Figure 4

Figure 4

LPA₃ is highly internalized and sorted to the lysosome degradation pathway in Progerin cells. (a) Western blot results show that treating Progerin HEK293 cells with 5 μM of MG132 for 6 hr showed no LPA₃ protein restoration. DMSO was used as solvent control (Ctrl). (b) Western blot results show that Progerin HEK293 cells were treated with 5 mM of NH₄Cl or 20 nM of Bafilomycin A1 (BA‐1) for 6 hr to block LPA₃ degradation through the lysosome pathway. ddH₂O was used as solvent control for NH₄Cl (Ctrl in upper panel). DMSO was used as solvent control for BA‐1 (Ctrl in lower panel). (c) Western blot results show that HGPS AG11 fibroblasts were treated with 5 mM of NH₄Cl and 5 μM of MG132 for 6 hr. NH₄Cl treatment rescued LPA₃, but not LPA₁ or LPA₂, in HGPS AG11 fibroblasts. ddH₂O and DMSO were both loaded to control sample as solvent control (Ctrl). (d) Counter‐immunofluorescent staining by LPA₃ and LAMP‐1 shows that LPA₃ localized into the lysosomes in Progerin HEK293 cells. (e) Western blot results show that NH₄Cl treatment rescued LPA₃ in HGPS AG03 fibroblasts. ddH₂O and DMSO were both loaded to control as solvent control (Ctrl). (f) Scheme representing the internalization assay of LPA receptors_. (g) Representative images of time‐dependent LPA₃ internalization. Internalization of LPA₃ was higher in Progerin HEK293 cells. (h) Integrated density of intracellular fluorescence was quantified to indicate internalized LPA₃. ANOVA and Student's t test; **p < .01, ***p < .001