Fig. 1: Identification of the C. elegans ortholog of SPT2.
From: The histone chaperone SPT2 regulates chromatin structure and function in Metazoa
a, Left, schematic representation of the three evolutionarily conserved regions found in SPT2 orthologs and in C. elegans T05A12.3. N-terminal, central and C-terminal (HBD) regions are shown shaded in red, yellow and violet, respectively. Right, multiple sequence alignments corresponding to the three conserved regions are shown inside colored boxes in red, yellow and violet, respectively. The amino acid coloring scheme indicates the average BLOSUM62 score (correlated to amino acid conservation) in each alignment column: black (greater than 3.5), gray (between 3.5 and 1.5) and light gray (between 1.5 and 0.5). Sequences are named according to their UniProt identifier. Species abbreviations: Q9GYK8_CAEEL, C. elegans; SPT2_HUMAN, Homo sapiens; SPT2_DROME, Drosophila melanogaster; SPT2_YEAST; Saccharomyces cerevisiae. b, Structural homology model for the putative HBD of C. elegans T05A12.3 (blue). This was generated using the crystal structure of the H. sapiens SPT2 HBD (yellow) in complex with the H3–H4 tetramer (shaded white) as a search template (PDB code 5BS7 (ref. 26)). The positions of the αC1 and αC2 helices and the connecting loop are shown. c, Same as b, except that only the CeSPT-2 HBD is shown, color-coded according to the degree of amino acid conservation. d, Coomassie gel staining of recombinant full-length HsSPT2 and CeSPT-2 produced in bacteria; one representative experiment of two. e, Pull-down of full-length recombinant CeSPT-2 or human HsSPT2 with beads covalently coupled to histone H3–H4 in the presence of 500 mM NaCl. One representative experiment of two is shown. f, H3–H4 pull-down with recombinant CeSPT-2 HBD (WT or HBM M627A), or HsSPT2 HBD (WT or HBM M641A). One representative experiment of three is shown. g, Coomassie gel staining of recombinant full-length CeSPT-2 (WT and HBM); one representative experiment of two. h,i, H3–H4 pull-down with full-length recombinant CeSPT-2 WT and HBM (h). Quantification (i) of the five independent replicates of the H3–H4 pull-down. n = 5, data are represented as mean ± s.d. normalized to WT. One-sided t-test with Welch’s correction, P = 0.0016.
