No description available.

Class I heavy chain (HC) must assemble with β-microglobulin (β2m) and acquire optimal peptide in order to be presented to cytotoxic T cells (CTLs). Calnexin is involved in the initial folding of class I HC and subsequent assembly with β2m. Incorporation of "empty" or suboptimally loaded class I molecules into the multimolecular loading complex is essential for them to acquire optimal peptides. The loading complex consists of several cofactors: TAP, tapasin, ERp57 and calreticulin. The precise role of calnexin and calreticulin in the regulated assembly and peptide loading and the significance of their physical interaction with other cofactors of the loading as well as preloading complex still remains unclear. Using mouse fibroblasts that lack calreticulin, I have studied the role of calreticulin in the assembly and loading of H2-K^b and H2-D^b expressed in these cells. MHC class I molecules in calreticulin-deficient cells are able to assemble with β2m normally, but their subsequent loading with optimal, stabilising peptides is defective despite their ability to interact with the TAP complex. The "empty" or suboptimally loaded class I molecules exit the ER rapidly. Reflecting the loading defect, presentation of endogenously processed antigens by class I molecules in calreticulin-deficient cells is impaired. I have used a human calnexin-deficient cell line CEM.NK^R to study assembly of class I in the absence of calnexin. The results demonstrate that contrary to current understanding, calnexin has an important role in class I HC assembly with 32- microglobulin. The role of heavy chain glycosylation in class I biogenesis is still controversial. My findings suggest asparagine (N)-linked glycosylation of human class I heavy chain at position 86 is optimal and any deviations from "normal" glycosylation results in poor loading with peptides and some defect in the assembly with β2m. