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. 2017 Feb 1;8(12):19684–19698. doi: 10.18632/oncotarget.14962

Figure 4. PHF14 and KIF4A formed a physical complex and co-localized extensively in the nucleus during mitosis.

Figure 4

A. FPLC strategy to identify protein complexes in nuclear extract from HeLa cells. PHF14, KIF4A, CENPE, and Cyclin D1 were detected in the same fractionations using western blotting analysis. B. Co-immunoprecipitation of endogenous PHF14 and KIF4A in A549 cells. C. and D. Mapping of the interaction site between PHF14 and KIF4A. Upper panels of (C): co-immunoprecipitation analysis of myc-tagged PHF14 or its mutants with GFP-tagged WT KIF4A. Myc-PHF14 or its mutants and GFP-KIF4A were co-overexpressed in HCT-116 cells. Lower panels of (C): schematic representation of PHF14 mutants. Upper panel of (D): co-immunoprecipitation analysis of GFP-tagged WT KIF4A or deletion mutants with PHF14. GFP-tagged WT KIF4A or deletion mutants were expressed in 293T cells. Lower panel of (D): schematic representation of KIF4Amutants. The affinity of the binding was rated as follows: +++: intensive; ++: moderate; +: weak; -: binding not detectable. E. Co-localization of endogenous PHF14 and KIF4A in A549 cells. A549 cells were stained with anti-PHF14 antibodies (green), anti-KIF4A antibodies (red) and DAPI (DNA, blue). Scale bar = 5 μm.