Fig. 1: PHGDH directly interacts with ASS1. | Cell Death & Disease

Fig. 1: PHGDH directly interacts with ASS1.

From: ASS1 inhibits triple-negative breast cancer by regulating PHGDH stability and de novo serine synthesis

Fig. 1

A Silver staining results of 293 T cells stably expressing the empty vector 3xFlag or 3xFlag-ASS1 that were subjected to co-IP assays using anti-Flag affinity gel beads. B The top ten ASS1-interacting proteins according to the number of identified unique peptides. C Validation of the mass spectrometry results of 293 T cells stably expressing the empty vector 3xFlag or 3xFlag-ASS1 by immunoblotting. D Co-IP and reciprocal co-IP assays of ASS1 and PHGDH in293T cells. Cell lysates were subjected to IP with the indicated antibodies and analyzed via IB. Normal rabbit IgG was used as a negative control. E, F Co-IP and reciprocal co-IP assays of ASS1 and PHGDH in MDA-MB-468 and Hs578 cells. Cell lysates were subjected to IP with the indicated antibodies and analyzed via IB. Normal rabbit IgG was used as a negative control. G, H Direct interaction of the recombinant GST-ASS1 and His-PHGDH proteins (G) or the recombinant GST-PHGDH and His-ASS1 proteins (H) analyzed by in vitro GST pull-down assays. The proteins were pulled down with the indicated recombinant protein and analyzed via IB. GST was used as a negative control. I Full-length PHGDH (PHGDH-F) and its truncated forms (N-314 and 317-C) with recombinant GST-ASS1 for the GST pull-down assay. GST was used as a negative control. J Immunofluorescence images showing the colocalization of ASS1 (red) and PHGDH (green) in MDA-MB-468 cells. The nuclei were counterstained with DAPI (blue). Scale bars: 10 μm.

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