New binding specificities evolve via point mutation in an invertebrate allorecognition gene

Abstract

Many organisms use genetic self-recognition systems to distinguish themselves from conspecifics. In the cnidarian, Hydractinia symbiolongicarpus, self-recognition is partially controlled by allorecognition 2 (Alr2). Alr2 encodes a highly polymorphic transmembrane protein that discriminates self from nonself by binding in trans to other Alr2 proteins with identical or similar sequences. Here, we focused on the N-terminal domain of Alr2, which can determine its binding specificity. We pair ancestral sequence reconstruction and experimental assays to show that amino acid substitutions can create sequences with novel binding specificities either directly (via one mutation) or via sequential mutations and intermediates with relaxed specificities. We also show that one side of the domain has experienced positive selection and likely forms the binding interface. Our results provide direct evidence that point mutations can generate Alr2 proteins with novel binding specificities. This provides a plausible mechanism for the generation and maintenance of functional variation in nature.

A family of unusual immunoglobulin superfamily genes in an invertebrate histocompatibility complex

“…In vitro studies have shown that the Alr1 protein is capable of trans (cell-to-cell) homophilic binding, which only occurs between allelic variants with similar extracellular sequences ( 17 ). The same is true for Alr2 ( 17 , 18 ). This variant-specific homophilic binding is hypothesized to be the mechanism of self/nonself discrimination in vivo.…”

“…These domains appear to have followed different evolutionary trajectories to arrive at their current sequences. As in vertebrates, the Hydractinia V-set domain plays a critical role in self/nonself recognition ( 17 , 18 ). However, it remains unclear whether this reflects a conserved, ancient function or is an example of convergent evolution.…”

Multiple Alr genes exhibit allorecognition-associated variation in the colonial cnidarian Hydractinia

“…Remarkably, haplotypes with two or three alr2 pseudogenes were identified, indicating haplotypic variation in this genomic region ). Recently, it has been shown that functional diversity of the alr2 gene is generated through a mechanism of point mutations, by which successive single nucleotide changes create new alr-binding specificities, with the feature that intermediate allele stages have broader alr-binding specificities (Huene et al 2021).…”

A family of unusual immunoglobulin superfamily genes in an invertebrate histocompatibility complex

“…In vitro studies have shown that the Alr1 protein is capable of trans (cell-to-cell) homophilic binding, which only occurs between allelic variants with similar extracellular sequences (17). The same is true for Alr2 (17,18). This variant-specific homophilic binding is hypothesized to be the mechanism of self/non-self discrimination in vivo.…”