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. 2019 Jul 2;2(4):e201800280. doi: 10.26508/lsa.201800280

Figure 6. Interactome validation of proteins involved in degradation pathways.

Figure 6.

(A) IP of endogenous p62 from cell lysates of wild-type N2a, N2a NM-HAsol, and N2a NM-HAagg cells using mAb anti-p62, followed by SDS–PAGE and Western blot. Total cell lysate (extract) was loaded as control. p62 was detected using mAb anti-p62, and NM-HA was detected using mAb anti-HA. Pull-downs using unspecific IgG served as controls. (B) Bulk N2a NM-HAagg cells were either not transfected (for p62 detection) or transfected with constructs encoding for VCP-EGFP, Keap1-GFP, or Ubiquilin-2-FLAG and subjected to immunofluorescence staining 48 h posttransfection. NM-HA was stained using mAb anti-HA (red), p62 was detected using mAb anti-p62 (green), and FLAG was stained using mAb anti-FLAG (green). GFP is shown in green. Nuclei were stained with Hoechst (blue). Scale bar: 5 μm.