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. 2016 Mar 22;7(18):25224–25240. doi: 10.18632/oncotarget.8279

Figure 7. mTOR directly phosphorylates ACL and Ser-455 phospho-mutants affect ACL function.

Figure 7

A. mTOR kinase assay using ACL as a substrate as described in Methods. The reaction mixtures were immunoblotted. B. 293T cells were transiently transfected with the indicated ACL constructs for 48 h. The cells were assayed for cellular acetyl-CoA (left panel) or subjected to immunoblotting (right panel). C. MDA453 stable cell clones overexpressing the indicated ACL constructs were each infected with GIPZ-lentivirus encoding Sh-NT or Sh-ACL#3. Puromycin-resistant cells were subjected to immunoblotting. D. and E. Cells as in C were plated in 24-well plates and then assayed for cell growth by counting live GFP-positive cells (D) or assessed for JC-1 staining (E). For cell growth, values Sh-ACL relative to Sh-NT are shown. Statistical analyses are performed as indicated.