NOVEL METHODS OF REPERTOIRE DEVELOPMENT IN THE LYMPHOCYTE ANTIGEN RECEPTORS OF BOS TAURUS AND GINGLYMOSTOMA CIRRATUM

The adaptive immune system of all jawed vertebrates relies on B and T cell effectors bearing immunoglobulin superfamily (IGSF) antigen receptors called B and T cell receptors (BCR, TCR). These antigen receptors are borne from distinct loci requiring genomic rearrangement of variable (V), diversity (D), and joining (J) gene segments, allowing a near infinite repertoire to be encoded within confined loci. Gene rearrangement events, carried out by the recombination activating genes (RAG), massively expands diversity in complementary determine regions (CDRs), loops forming the receptor-ligand interface. IGSF receptors can also be altered by activation induced cytidine deaminase (AID) activity to hone response specificity or to expand the primary repertoire. Bovine BCRs undergo AID mediated somatic hypermutations (SHM) to diversify the naïve repertoire. The Bos taurus antibody repertoire is characterized by a subset of immunoglobulin heavy (IgH) receptors encoding ultralong CDR3 regions greater than forty amino acid residues in length. Ultralong CDR3 BCR rearrangements are genetically constructed with a propensity to yield Cys residues when mutated. The Cys residues are integral in facilitating the unique ‘knob’ domain which protrudes from a β-ribbon stalk. This protruding knob is held far from the other CDRs and forms the primary paratope of the molecule. This miniscule paratope can be further honed by AID mediated deletion events conferring unique structural motifs compared to full length knobs. The TCR repertoire of Ginglymostoma cirratum employs Ig-TCRδ rearrangements that blend gene segments from distinct BCR and TCR loci. These receptors make use of IgH V segments to diversify the existing TCRδ repertoire. Sequencing of the Ig-TCRδ repertoire identified these rearrangements stemmed from a number of IgH mini-cluster loci as well as a distinct lineage of Ig-like V segments located within the TCRδ locus termed TAILV segments. Quantification of Ig-TCRδ transcripts determined they were just as common as canonical TCRδ receptors in this ancient iteration of adaptive immunity, and could be found localized on the cell surface presumptively playing an active role in shark immunity. These novel methods of repertoire development showcase the malleability of the IGSF system in developing a massively diverse repertoire to protect an organism from the antigenic onslaught of life

The Marine Mammal Class II Major Histocompatibility Complex Organization

Sirenians share with cetaceans and pinnipeds several convergent traits selected for the aquatic lifestyle. Living in water poses new challenges not only for locomotion and feeding but also for combating new pathogens, which may render the immune system one of the best tools aquatic mammals have for dealing with aquatic microbial threats. So far, only cetaceans have had their class II Major Histocompatibility Complex (MHC) organization characterized, despite the importance of MHC genes for adaptive immune responses. This study aims to characterize the organization of the marine mammal class II MHC using publicly available genomes. We located class II sequences in the genomes of one sirenian, four pinnipeds and eight cetaceans using NCBI-BLAST and reannotated the sequences using local BLAST search with exon and intron libraries. Scaffolds containing class II sequences were compared using dotplot analysis and introns were used for phylogenetic analysis. The manatee class II region shares overall synteny with other mammals, however most DR loci were translocated from the canonical location, past the extended class II region. Detailed analysis of the genomes of closely related taxa revealed that this presumed translocation is shared with all other living afrotherians. Other presumptive chromosome rearrangements in Afrotheria are the deletion of DQ loci in Afrosoricida and deletion of DP in E. telfairi. Pinnipeds share the main features of dog MHC: lack of a functional pair of DPA/DPB genes and inverted DRB locus between DQ and DO subregions. All cetaceans share the Cetartiodactyla inversion separating class II genes into two subregions: class IIa, with DR and DQ genes, and class IIb, with non-classic genes and a DRB pseudogene. These results point to three distinct and unheralded class II MHC structures in marine mammals: one canonical organization but lacking DP genes in pinnipeds; one bearing an inversion separating IIa and IIb subregions lacking DP genes found in cetaceans; and one with a translocation separating the most diverse class II gene from the MHC found in afrotherians and presumptive functional DR, DQ, and DP genes. Future functional research will reveal how these aquatic mammals cope with pathogen pressures with these divergent MHC organizations