Journal: Brain : a journal of neurology

Article Title: Inhibition of YAP/TAZ-driven TEAD activity prevents growth of NF2-null schwannoma and meningioma.

doi: 10.1093/brain/awac342

Figure Lengend Snippet: Figure 1 Proliferation of schwannoma cells in the DRG and VG is dependent upon both YAP and TAZ proteins. (A–H) Images of DRG (A–D) and VG (E–H) from 5-month-old mice stained for EdU and neurofilament (NF); sections were counterstained with Hoechst to reveal nuclei (Ho). Arrows indicate EdU-positive nuclei in the areas of the ganglia in close proximity to the neuronal cell bodies; note fewer proliferating cells in NF2/YAP (C and G) and NF2/TAZ (D and H) than in NF2 single null (B and F) ganglia. (I and J) Quantification of EdU-positive cells per area of ganglion tissue of DRG (I) and VG (J). Note significant decreases in proliferation in both NF2/YAP and NF2/TAZ ganglia. (K–R) Staining of DRG sections from 9-month-old control (NF2fl/

Article Snippet: Primary antibodies used for western blotting: YAP (1:1000; #14074; CST), TAZ (1:500; sc-48805; Santa Cruz), Pan-TEAD (1:1000; #13295; CST), GAPDH (1:5000; AB2302; Merck), ALDH1A1 (1:1000, ab52492; Abcam) and CTGF (1:500, ab6992, Abcam).

Techniques: Staining, Control

Journal: Brain : a journal of neurology

Article Title: Inhibition of YAP/TAZ-driven TEAD activity prevents growth of NF2-null schwannoma and meningioma.

doi: 10.1093/brain/awac342

Figure Lengend Snippet: Figure 4 Increased numbers of macrophages in NF2-null DRG tissue is dependent upon YAP and TAZ. (A–L) Staining of DRG sections from NF2fl/fl-CRE- (A, E and I), NF2fl/fl-CRE+ (B, F and J), NF2fl/flYAPfl/fl-CRE+ (C, G and K) and NF2fl/flTAZfl/fl-CRE+ (D, H and L) animals at 3, 5 and 9 months of age with pan- macrophage marker Iba1 antibody. Note time-dependent increase in numbers of Iba1-positive macrophages in NF2fl/fl-CRE+ DRG between 3 (B), 5 (F) and 9 (J) months. (M–P) Staining of sections of VG tissue from NF2fl/fl-CRE- (M and O) and NF2fl/fl-CRE+ (N and P) at 3 and 5 months of age with Iba1 anti body. (Q) Quantification of % macrophages of total cell number in DRG tissues. (R) Quantification of percentage of macrophages at 3 and 5 months in NF2fl/fl-CRE- and NF2fl/fl-CRE+ VG tissue. For data presented, n = 3 mice for each genotype and age. Data presented in graphs are means ± SEM; in Q and R, two-way ANOVA was used with Tukey’s multiple comparison test. *P < 0.05; **P < 0.01; ***P < 0.001. Scale bars = 25 μm.

Article Snippet: Primary antibodies used for western blotting: YAP (1:1000; #14074; CST), TAZ (1:500; sc-48805; Santa Cruz), Pan-TEAD (1:1000; #13295; CST), GAPDH (1:5000; AB2302; Merck), ALDH1A1 (1:1000, ab52492; Abcam) and CTGF (1:500, ab6992, Abcam).

Techniques: Staining, Marker, Comparison

Journal: Brain : a journal of neurology

Article Title: Inhibition of YAP/TAZ-driven TEAD activity prevents growth of NF2-null schwannoma and meningioma.

doi: 10.1093/brain/awac342

Figure Lengend Snippet: Figure 5 Knockdown of either YAP or TAZ or use of TEAD auto-palmitoylation inhibitors inhibits human schwannoma cell proliferation. (A–G) Lentiviral knockdown of either YAP or TAZ significantly reduces proliferation of human NF2-null schwannoma cells. (A–C) Representative images of EdU-positive cells, counterstained with Hoechst (Ho) from scrambled control (Scr, A), YAP knockdown (shYAP, B) and TAZ knockdown (shTAZ, C). (D) Quantification of percentage positive EdU cells for each condition. (E–G) Western blot (E) and quantification (F and G) confirming YAP or TAZ knockdown in cells; note that knockdown of TAZ does not significantly affect levels of YAP (F), but knockdown of YAP does significantly lower levels of TAZ (G). (H–K) TEAD auto-palmitoylation inhibitor (VT3) decreases human schwannoma cell proliferation in a dose-dependent manner. (L) Expression of four TEAD isoforms (TEAD1–4) in cells from three human schwannoma tumours S1, S2 and S3 treated with either vehicle (left) or 2 μm VT3 for 7 days (VT, right). (M) Quantification of schwannoma cell proliferation with increasing concentrations of auto-palmitoylation inhibitor VT3. N = 3 for all data shown. Data presented in graphs are means ± SEM. Data analysis in D was one-way ANOVA with Bonferroni’s correction, in F and G matched one-way ANOVAs with the Geisser–Greenhouse correction and Tukey’s multiple comparisons test and M one-way ANOVA with Bonferroni’s correction. *P < 0.05; **P < 0.01; ***P < 0.001. Scale bars = 25 μm.

