Fig. 2
From: Multilevel interrogation of H3.3 reveals a primordial role in transcription regulation
Tetrahymena Cac2 and Hir1 have highly conserved Asf1-interacting B-domain-like sequences. A Left, Neighbor-joining phylogenetic analysis of HIRA and Cac2 proteins. Different subfamilies are highlighted in different colors. The numbers on the branches represent confidence values based on 1000 bootstrap replicates. Right, Comparative domain analysis of Tetrahymena Cac2Tt and Hir1Tt proteins against H. sapiens, and S. cerevisiae orthologs. Highly conserved B-domain sequences are shown as multiple sequence alignments for both Cac2Tt and Hir1Tt proteins. B Visualization of the predicted binding interface between TTHERM_00219420 (Cac2) and TTHERM_00442300 (Asf1). Cac2Tt is colored in cyan; Asf1Tt is colored green. The B-domain of Cac2Tt is highlighted in red. Labeled residues (K87-G531, D89-K534, R146-D372) are predicted to form polar intermolecular contacts between Asf1Tt and Cac2Tt within 3 Å, as well as an intramolecular π interaction (F393-K535) involving a Lysine residue within the B-domain of Cac2Tt (T527–Y545). All interactions are shown as dashed yellow lines. C Western blotting analysis using whole cell lysates prepared from growing Tetrahymena cells expressing Cac2Tt -FZZ (left; Cac2 ∼ 63 kDa + FZZ ∼ 18 kDa) and Hir1Tt-FZZ (right; Hir1 ∼ 117 kDa + FZZ ∼ 18 kDa). The blots were probed with the indicated antibodies. D Dot plot representation of the interaction partners identified with Cac2Tt, Hir1Tt, Hat1Tt, and RebL1 in growing Tetrahymena cells. Inner circle color shows the average spectral count, the circle size indicates the relative prey abundance, and the circle outer edge is the SAINT FDR. See Additional file 1: Tables S5, S6, and S8 for complete AP-MS data