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. 2021 Oct 16;24(11):103296. doi: 10.1016/j.isci.2021.103296

Figure 5.

Figure 5

ATG5 negatively regulates c-Myc independent of its conjugation with ATG12

(A) 293T cells were transfected with a Flag-tagged ATG5K130R and an HA-tagged c-Myc construct for 48 h. Cells were lysed and subjected to co-immunoprecipitation (co-IP) assays with antibodies against Flag (left) or HA (right) as indicated.

(B) 293T cells transfected with a Flag control and a Flag-tagged ATG5K130R construct were further cultured under normal conditions or serum starvation conditions for 48 h. Western blot analyses were then performed.

(C) 293T cells were transfected with a Flag-tagged ATG5 construct or an ATG12-specific siRNA or together for 48 h. Western blot analyses were then performed.

(D) An HA-tagged c-Myc and an His-tagged ubiquitin construct were transfected with a Flag-tagged ATG5 or ATG5K130R mutant construct into 293T cells as indicated for 48 h. Cells were lysed under denaturing conditions, and the c-Myc protein was immunoprecipitated (IP) with HA-tag-specific antibodies. IP with IgG antibodies was performed as a negative control. The precipitated proteins were subjected to western blot analyses with the indicated antibodies.