Article Snippet: Primary antibodies used for western blotting: YAP (1:1000; #14074; CST), TAZ (1:500; sc-48805; Santa Cruz), Pan-TEAD (1:1000; #13295; CST), GAPDH (1:5000; AB2302; Merck), ALDH1A1 (1:1000, ab52492; Abcam) and CTGF (1:500, ab6992, Abcam).

Techniques: Knockdown, Control, Western Blot, Expressing

Journal: Brain : a journal of neurology

Article Title: Inhibition of YAP/TAZ-driven TEAD activity prevents growth of NF2-null schwannoma and meningioma.

doi: 10.1093/brain/awac342

Figure Lengend Snippet: Figure 8 ALDH1A1 is upregulated in NF2-null human meningioma tissue and cells. (A and B) Staining of sections of NF2-postive (NF2+/+, A) and NF2-null (NF2−/−, B) meningioma tissue shows higher ALDH1A1 in NF2-null tumours. Boxes show enlarged section of the tumour with strong cytoplas mic stain of ALDH1A1 protein. (C) Western blot of control human meningeal cells (HMC), BenMen-1 (BM1) and KT21-MG1 (KT21) meningioma cells showing elevated ALDH1A1 expression in both human NF2-null cell lines. (D) Quantification of western blot in C. (E) Lentiviral shRNA knockdown of YAP or TAZ in BenMen-1 cells showing that ALDH1A1 expression is dependent upon TAZ. (F) Quantification of western blot in E. (G–I) ALDH1A1 in hibitor 1 reduces proliferation of BenMen-1 cells. (J) Quantification of percentage Ki-67-positive cells with increasing ALDH1A1 inhibitor concentra tions. (K–M) Comparison of the effects of shRNA knockdown of ALDH1A1 (L) and TAZ (M) upon cell proliferation in BenMen-1 cells. (N) Quantification of percentage EdU-positive cells. For data presented, n = 3. Data presented in graphs are means ± SEM. Statistical analysis shown is a one-way ANOVA with Bonferroni’s correction. *P < 0.05; **P < 0.01; ***P < 0.001; ns = not significant. Scale bars = 25 μm.

Article Snippet: Primary antibodies used for western blotting: YAP (1:1000; #14074; CST), TAZ (1:500; sc-48805; Santa Cruz), Pan-TEAD (1:1000; #13295; CST), GAPDH (1:5000; AB2302; Merck), ALDH1A1 (1:1000, ab52492; Abcam) and CTGF (1:500, ab6992, Abcam).

Techniques: Staining, Western Blot, Control, Expressing, shRNA, Knockdown, Comparison

Journal: Cell death & disease

Article Title: The microprotein encoded by exosomal lncAKR1C2 promotes gastric cancer lymph node metastasis by regulating fatty acid metabolism.

doi: 10.1038/s41419-023-06220-1

Figure Lengend Snippet: Fig. 6 Pep-AKR1C2 promotes the expression of FAO-related CPT1A by decreasing YAP phosphorylation. A Silver staining analysis of the proteins interacting with pep-AKR1C2. B Pep-AKR1C2 encoded by exo-lncAKR1C2 decreases p127-YAP levels in HLECs cells. C Pep-AKR1C2 promotes the nuclear translocation of YAP. D Immunofluorescence colocalization analysis was used to determine the interaction between YAP and pep-AKR1C2. E, F Determination of CPT1A mRNA (E) and protein levels (F) in HLECS treated as above. G. Knockdown of CPT1A rescues the high-consumption level of fatty acids mediated by overexpressed lnc-AKR1C2 (n = 3). H Knockdown of CPT1A rescues the high ATP production levels mediated by overexpressed lnc-AKR1C2 (n = 3). ** indicates p < 0.01.

Article Snippet: Protein extraction was blocked with PBS-5% fat-free dried milk at room temperature for 1 h and incubated at 4 °C overnight with anti-CPT1A (1:1000; Abcam, Cambridge, UK; ab234111), anti-Flag (1:5,000; Abcam, Cambridge, UK; ab205606), antiYAP (1:1000, Abcam; ab205270), anti p127-YAP (1:1000, Abcam; ab76252), anti-pep-AKR1C2 (1:1000, CST, 13035 S), anti-VEGFC (1:1000, Abcam; ab9546), anti-HIF-1α (Santa Cruz, sc-13515) anti-TSG101 (1:200; Santa Cruz, sc-7964), anti-Alix (1:1000, Abcam, ab275377), anti-CD9 (1:1000, Abcam, ab223052), anti-H3 (1:1000, Abcam, ab1791) and anti-GAPDH (1:3000, Santa Cruz, sc-365062) antibodies respectively.

Techniques: Expressing, Phospho-proteomics, Silver Staining, Translocation Assay, Knockdown