Xenopus augmin subunit assembly and interactions. (A) Augmin IP was performed with antibodies IgG, anti-H1 [αH1], αH2, αH3, αH6, and αH8, and checked by Western blot. (B) Proteins in IP samples were identified and quantified by SixplexTMT labeling and liquid chromatography tandem mass spectrometry analyses. (C) Quantified peptides were normalized, providing relative intensities of augmin subunits (H1–H7) in each IP sample. Data were visualized by a heatmap with hierarchical clustering, generated using the heapmaply package of R. (D) Hierarchical clustering dendrograms were drawn to include H8 that directly binds to H6, showing the assembly of all augmin subunits. (E) In vitro pull-down assays were performed to identify all interactions between augmin subunits (Fig. S3). All interactions are summarized; a green square stands for a strong interaction and blue for a weak. Strep-GFP tagged subunits were used as bait and the other seven subunits as prey. (F) All data resulted in a model of augmin assembly in which the direction of arrows point from bait to prey subunits.