“…The impact of U2AF1 mutations on the transcriptome raises interesting hypotheses for an oncogenic role through mRNA dysregulation. U2AF1 S34F is known to alter alternative-splicing and polyadenylation of cancer-relevant genes (Przychodzen et al, 2013;Brooks et al, 2014;Ilagan et al, 2015;Okeyo-Owuor et al, 2015;Shirai et al, 2015;Yip et al, 2015Yip et al, , 2017Fei et al, 2016;Park et al, 2016;Smith et al, 2019).…”

“…In myelodysplastic syndromes, U2AF1 S34F alters pre-mRNA splicing of Interleukin-1 receptor-associated kinase 4 (IRAK4), producing isoforms that promote activation of kappa-light-chain-enhancer of B cells (NF-kB), a factor known to promote leukemic cell growth (Smith et al, 2019). In addition to splicingdependent functions of U2AF1 S34F, recent studies show a splicing-independent post-transcriptional role for U2AF1 S34F in modulating translational efficiency of genes involved in inflammation and metastasis in human bronchial epithelial cells (Palangat et al, 2019).…”

Ribosome biogenesis is a downstream effector of the oncogenic U2AF1-S34F mutation

“…The mutation is heterozygous, occurs early in disease progression, and is a hotspot mutation in the RNA binding domain of the protein, strongly suggesting a gain of function, oncogenic mutation. While several groups have examined alternative splicing alterations that are mediated by the S34F mutation (Brooks et al, 2014;Fei et al, 2016;Smith et al, 2019;Yoshida et al, 2011), there is no clear set of splicing events that can explain the tumorigenic role of the S34F mutation. Previously, we reported that U2AF1, in complex with U2AF2, plays a noncanonical role in translation regulation through direct interactions with target RNA in the cytosol (Palangat et al, 2019).…”