1. RIF1 interacts with protein phosphatase 1 isoforms
RIF1 interacts with protein phosphatase 1 isoforms Construction of RIF1 cDNA mutated at its PP1 interaction motifs. Critical residues in all three potential PP1 interaction motifs are substituted with alanine, to create a RIF‐pp1bs allele.Expression and localization of GFP‐RIF1 fusion proteins in stably transfected cells. Flp‐In T‐REx 293 cells with GFP, GFP‐RIF1, or GFP‐RIF1‐pp1bs were cultivated with 1 μM doxycycline (DOX) for 3 days, and expression and localization of GFP proteins were confirmed by fluorescence microscopy. Phase‐contrast, DAPI‐stain, and GFP images are shown. Scale bar indicates 25 μm.RIF1 binds PP1 protein isoforms through its PP1 interaction motifs. GFP, GFP‐RIF1, and GFP‐RIF1‐pp1bs proteins were recovered from cell extracts using GFP‐Trap beads, and co‐purifying proteins were analyzed by Western blotting with anti‐GFP (upper two panels) or isoform‐specific PP1 antibodies (lower panels).
Shin‐ichiro Hiraga et al. EMBO Rep. 2017;embr
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