Fig. 3

?N-p73? Containing ALS Patient Mutations Expressed in Neuro-2A Cells Displayed Impeded Binding to Proapoptotic p53 (A) Western blots show equal input level of protein (proliferating cell nuclear antigen [PCNA], ?N-p73?, and p53) before immunoprecipitation. Protein was immunoprecipitated with ?N-p73? or PCNA antibody. Postimmunoprecipitation lysate demonstrates that all ?N-p73? protein was immunoprecipitated with ?N-p73? antibody. Blots were probed for ?N-p73?, p53, and PCNA. PCNA was used as a loading control and was immunoprecipitated in tandem to ?N-p73?. Red arrows denote less intense p53 protein bands immunoprecipitated by mutant ?N-p73? compared to wild-type (WT) ?N-p73? and enhanced green fluorescent protein (EGFP) (in EGFP lane, ?N-p73? band is endogenous protein). (B) Quantification of p53 band intensity of Western blots performed after ?N-p73? immunoprecipitation. Shown are mean intensity and SD for p53 bands from coimmunoprecipitations performed in biological duplicate for Neuro-2A cells expressing ST ?N-p73? or mutant ?N-p73? on doxycycline (dox) exposure. ALS = amyotrophic lateral sclerosis.