Figure 2: Low PHGDH expression promotes integrin α_vβ₃-mediated invasion and migration.

a. Percentage of migratory cells per migratory position (n=14) in primary tumors derived from orthotopically injected (m.f.p.) 4T1 (mixture of shSCR mTurquoise and shPHGDH Dendra) cells assessed by time-lapse intravital imaging (n=8). Unpaired t test with Welch’s correction, two-tailed.

b. Representative track length of migratory shSCR-mTurquoise (n=9 within one position) and shPHGDH-Dendra (n=10 within one position) cells in primary tumors from the orthotopic (m.f.p.) 4T1 mouse model, assessed by time-lapse intravital imaging.

c. Number of lung metastases per mouse in the orthotopic (m.f.p.) 4T1 (left panel) and EMT6.5 (right panel) mouse model, injected with either shSCR (n=11 and n=18) or shPHGDH cells (n=10 and n=19). Unpaired t test with Welch’s correction, two-tailed.

d. Correlation plot of GSVA-derived Z-scores for the Integrin-1 pathway, versus the scores for the Hallmark EMT gene signature, based on scRNA-seq data from primary tumors of 13 TNBC patients. Color code indicates the Z-Score for a gene expression signature indicative of low PHGDH protein expression. Total least-squares regression line and confidence intervals are overlaid on top, and Pearson correlation coefficient (R) values are shown on the top-left corners.

e. Invasive ability of 4T1 cells pre-treated (24h) with an antibody against integrin α_vβ₃ or control IgG (2.5 μg/ml) and upon Phgdh knockdown (shPHGDH) compared to control (shSCR) cells in a 3D matrix. The invasive area was stained with calcein green. Each dot represents a different microscopy field (n=5). Welch and Brown-Forsythe ANOVA with Dunnett’s multiple comparison.

For panels a, c, and e, the solid lines indicate the median, the boxes extend to the 25th and 75th percentiles, the whiskers span the minimum and maximum values.