Lymphocyte evolution and ontogeny in non-eutherian mammals

Based on their different reproductive modes and evolutionary history, extant mammals are divided into three lineages: eutherians, marsupials and monotremes. Marsupials are the closest relatives to eutherians. They are born immune-incompetent. The characteristic that they develop their immune system postnatally makes them unique models to study newborn immune development and maternal immunology. As monotremes comprises the basal lineage of mammals, the study of monotreme immunology will provide unique insights regarding the origin of the mammalian immune system. Unfortunately, compared to our in-depth knowledge of eutherian immunology, especially humans and mice, knowledge of marsupial and monotreme immunology is scarce. The goal of the first part of this research was to investigate the development of immune-competence in a model marsupial, the gray short-tailed opossum, Monodelphis domestica. This research specifically looked at the ontogeny of B cells with emphasis on diversity of immunoglobulin genes during postnatal development. It takes advantage of genomic resources available for the opossum and a captive colony at the UNM Biology Department. To achieve these goals, the content and genomic organization of Ig heavy chain and light chain loci was determined. Opossum Ig heavy chain are of low germ-line diversity while light chains have high germ-line diversity. This suggests that opossums rely more on light chains than heavy chains for repertoire diversity (Wang et al. 2009). Using the detailed genomic information available of Ig loci, the timing of B cell ontogeny and Ig repertoire diversity was then determined. Opossum newborns start heavy chain VDJ recombination within the first 24 hours postpartum. The expression of the surrogate L chains occurs at day 6 postnatally. The subsequent rearrangement of the Igλ and Igκ L chain genes occur at days 7 and 8 postnatal, respectively. The diversity of early B cell H chains is limited and reduced in N region additions, as has been seen in fetal humans and mice, but lacks bias in the V, D and J segments used. Different from H chains, L chains develop much more diverse VJ recombinations and high IgL repertoire diversity when first expressed. Collectively the results demonstrate that B cell development is entirely postnatal in the opossum. The earliest time-point that an opossum has mature B cells is at the starting the second week of life (Wang et al. in preparation). These results are consistent with earlier work demonstrating that most marsupial species, including opossums are unable to generate an antibody response until the second week. A second goal of my research was the characterization of a novel T cell receptor in the duckbill platypus Ornithorhynchus anatinus. Although different from the previous goal, it nonetheless uses a non-eutherian model to address broader questions regarding immunoglobulin and T cell receptor gene evolution. TCRμ is a new T cell receptor that was first identified in marsupials and does not exist in eutherians (Parra et al. 2007). Homology searches of the platypus genome with opossum TCRμ sequence have identified a homologue of this unconventional TCRμ in platypus. Platypus TCRμ is expressed in a double V domains structure and these resemble Ig V more than conventional TCR V domains. Different from opossum TCRμ, platypus TCRμ requires two rounds of somatic recombination to assemble both V domains. The identification of TCRμ in platypus indicates that TCRμ as an ancient T cell receptor has been lost in eutherians.PhilosophyDoctoralUniversity of New Mexico. Biology Dept.MILLER, ROBERTLOKER, ERICMILLER, ROBERTAdema, CoenraadCriscitiello, Michae