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. Author manuscript; available in PMC: 2021 Oct 5.
Published in final edited form as: Cell Rep. 2021 Sep 21;36(12):109706. doi: 10.1016/j.celrep.2021.109706

Figure 2. Knockout of ATF3 expression impairs cancer cell growth under serine-deprived conditions.

Figure 2.

(A) ATF3-knockout (KO) cells developed with CRISPR-Cas9 tools and control wild-type (WT) cells were subjected to western blotting to confirm the loss of ATF3 expression.

(B–E) Indicated WT or ATF3KO cells were cultured in the serine/glycine-deprived (Ser(−)) or the complete (Ser(+)) medium for different days. Cell numbers were counted, and growth curves were plotted. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001; Student’s t test; ATF3 KO compared to WT in the absence of serine (Ser(−)); n = 3 (biological replicates).

(F) ATF3KO HCT116 cells developed with an AAV-based precise genome-editing technology and control WT cells were cultured in the serine/glycine-deprived (Ser(−)) or the complete (Ser(+)) medium for different days. The inset shows western blotting results that confirm the loss of ATF3 expression in the KO cells. **p < 0.01; ***p < 0.001; Student’s t test; ATF3 KO compared to WT in the absence of serine (Ser(−)); n = 3 (biological replicates).

(G) WT and ATF3-KO HCT116 cells were implanted in nude mice (5 mice each group) fed with a diet deprived of serine and glycine (SG(−)) or a normal control diet (SG(+)). Tumor volumes were calculated and presented in the plot. *p < 0.05; #p < 0.01; &p < 0.001; Student’s t test.

(H) A representative image of tumors dissected from the nude mice.

(I) Tumor weights were presented in the plot.

(J and K) Tumor sections were stained for Ki67 expression (J), and positive cells were counted (K). *p < 0.05; ***p < 0.001; ns, not significant; Student’s t test.

See also Figure S2.