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. 2022 Feb 15;9(12):2105222. doi: 10.1002/advs.202105222

Figure 6.

Figure 6

VCP, UBXD8, AP2A, and ACSL3 are required for lipid droplet‐mediated Numb ubiquitination. A) Immunoprecipitation and Western blot showing the interaction of VCP and MDM2 in HCT116 and HT29 cells. B,C) Fluorescent staining (3D‐SIM) showing the influence of UBXD8 knockdown (shUBXD8‐1 and shUBXD8‐2) on MDM2 (B) and VCP (C) localization on lipid droplets in F. nucleatum‐infected HCT116 cells. Scale bar, 5 µm. D) Western blot showing the influence of UBXD8 knockdown (shUBXD8‐1 and shUBXD8‐2) on Numb and NICD levels in HCT116 and HT29 cells. E) Immunoprecipitation and Western blot showing the interaction of AP2A and Numb in HCT116 and HT29 cells. F) Fluorescent staining (3D‐SIM) showing localization of AP2A on lipid droplets. Scale bar, 5 µm. G) Fluorescent staining (3D‐SIM) showing the influence of AP2A knockdown (shAP2A‐1 and shAP2A‐2) on Numb localization on lipid droplets in F. nucleatum‐infected HCT116 cells. Scale bar, 5 µm. H) Western blot showing the influence of AP2A knockdown (shAP2A‐1 and shAP2A‐2) on Numb and NICD levels in HCT116 and HT29 cells. I) Immunoprecipitation and Western blot showing the interaction of lipid protein ACSL3 and AP2A in HCT116 and HT29 cells. J,K) Fluorescent staining (3D‐SIM) showing the influence of ACSL3 knockdown (shACSL3‐1 and shACSL3‐2) on the localization of Numb (J) and AP2A (K) on lipid droplets in F. nucleatum‐infected HCT116 cells. Scale bar, 5 µm. L) Western blot showing the influence of ACSL3 knockdown (shACSL3‐1 and shACSL3‐2) on Numb and NICD levels in HCT116 and HT29 cells.