The biophysical and structural basis of allorecognition specificity

How do alr1 and alr2, by themselves, distinguish self from non-self? The answer lies in discovering the ligand(s) of the highly polymorphic extracellular domains of alr1 and alr2. We hypothesize that alr1 and alr2 function by allele-specific homophilic binding. This hypothesis is based on the observation that allelic variation at alr1 and alr2 is sufficient to predict allorecognition phenotypes in laboratory strains and no other allodeterminants are present in the genomes of these animals. We are currently testing the homophilic binding hypothesis using a combination of in vitro and in vivo methods, the latter relying on recently established techniques for the production of transgenic Hydractinia.

Discovery of allele-specific binding would be truly remarkable. Natural populations of Hydractinia maintain hundreds of unique alleles at alr1 and alr2, resulting in natural fusion rates of 0-2% between randomly selected colonies. How natural selection could create hundreds of unique specificities within a single locus will be a puzzle for population geneticists, molecular evolutionists, and structural biologists alike.

The allorecognition signaling pathway

In rejection, tissues contacting another colony become swollen with nematocysts (the stinging organelles characteristic of cnidarians) and differentiate into specialized fighting stolons that grow over their opponent. In fusion, these same tissues adhere and reorganize themselves to create a continuous epidermis and gastrovascular system, all within minutes. How are these disparate responses regulated? To answer this question, one must first define the allorecognition signal transduction pathway. We hypothesize that the cytoplasmic tails of alr1 and alr2 initiate allorecognition responses because they are relatively large and bear a number of potential phosphorylation sites. Our current work focuses on determining whether phosphorlyation of alr1 and alr2 does, indeed, regulate allorecognition. We are also working to identify identify the intracellular binding partners.

Characterizing the Hydractinia allorecognition gene complex

alr1 and alr2 predict allorecognition phenotypes in inbred strains of Hydractinia, but wild-type animals sometimes display unexpected phenotypes. This raises the question: Are there additional allorecognition loci in Hydractinia? Systematic attempts to identify unlinked modifiers of allorecognition have revealed none. However, the allorecognition complex is incompletely sequenced and alr1 is part of a multi-gene family of structurally similar genes, the extent of which is unknown. It is possible that these genes—or others in the complex—have been homogenized in our laboratory strains but remain variable in wild-type animals, giving rise to unexpected phenotypes. Our goal is to generate a reference sequence of the allorecognition complex and subsequently sequence a selection of wild-type haplotypes in order to identify all linked allorecognition loci.