Diversification of AID/APOBEC-like deaminases in metazoa: multiplicity of clades and widespread roles in immunity

Abstract

AID/APOBEC deaminases (AADs) convert cytidine to uridine in single-stranded nucleic acids. They are involved in numerous mutagenic processes, including those underpinning vertebrate innate and adaptive immunity. Using a multipronged sequence analysis strategy, we uncover several AADs across metazoa, dictyosteliida, and algae, including multiple previously unreported vertebrate clades, and versions from urochordates, nematodes, echinoderms, arthropods, lophotrochozoans, cnidarians, and porifera. Evolutionary analysis suggests a fundamental division of AADs early in metazoan evolution into secreted deaminases (SNADs) and classical AADs, followed by diversification into several clades driven by rapid-sequence evolution, gene loss, lineage-specific expansions, and lateral transfer to various algae. Most vertebrate AADs, including AID and APOBECs1-3, diversified in the vertebrates, whereas the APOBEC4-like clade has a deeper origin in metazoa. Positional entropy analysis suggests that several AAD clades are diversifying rapidly, especially in the positions predicted to interact with the nucleic acid target motif, and with potential viral inhibitors. Further, several AADs have evolved neomorphic metal-binding inserts, especially within loops predicted to interact with the target nucleic acid. We also observe polymorphisms, driven by alternative splicing, gene loss, and possibly intergenic recombination between paralogs. We propose that biological conflicts of AADs with viruses and genomic retroelements are drivers of rapid AAD evolution, suggesting a widespread presence of mutagenesis-based immune-defense systems. Deaminases like AID represent versions "institutionalized" from the broader array of AADs pitted in such arms races for mutagenesis of self-DNA, and similar recruitment might have independently occurred elsewhere in metazoa.

Keywords: DNA/RNA editing; arms race; biological conflicts; immunity; retroelements.

Fig. 1.

Characterization of the AAD protein family. (A) MSAs of representatives of AAD clades labeled with accessions, clade names, and species abbreviations (full species names in SI Appendix, Fig. S1). Zn-chelating active site residues are highlighted in black, and known substrate-interacting residues are shown in red. The SNAD1/2-specific helix between strands 1 and 2 and neomorphic strands are shown in gray rectangles. (B) Topology diagram depicting the conserved core common to all AADs, the nucleic acid substrate, and key residues. (C) Topologies of clades showing Zn-chelating features. Homologous Zn-chelating residues between APOBEC4 and the Cnidaria-algae clade are highlighted in purple, green, and blue, whereas other Zn-chelating residues are colored gray. (D) Topologies of SNADs1–3: inserts embedded within the core domain are in blue, while N- and C-terminal extensions are in gray.

Fig. 2.

Phylogenetic relationships among AADs. (A) Higher order relationships of deaminases with a focus on AADs. Clade-specific features are on the Right. Zn-chelating residues are shown within parentheses and homologous residues shared by members of the cnidaria-algae-APOBEC4 clade are colored. (B) Phylogenetic tree illustrating LSEs of AADs in various metazoan lineages. Clades entirely composed of monospecific representatives are collapsed and labeled with species abbreviations and number of sequences in the LSE. Nodes supported by bootstrap values >80% and >90% are marked with green-outlined and red-outlined circles, respectively (also see SI Appendix, Fig. S21). Hel, helix; L, loop; SP, signal peptide; Str, strand.

Fig. 3.

(A) Illustration of human APOBEC3A bound to ssDNA substrate (PDB ID code 5ssw) showing key APOBEC3A residues involved in Zn chelation and interactions with the target and neighboring bases (labeled −2, −1, +1, and +2). (B) Surface view of APOBEC3F (PDB: 4j4j) depicting Vif1-binding residues obtained from various studies. Buried residues: red; solvent exposed: blue. (C) Positional entropy values for various AAD clades with mean entropy values (blue horizontal lines) and secondary structures on top. Loop-1 and -7 residues are shown in E: blue dots. Refer to SI Appendix, Fig. S22, for details. (D) Boxplots comparing global entropy values. Mean entropy values of >2, <2, and <1.5 are colored deep-brick, blue, and green, respectively. (E) Sequence logos of key substrate-binding residues in loops 7 and 1. Loop-7 residues are equivalents of APOBEC3A (PDB: 5ssw) Y130, D131, and Y132. Loop-1 residues are equivalents of H29, K30, and T31.

Fig. 4.

Length diversity of substrate-binding loops in AADs. AAD clades are listed on the y axis. For each clade, the range of the loop lengths in amino acids (x axis) is shown. The sequences corresponding to a loop of a particular length are shown as a circle centered at that length, with radius scaled by number of sequences normalized for a clade. The coloring scheme is a rainbow spectrum ranging from red (small number of sequences) to violet (large number of sequences), with the vertical blue lines representing the median loop length.

Fig. 5.

Evolutionary reconstruction and origins of various AAD families. The provenance/distribution of AAD clades is superimposed on a simplified eukaryotic tree. Presence of AADs (green), LSEs of AADs (red circles with outer glow), and absence/potential loss of AADs (yellow markers/markers with cross). Tunicate inactive versions (gray circle); dotted lines indicate alternative lateral transfer scenarios from bacteria to stems of metazoa and dictyosteliids or to stems of ophisthokonta-amoebozoa with loss in fungi (“?”).