1. b c * a
Supplementary Figure 1. NLRP1 knockdown suppresses tumor growth in vivo. (a) 1205Lu tumors with NLRP1 shRNA from Santa Cruz Biotechnology. (b) 1205Lu tumors with NLRP1 shRNA from Origene. (c) HS294T tumors with NLRP1 shRNA from Santa Cruz Biotechnology. Mean ± s.e.m., n = 10 (a), 6 (b) or 8 (c). *P < 0.05.

2. 1205Lu
HS294T
Supplementary Figure 2. NLRP1 knockdown increases apoptosis analyzed by flow cytometry. Cells were exposed to 1 μM AMD for 18 h. (a) Control shRNA-transduced cells treated with DMSO as vehicle. (b) NLRP1 shRNA-transduced cells with DMSO. (c) Control shRNA-transduced cells with AMD. (d) NLRP1 shRNA-transduced cells with AMD. a b c
/1.69/1 1.1
1.69/1.41 d
Annexin V PI

3. THP-1
Supplementary Figure 3. Annexin V staining shows that NLRP1 knockdown by NLRP1 siRNA had no effects on apoptosis in THP-1 cells in the absence or presence of AMD. Mean ± s.e.m., n = 3. KD, knockdown.

4. IgG
NLRP1
1205Lu
NLRP3 IP
HS294T
Heavy chain
Light chain
150 kDa
100 kDa
37 kDa
50 kDa
75 kDa
25 kDa
20 kDa
15 kDa
250 kDa M
Supplementary Figure 4. Ponceau S (Sigma) staining of the blot depicted in Figure 5a shows the ~55 kDa heavy chains and ~23 kDa light chains of the immunoprecipitating antibody. M, molecular weight marker.

5. WM35
WM115 *
*** * **
Supplementary Figure 5. NLRP1 knockdown enhances caspase-3/7 activity in WM35 and WM115 treated with DMSO as vehicle, AMD or CPT. Mean ± s.e.m., n = 3. *P < 0.05, **P < 0.01, and ***P < KD, knockdown.

6. *** *
1205Lu
*** *
HS294T
Supplementary Figure 6. NLRP1 knockdown results in decreased caspase-1 activity in 1205Lu and HS294T cells treated with AMD for 18 h. *P < 0.05 and ***P < Mean ± s.e.m., n = 3. KD, knockdown.

7. 1205Lu
HS294T
Supplementary Figure 7. NLRP1 inflammasome activation by LT reduces apoptosis assayed by flow cytometry. Cells were stimulated with LT for 2 h, then AMD for another 18 h. (a) Vehicle. (b) LT. (c) AMD. (d) LT plus AMD. a b c
/1.69/1 1.1
1.69/1.41 d
Annexin V PI

8. THP-1 **
*** *
1205Lu ** *
Supplementary Figure 8. LT enhances caspase-1 activity in differentiated THP-1, but not in 1205Lu cells, treated with LT for 2 h and then with AMD or CPT for 18 h. MSU and IL-1α were used as controls. Mean ± s.e.m., n = 3. *P < 0.05, **P < 0.01, and ***P <

9. Caspase-2
Caspase-1
Caspase-9
CyPA
HSP90
Lamin B
Untreated LT
PMA
PMA + LT
THP-1
Cytoplasmic
Untreaed
Nuclear
1205Lu
Supplementary Figure 9. Cytoplasmic localization of caspase-2 and -9 in THP-1 and 125Lu cells assayed by Western blot. For THP-1 cells, the subcellular fractions were from Figure 1d. CyPA, cyclophilin A.
20 μm