A mitochondrial surveillance mechanism activated by SRSF2 mutations in hematologic malignancies
Xiaolei Liu, … , Omar Abdel-Wahab, Peter S. Klein
Xiaolei Liu, … , Omar Abdel-Wahab, Peter S. Klein
Published May 7, 2024
Citation Information: J Clin Invest. 2024;134(12):e175619. https://doi.org/10.1172/JCI175619.
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A mitochondrial surveillance mechanism activated by SRSF2 mutations in hematologic malignancies

Abstract

Splicing factor mutations are common in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML), but how they alter cellular functions is unclear. We show that the pathogenic SRSF2P95H/+ mutation disrupts the splicing of mitochondrial mRNAs, impairs mitochondrial complex I function, and robustly increases mitophagy. We also identified a mitochondrial surveillance mechanism by which mitochondrial dysfunction modifies splicing of the mitophagy activator PINK1 to remove a poison intron, increasing the stability and abundance of PINK1 mRNA and protein. SRSF2P95H-induced mitochondrial dysfunction increased PINK1 expression through this mechanism, which is essential for survival of SRSF2P95H/+ cells. Inhibition of splicing with a glycogen synthase kinase 3 inhibitor promoted retention of the poison intron, impairing mitophagy and activating apoptosis in SRSF2P95H/+ cells. These data reveal a homeostatic mechanism for sensing mitochondrial stress through PINK1 splicing and identify increased mitophagy as a disease marker and a therapeutic vulnerability in SRSF2P95H mutant MDS and AML.

Authors

Xiaolei Liu, Sudhish A. Devadiga, Robert F. Stanley, Ryan M. Morrow, Kevin A. Janssen, Mathieu Quesnel-Vallières, Oz Pomp, Adam A. Moverley, Chenchen Li, Nicolas Skuli, Martin Carroll, Jian Huang, Douglas C. Wallace, Kristen W. Lynch, Omar Abdel-Wahab, Peter S. Klein

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Figure 6

The SRSF2 mutation is associated with accumulation of defective mitochondria.

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The SRSF2 mutation is associated with accumulation of defective mitochon...
(A) Bubble plot for GO enrichment analysis of differentially spliced genes in SRSF2-mutant versus WT K562 cells. The y axis shows the number of genes; the x axis denotes –log[q value]. The size of each circle represents –log[P value]. (B) Bar graph for GO enrichment analysis of differentially spliced genes in SRSF2-mutant cells versus SRSF2+/+ primary CMML and AML patient samples shows significantly dysregulated pathway associated with mitochondrion organization, with Fisher’s exact test –log[q value] on x axis. (C) Venn diagram showing overlap of proteins differentially expressed in SRSF2P95H/+ versus WT K562 cells and MitoCarta3.0 database. (D) Bar graph shows log2 ratio of selected mitochondrial protein in SRSF2P95H/+ versus SRSF2+/+ K562 cells. (E–H) Quantification of mitochondrial parameters in WT and SRSF2P95H/+ cells (mean ± SD). (E) MTG. (F) Mitochondrial DNA copy number. Data are shown as ratio of mitochondrial DNA (mt-ND1) to nuclear DNA (B2M) in WT and SRSF2P95H/+ cells. (G) MMP per mitochondrion. (H) Mitochondrial ROS (mtROS). (I) Basal, maximal, and complex I–linked mitochondrial respiratory capacity was measured in WT and SRSF2P95H/+ cells. Maximal respiratory capacity was measured after FCCP injection. Complex I–linked respiration was measured by sequential addition of the complex I–linked substrates pyruvate, malate, and glutamate (P/M/G) and ADP. Oxygen flux is expressed as respiration per million cells [pmol/(s·106 cells)], mean ± SD of n = 3 independent cultures. Each sample was measured in duplicate. P values were determined by 2-tailed Student’s t test (E–I). (J) Analysis of mitochondrial depolarization in WT and SRSF2P95H/+ cells treated with DMSO or 2 μM CCCP for 24 hours using JC-1 staining, in which a high red+/green+ ratio indicates high MMP (n = 3). Statistical analysis was by 2-tailed χ2 test. (K) Representative RT-PCR results of PINK1 splicing in WT and SRSF2P95H/+ cells treated with DMSO, 3 μM CHIR, or indicated concentrations of CCCP for 24 hours. (L) RT-qPCR analysis of PINK1 mRNA levels in WT and SRSF2P95H/+ cells treated with DMSO, 3 μM CHIR, or 2 μM CCCP for 24 hours (mean ± SD). **P < 0.01, ***P < 0.001, and ****P < 0.0001 (2-way ANOVA with Šidák’s multiple-comparison test).