Figure 1.
Localization of synovial sarcoma-related proteins in synovial sarcoma SYO-1 cells.
GFP fused proteins were observed using a fluorescence microscope. A, GFP-SS18; B, GFP-SSX1; C, GFP-SSX2; D, GFP-tSS18; E, GFP-tSSX1; F, GFP-tSSX2; G, GFP-SS18-SSX1; H, GFP-SS18-SSX2; I, GFP. Scale bars indicate 5 µm.
Figure 2.
Representative images showing the co-expression of GFP-tagged SS18-SSX2 and of DsRed-monomer-tagged truncated proteins in transfected SYO-1 cells.
Fluorescent proteins were observed using a fluorescence microscope. A, co-expression of GFP-SS18-SSX2 (A1) and DsRed-monomer-tSS18 (A2); B, co-expression of GFP-SS18-SSX2 (B1) and DsRed-monomer-tSSX2 (B2); C, co-expression of GFP-SS18-SSX2 (C1) and DsRed-monomer empty vector (C2); D, co-transfection of GFP-SS18-SSX2 (D1) and DsRed-monomer-tSSX2 (D2) SYO-1 cell in which tSSX2 was not expressed after co-transfection. Scale bars indicate 5 µm.
Figure 3.
Effect of increasing expression of tSSX on the localization of SS18-SSX.
Plasmid pEGFP-SS18-SSX2 (2 µg) was co-transfected in HEK293 cells with pDsRedmonomer-tSSX2 (0, 2, 4, and 6 µg) (A), or DsRedmonomer-tSS18 (0, 2, 4, and 6 µg) (B). The total DNA amount of transfection was complimented by pCMV-Tag2B empty plasmid without fluorescent protein (6, 4, 2, and 0 µg). The cells showing either a speckled pattern or a diffuse pattern of SS18-SSX2 localization were counted in 30 fields of fluorescence microscope, and the relative ratio of cells with speckled pattern was calculated (C). Scale bars indicate 10 µm.
Figure 4.
Exogenous expression of tSSX2 suppresses the proliferation of synovial sarcoma SYO-1 cells.
SYO-1 cells were transfected with pEGFP-tSSX2 or pEGFP vector, split 48 h after transfection, and the cells expressing the GFP-tagged fusion proteins were counted on day 4, 6 and 8 after transfection. The cell proliferation ratio was normalized by dividing the number of cells expressing GFP-tagged proteins on days 6 and 8 by the number on day 4. Error bars indicate standard deviation, *p < 0.001, **p < 0.0001.
Figure 5.
Representative images showing changes in proliferation of SYO-1 cells expressing GFP-tSSX2 monitored for a period of 18 days.
SYO-1 cells were transfected with pEGFP-tSSX2 (A) or pEGFP vector (B), split 48 h after transfection, and the cells expressing GFP-tagged proteins were observed under fluorescence microscope on day 4, 6, 8, 10, and 18 after transfection. 1, day 4; 2, day 6; 3, day 8; 4, day 10; 5, day18. Scale bars indicate 20 µm.
Figure 6.
A. Exogenous expression of tSSX suppresses the colony formation in SYO-1 cells.
SYO-1 cells cultured in 90 mm dishes were transfected with pEGFP empty vector (A1) or pEGFP-tSSX2 (A2), split into 10 plates of 60 mm dishes 48 h after transfection and selected with G418 for three weeks. The cells were fixed in 4% formaldehyde and stained with Giemsastain solution. B, YaFuSS cells were transfected with pEGFP empty vector (B1) or pEGFP-tSSX2 (B2), and assessed for colony formation activity. The colonies were counted and the average number of colonies formed is shown as bar graphs for SYO-1 (A3) and YaFuSS (B3). Error bars indicate standard deviation, * p < 0.01.