Figure 2.

CDYL2a promotes breast cancer cell proliferation and tumor growth while CDYL2b suppresses breast cancer cell migratory, invasive, and metastatic potential. (A) Establishment of stable MDA-MB-231 and Hs578T cells expressing pCDH, Flag-CDYL2a, and Flag-CDYL2b by lentiviral infection. The expression status of Flag-CDYL2a and Flag-CDYL2b in these cell lines was validated by immunoblotting analysis with an anti-Flag antibody. Vinculin was used as a loading control. (B-D) MDA-MB-231 and Hs578T cells stably expressing pCDH, Flag-CDYL2a, or Flag-CDYL2b were subjected to cell proliferation assays using CCK-8 (B) and colony formation assays (C-D). Representative images of the colonies (C) and quantitative results (D) are shown. (E-G) MDA-MB-231 cells stably expressing pCDH, Flag-CDYL2a, or Flag-CDYL2b were injected into mammary fat pads of 6-week-old female BALB/c nude mice (n = 6). After 8 weeks of injections, xenograft tumors were harvested. Photographs of harvested tumors (E), tumor growth curves (F), and tumor weights (G) are shown. (H-J) MDA-MB-231 and Hs578T cells stably expressing pCDH, Flag-CDYL2a, or Flag-CDYL2b were subjected to wound-healing assays (H), Boyden's chamber migration assays and Matrigel-coated invasion assays (I and J). Representative images of the wound-healing assays are included in Supplementary Figure S8A and the corresponding quantitative results are shown in H. Representative images of Transwell migration and invasion assays are shown in I and the corresponding quantitative results are shown in J. (K-L) MDA-MB-231 cells stably expressing pCDH, Flag-CDYL2a, or Flag-CDYL2b were injected into 6-week-old female BALB/c nude mice (n = 6) through the tail vein. After 6 weeks of injections, the lungs were harvested. Representative images of lung metastasis (K) and quantitative results of lung nodules (L) are shown. NS, are not significant differences; *, **, and *** are significant differences at p<0.05, p< 0.01, and p<0.001 levels, respectively.