1. A B Supplemental Fig. 1A-C Bap1Δ/Δ NMC MM 526 Bap1 Ub-H2A Nf2 Aldh1a2
p-Yap
Total Yap
NMC
Bap1Δ/Δ
MM 526
Actin
(longer exp.)
Bap1
Nf2
p-Akt
Total Akt
Gapdh
p16Ink4a

2. C Control WT Δ/Δ Bap1Δ/Δ Tail 526 Bap1 Cdkn2a Nf2 MM 526
Supplemental Figure 1. Characterization of malignant mesothelioma (MM) from Bap1f/f mouse, which was detected 45 weeks after intrathoracic injection of Adeno-Cre virus. A, Immunoblot confirming loss of Bap1 expression in MM from Bap1 conditional knockout mouse 526, with tumor also showing greatly reduced expression of Nf2 and p16Ink4a as well as Akt activation when compared to normal mesothelial cells (NMC). B, Immunoblots of same lysates as in panel A showing Bap1-related loss of deubiquitination activity in MM 526, as indicated by presence of Ub-H2A in tumor but not in NMC, along with overexpression of product of PRC2 target gene Aldh1a2. In addition to acquired loss of Nf2 expression (panel A), MM 526 shows loss of p-Yap activity, indicative of aberrant Hippo signaling. C, Demonstration that loss of expression of Nf2 and p16Ink4a is acquired, not due to excision of floxed Nf2 and Cdkn2a alleles. Control WT and control Δ/Δ lanes depict sizes of wild type and homozygously floxed (Δ/Δ) Bap1, Nf2, and Cdkn2a alleles. DNA from Bap1Δ/Δ mouse tail and matching MM from animal 526 verify that Bap1, but not Nf2 or Cdkn2a, has floxed alleles. Note that the primers used to detect mutant Bap1 allele were designed to bind outside the flox sites, such that a smaller band is seen when Cre recombinase excises Bap1 exon 7. C
Control WT
Δ/Δ
Bap1Δ/Δ
Tail 526
Bap1
Cdkn2a
Nf2
MM 526