Figure 6. LRP5.Q89R did not affect binding
to MESD or protein trafficking in GTM3 cells. LRP5.WT-MycHis,
LRP5.Q89R-MycHis or LRP5.G171V was transiently expressed in GTM3 cells
with (A) or without (B) MESD-FLAG by transfection. A:
Co-IP
of LRP5 and MESD protein complex. Input: LRP5.WT, LRP5.Q89R,
LRP5G171V and MESD protein levels were studied by western
immunoblotting with anti-cMyc or anti-FLAG antibodies, respectively.
Whole cell lysate was immunoprecipitated with an anti-FLAG antibody
(IP: anti-FLAG). The enriched protein was analyzed with SDS–PAGE and
detected with anti-cMyc antibodies. B: Pull-down of cell
surface LRP5 proteins. Proteins on the cell surface were labeled with
biotin and pulled down with Neutravidin beads. Protein samples from
input and after pull-down were subjected to Western immunoblotting.
LRP5.WT, LRP5.Q89R and LRP5.G171V were probed with an anti-cMyc
antibody. Experiments were performed in biologic replicates and
representative data were shown.