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. Author manuscript; available in PMC: 2022 Aug 12.
Published in final edited form as: Nat Genet. 2022 May 9;54(5):649–659. doi: 10.1038/s41588-022-01061-8

Figure 2. NF2/Merlin drives meningioma apoptosis and susceptibility to cytotoxic therapy.

Figure 2.

a, Meningioma DNA methylation analysis of chromosome 22q segment copy number deletions of any size containing the entire NF2 locus across Merlin-intact (n=32 of 192 meningiomas, 17%), Immune-enriched (n=165 of 216 meningiomas, 76%), and Hypermitotic (n=154 of 157 meningiomas, 98%) DNA methylation groups (n=565, Chi-squared test, two-sided). b, Meningioma NF2 transcripts per million (TPM) expression across Merlin-intact (n=72), Immune-enriched (n=65), and Hypermitotic (n=63) DNA methylation groups (n=200, ANOVA, one-sided). c, Immunoblot for Merlin or GAPDH in 3 meningiomas with loss of at least one copy of the NF2 locus from each meningioma DNA methylation group. d, Confocal microscopy and quantification of Annexin V in M10GdCas9-KRAB cells stably expressing a non-targeting control single-guide RNA (sgNTC) or a single-guide RNA suppressing NF2 (sgNF2) after 24 hours of actinomycin D or vehicle control treatment. DNA is marked with DAPI. Scale bar 10 μM. From left to right, 53, 88, 69, or 56 cells are shown (ANOVA, one-sided). e, Immunoblot for FLAG, cleaved Caspase-7 (cCaspase-7), or GAPDH from CH-157MN xenografts stably expressing doxycycline-inducible Merlin encoding a FLAG tag (NF2-FLAG) in NU/NU mice after 7 days of doxycycline or vehicle treatment, and 24 hours after 4 Gy ionizing radiation or control treatment. f, Immunoblot for ARHGAP35 or FLAG after FLAG immunoprecipitation from CH-157MN cells stably expressing Merlin encoding a FLAG tag (NF2FLAG). EV, empty vector. g, QPCR for NF2 or NR3C1 in M10GdCas9-KRAB cells stably expressing sgNTC, sgNF2, or sgNF2 with NF2 rescue (sgNF2+NF2HA). 3 biological replicates per condition (Student’s t test, one-sided). h, Quantification of Annexin V confocal microscopy in IOMM-Lee cells stably expressing a short-hairpin RNA suppressing NF2 (sgNF2-2) and transiently expressing a non-targeting control siRNA (siNTC) or siRNAs suppressing NR3C1 (siNR3C1). Cells were treated as in d. From left to right, 39, 80, 58, or 52 cells are shown (ANOVA, one-sided). i, NR3C1 TPM expression in euploid meningiomas (n=52) or meningiomas with loss of NF2 as the only CNV (n=28) (Student’s t test, one-sided). j, Model of Merlin pro-apoptotic tumor suppressor function in meningioma cells. Lines represent means, and error bars represent standard error of the means. ***p≤0.0001.