SF3B1 mutation and ATM deletion codrive leukemogenesis via centromeric R-loop dysregulation
Martina Cusan, … , Ren-Jang Lin, Lili Wang
Martina Cusan, … , Ren-Jang Lin, Lili Wang
Published July 18, 2023
Citation Information: J Clin Invest. 2023;133(17):e163325. https://doi.org/10.1172/JCI163325.
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SF3B1 mutation and ATM deletion codrive leukemogenesis via centromeric R-loop dysregulation

Abstract

RNA splicing factor SF3B1 is recurrently mutated in various cancers, particularly in hematologic malignancies. We previously reported that coexpression of Sf3b1 mutation and Atm deletion in B cells, but not either lesion alone, leads to the onset of chronic lymphocytic leukemia (CLL) with CLL cells harboring chromosome amplification. However, the exact role of Sf3b1 mutation and Atm deletion in chromosomal instability (CIN) remains unclear. Here, we demonstrated that SF3B1 mutation promotes centromeric R-loop (cen-R-loop) accumulation, leading to increased chromosome oscillation, impaired chromosome segregation, altered spindle architecture, and aneuploidy, which could be alleviated by removal of cen-R-loop and exaggerated by deletion of ATM. Aberrant splicing of key genes involved in R-loop processing underlay augmentation of cen-R-loop, as overexpression of the normal isoform, but not the altered form, mitigated mitotic stress in SF3B1-mutant cells. Our study identifies a critical role of splice variants in linking RNA splicing dysregulation and CIN and highlights cen-R-loop augmentation as a key mechanism for leukemogenesis.

Authors

Martina Cusan, Haifeng Shen, Bo Zhang, Aijun Liao, Lu Yang, Meiling Jin, Mike Fernandez, Prajish Iyer, Yiming Wu, Kevyn Hart, Catherine Gutierrez, Sara Nik, Shondra M. Pruett-Miller, Jeremy Stark, Esther A. Obeng, Teresa V. Bowman, Catherine J. Wu, Ren-Jang Lin, Lili Wang

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Figure 3

SF3B1 mutation–associated cen-R-loop dysregulation leads to aneuploidy.

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SF3B1 mutation–associated cen-R-loop dysregulation leads to aneuploidy....
(A–C) Analysis of 2-dimensional cross-sectional area of the entire body of chromosomes (A) and aberrant mitosis frequency (B) and spindle length and width (C) in HEK293T SF3B1 WT and MT overexpressing either empty vector (EV) or WT RNH1. (D) DRIP-qPCR in HEK293T SF3B1 WT and MT overexpressing either dCas9–GFP–RNaseH1 WKKD (WKKD RNH1) or dCas9–GFP–RNaseH1 WT (WT RNH1) in combination with either sgRNA guide control (sgCTRL) or sgRNA targeting α-satellite centromeric repeats (sgAlphaSat). Centromeric arrays: The chromosome is specified by the number following the “D”; and the array is specified by the number following the “Z.” Two-way ANOVA test. (E and F) Analysis of 2-dimensional cross-sectional area of the entire body of chromosomes (E) and aberrant mitosis frequency (F) in cells from D. Box plots show the median and 25th and 75th percentiles, with whiskers extending to minimum and maximum values. Bar graphs represent mean ± SD. Each dot represents a biological replicate. Two-tailed unpaired t test followed by Bonferroni’s post hoc test, except in D.