A case of convergence: why did a simple alternative to canonical antibodies arise in sharks and camels?

Abstract

The success of our adaptive humoral immune response, mediated by B cells, relies on its capacity to produce multiple antigen-recognition specificities against any possible invader. This antigen-recognition specificity is attributed to antibodies, which display remarkable flexibility and diversity. That diversity is generated by cutting and pasting of immunoglobulin (Ig) gene segments during B-cell development to generate functional variable (V) genes, which are selected from two separate regions of the genome to yield so-called “heavy” (H) and “light” (L) chains (Figure 1; see Section 1 in Text S1). Ig H and L chain V genes have been expanded in various vertebrate taxa to yield multiple families and subfamilies [1]. Given the obvious advantage of the diversity afforded by such multichain antibodies, it was extremely surprising to discover that some species—notably cartilaginous fish (holocephalins, e.g., chimeras, and elasmobranchs, e.g., sharks, skates, and rays) and camelids (e.g., camels and llamas)—developed functional H-chain-only antibodies (HCAbs) (Figures 1 and ​and2)2) [2],[3]. Loss of the combinatorial H-L diversification of heterotetrameric (H2L2) antibodies should result in a handicapped, less diversified antigen-binding repertoire in HCAbs; yet, contrary to expectations, high-affinity, antigen-reactive HCAbs can be elicited to all studied antigens [4],[5]. Even more peculiar, the HCAbs function alongside conventional H2L2 antibodies. What explains this remarkable evolutionary convergence of functional HCAbs in sharks and camels? What forces drove such distantly related species to produce antibodies with this simple structure? And, if they are so effective why is their prevalence not more widespread in evolution? Figure 1 Schematic of classical H2L2 and H2 homodimeric HCAbs. Figure 2 Simplified vertebrate phylogenetic tree emphasizing those taxa possessing single-domain Igs or TCRs. Occurrence of Functional Heavy Chain-Only Antibodies in Camelids and Shark The antigen-binding fragment of camelid HCAbs consists of a single domain known as a variable domain of the heavy chain of HCAbs (VHH) (Figure 1) [2]. It was demonstrated that HCAbs from dromedaries (one-humped camels) infected with trypanosomes are capable of associating tightly and specifically with these parasite antigens [2]. Subsequently, HCAbs were elicited to many different foreign antigens. HCAbs are present in all living species of the mammalian family Camelidae, but not in other ungulates (hoofed mammals). Approximately equal levels of HCAbs and H2L2 antibodies are present in camel blood [6]. The origin of camelid Ig genes encoding HCAbs is unambiguous. Phylogenetic analyses have demonstrated that the HCAb-dedicated γ genes were derived from γ genes coding for conventional antibodies. The genes emerged and diverged ∼25 million years ago [6],[7], after Tylopoda split from other mammals (60–80 million years ago) and before the camel and llama speciation (∼11 million years ago) (Figure 2) [6]. Also, the VHH germline genes descended from the classical variable region of a heavy-chain (VH) genes, and the diversity (D) and JH genes are even shared in the formation of the VH or VHH domains. Thus, the emergence of HCAbs was a relatively recent event in these species. In the nurse shark (Ginglymostoma cirratum), a novel secreted antigen receptor was reported and named the new antigen receptor (NAR) [3]. It too is composed of two covalently associated H-chains with no associated L-chains, but having six domains (Figure 1). Since the molecule shares several functional features with Ig isotypes, it was renamed IgNAR [8], and its N-terminal V domain is known as V-NAR. Later studies showed IgNAR to be induced by immunization with protein antigens with similar kinetics as in a typical IgM response [9]. Although the origin of shark IgNAR is poorly understood, IgNAR is found in all elasmobranchs and thus emerged at least 220 million years ago (Figure 2). However, the single-domain V is also found as a T-cell receptor (TCR) in all living cartilaginous fish (so-called NAR-TCR) [10],[11], showing that this immune system feature appeared at least 350 million years ago (Figure 2). In addition, V-NAR gene clusters actually are present in the TCR-δ loci of all cartilaginous fish and thus it remains uncertain whether they originated as a TCR, an Ig, or perhaps a common antigen receptor ancestor [10].