Li et al., Supplemental Figures

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1 Li et al., Supplemental Figures Fig. S1. Suppressing TGM2 expression with TGM2 sirnas inhibits migration and invasion in A549-TR cells. A, A549-TR cells transfected with negative control sirna (NC sirna) or two individual TGM2 sirna. Knockdown of TGM2 expression was confirmed by Western blot. B, The migration assay was performed 48 h post transfection. C, Invasion assay. Data shown are mean ± S.D; **P< 0.01 (compared to NC). Fig. S2. Overexpression of TGM2 in A549-WT cells increases migration and invasion. A, A549-WT cells were transfected with an empty vector (pcdna) or pcdna-tgm2. Stable transfectant clones were selected and TGM2 expression was confirmed by Western blot. Note that the ectopic expressed TGM2 has a similar size as that of the endogenous TGM2. B, Migration assay. C, Invasion assay. Data shown are mean ± S.D; **P< 0.01 (compared to NC).

Fig. S3. Restoration of TGM2 expression alleviates TGM2 sirna s inhibition on migration in A549-TR cells. A, A549-TR cells were stably transfected with TGM2-expressing or an empty vector.

2 Fig. S3. Restoration of TGM2 expression alleviates TGM2 sirna s inhibition on migration in A549-TR cells. A, A549-TR cells were stably transfected with TGM2-expressing or an empty vector. The cells were transiently transfected with negative control sirna (NC sirna) or an individual TGM2 sirna (#4). Overexpression and knockdown of TGM2 expression were confirmed by Western blot. Note that the ectopically expressed TGM2 overlaps the endogenous TGM2. The slight reduction of TGM2 level in the TGM2-transfected cells is likely due to removal of endogenous TGM2 by the sirna. B, Migration assay was performed 48 h post transfection. Data shown are mean ± S.D; ** P< Fig. S4. Knockdown of TGM2 expression with individual TGM2 sirna results in MMP-9 suppression in A549-TR cells. A549-TR cells were transfected with negative control sirna (NC sirna) or two individual TGM2 sirna. Expression of the indicated proteins was detected by Western blot. β-actin was detected as a loading control.

3 Fig. S5. Knockdown of TGM2 expression with individual TGM2 sirna sensitizes TRAIL s cytotoxicity in A549-TR cells. A549-TR cells were transfected with negative control sirna (NC sirna) or two individual TGM2 sirna. Expression of the indicated proteins was detected by Western blot as shown in Fig. S1. The cells were treated with TRAIL (150 ng/ml) for 30h. Cell death was detected by LDH leakage assay. Fig. S6. Restoration of TGM2 expression inhibits TGM2 sirna s potetiation on TRAIL s cytotoxicity in A549-TR cells. A549-TR cells were transfected with negative control sirna (NC sirna) or two individual TGM2 sirna. Knockdown of TGM2 expression of TGM2 was shown in Fig. S3. The cells were treated with TRAIL (150 ng/ml) for 30h. Cell death was detected by LDH leakage assay. Data shown are mean ± S.D; ** P< 0.01.

4 Fig. S7. Overexpression of TGM2 in A549-WT cells suppresses TRAIL s cytotoxicity in A549-WT cells. Stable transfectant clones shown in Fig. S2 were treated with TRAIL (150 ng/ml) for 30h. Cell death was detected by LDH leakage assay. Data shown are mean ± S.D; **P< 0.01, *P<0.05 (compared to NC).

5 Fig. S8. Confirmation of the effectiveness of inhibitors in blocking the respective signaling pathways. A, A549-WT cells were pretreated with the EGFR inhibitor (10 µm) for 30 min or left untreated followed by exposure to TGF-α (10 ng/ml) for the indicated times. Phosphorylated and total Akt were detected by Western blot. B, A549-WT cells were pretreated with LY (10 µm) for 30 min or left untreated followed by exposure to TGF-α (10 ng/ml) for the indicated times. Phosphorylated and total Akt were detected by Western blot. C, A549-WT cells were pretreated with U0126 (10 µm) for 30 min or left untreated followed by exposure to TGF-α (10 ng/ml) for the indicated times. Phosphorylated and total ERK were detected by Western blot. D, A549-WT cells were pretreated with SB (10 µm) for 30 min or left untreated following by exposure to UVB (100 mj/cm 2 ) and culture for 30 min. Phosphorylated and total p38 were detected by Western blot. E, A549-Luc (a cell line with a NF-κB-driven luciferase reporter) cells were pretreated with IKK inhibitor II (10 µm) for 30 min or left untreated following by exposure to TNF-α (10 ng/ml) for 6 h. Luciferase was measured. Data shown are mean ± S.D. D, A549- WT cells were pretreated with SP (10 µm) for 30 min or left untreated followed by exposure to TNF-α (10 ng/ml). Phosphorylated and total JNK were detected by Western blot.

Supplementary Fig. 1 Kinetics of appearence of the faster migrating form of Bcl-10.

α-cd3 + α-cd28: Time (min): + + + + + + + + + 0 5 15 30 60 120 180 240 300 360 360 n.s. Supplementary Fig. 1 Kinetics of appearence of the faster migrating form of. Immunoblot of lysates from Jurkat cells