DNA damage repair proteins across the Tree of Life

Abstract

Genome maintenance is orchestrated by a highly regulated DNA damage response with specific DNA repair pathways. Here, we investigate the phylogenetic diversity in the recognition and repair of three well-established DNA lesions, primarily repaired by base excision repair (BER) and ribonucleotide excision repair (RER): (1) 8-oxoguanine, (2) abasic site, and (3) incorporated ribonucleotide in DNA in 11 species: Escherichia coli, Bacillus subtilis, Halobacterium salinarum, Trypanosoma brucei, Tetrahymena thermophila, Saccharomyces cerevisiae, Schizosaccharomyces pombe, Caenorhabditis elegans, Homo sapiens, Arabidopsis thaliana, and Zea mays. Using quantitative mass spectrometry, we identified 337 binding proteins across these species. Of these proteins, 99 were previously characterized to be involved in DNA repair. Through orthology, network, and domain analysis, we linked 44 previously unconnected proteins to DNA repair. Our study presents a resource for future study of the crosstalk and evolutionary conservation of DNA damage repair across all domains of life.

Keywords: Evolutionary biology; Molecular biology; Phylogenetics.

Graphical abstract

Figure 1

Overview of screen for proteins interacting with DNA damage marks (A) Phylogenetic tree and overview of the eleven species included in this study. (B) Experimental setup of the interactomics screen. Pull downs were performed for a control, and for an 8-oxoG, abasic, and RNA base lesion. Pull downs of the respective DNA damage lesion were compared to the common control to calculate enriched interaction partners passing a fold change threshold > 2 with Welch t-test p values < 0.05 (dashed gray line).

Figure 2

Interactors of the DNA damage lesions per species (A) Number of proteins enriched at each lesion in each species highlighted for Gene Ontology annotation “DNA repair” (GO:0006281) (blue) and presence of orthologs in OrthoMCL (yellow). (B) KEGG term overrepresentation of enriched proteins at each lesion across species. Conditions with no enriched KEGG terms are not shown or presented in gray. Gene ratio refers to genes in the dataset (enriched proteins at lesion) over genes in the background (whole genome).

Figure 3

Interactors of the different lesions across phylogenetic branches (A) Bar plot of the total number of enriched proteins at 8-oxoG across species. (B) UpSet plot showing overlap of enriched proteins at the 8-oxoG lesion for the different species based on assigned orthology groups via OrthoMCL. (C) Bar plot of the total number of enriched proteins at abasic lesions per species. (D) UpSet plot showing overlap of enriched proteins at the abasic lesion for the different species based on assigned orthology groups via OrthoMCL. (E) Bar plots of the total number of enriched proteins at the uracil RNA base per species. (F) UpSet plot showing overlap of enriched proteins at the RNA base lesion for the different species based on assigned orthology groups via OrthoMCL.

Figure 4

Conserved interaction partners across the lesions Venn diagrams showing the overlapping enriched proteins at the RNA base, abasic site, and 8-oxoG lesions for (A) A. thaliana, (B) S. cerevisiae, (C) E. coli, and (D) B. subtilis. Overlap in other species is detailed in Table S11.

Figure 5

Conservation of DNA repair orthologs across the Tree of Life (A) Heatmap representing enrichment levels of OrthoMCL orthology groups with GO annotation “DNA repair” (GO:0006281) with two or more enriched proteins across eleven species and 8-oxoG (black), abasic (white) and RNA base (gray) lesions. The color scale represents the fold change in comparison to control samples. Abbreviations: hsap, Homo sapiens; scer, S. cerevisiae; cele, C. elegans; atha, A. thaliana; spom, S. pombe. (B) Maximum likelihood phylogenetic tree of the photolyase gene family including information on detection and enrichment (fold change > 2, Welch t-test p value < 0.05) for the different lesions. White boxes represent proteins that were not detected in the respective experiment. The scale bar in the plots indicates the number of amino acid substitutions per site. (C) Maximum likelihood phylogenetic tree of the MUTY glycosylase gene family. Same as (B).

Figure 6

Network, domain, and phylogenetic analyses implicate novel proteins in DNA repair (A) Networks of enriched proteins across lesions for S. cerevisiae, C. elegans, and T. thermophila. Interactions as established in the STRING database. (B) Classification of non-DNA repair proteins based on Pfam domain annotation. The total number of proteins classified at 8-oxoG was 29, at abasic 75, and at the RNA base 74. (C) Heatmap representing enrichment levels of OrthoMCL orthology groups without GO annotation “DNA repair” (GO:0006281) with two or more enriched proteins across all eleven species and 8-oxoG (black), abasic (white) and RNA base (gray) lesions. The color scale represents the fold change in comparison to control samples. Abbreviations: hsap, Homo sapiens; cele, C. elegans; spom, S. pombe.