Figure 6 | Scientific Reports

Figure 6

From: Physiologically generated presenilin 1 lacking exon 8 fails to rescue brain PS1−/− phenotype and forms complexes with wildtype PS1 and nicastrin

Figure 6

PS1 and PS1∆exon8 co-immunoprecipitate in vitro.

Co-IP immunoblots of different epitope-tagged PS1 and PS1∆exon8 proteins. Constructs were cotransfected in six different combinations: (A) (1) empty vectors containing either HA or FLAG only, (2) HA-tagged PS1 and FLAG-tagged PS1 and (3) HA-tagged PS1∆exon8 and FLAG-tagged PS1∆exon8. (B) (4) No transfection, (5) HA-tagged PS1 and FLAG-tagged PS1∆exon8 and (6) FLAG-tagged PS1 and HA-tagged PS1∆exon8. Transfected constructs were co-immunoprecipitated with anti-HA antibody (1.5 μg per 300 μg cell lysate) and blotted with the anti-FLAG antibody (top immunoblot) and the anti-human PS1 antibody NT.1 (directly below FLAG immunoblot). The co-immunoprecipitated band is indicated by arrows and corresponds to full-length PS1 (~50 kDa). Note there is no band present in the eluate lane for Empty Vectors and No Transfection. (C,D) For pooled lysates, constructs were transfected in separate wells and then pooled after lysis. The pooled lysed cells were co-immunoprecipitated with anti-HA antibody and blotted for anti-FLAG as described above. No PS1 band was detected in the eluate (indicated by arrows). (E,F) The experiment in (A,B) was repeated but transfected constructs were co-immunoprecipitated with either IgG rabbit as a negative control or the anti-HA antibody. No PS1 band was detected in the IgG control but was detected in the HA lane, except in the Empty Vectors and No Transfection lanes (indicated by arrows, asterisk marks non-specific bands corresponding to the IgG heavy chain). I = input, E = eluate, IgG = rabbit IgG and HA = HA antibody.

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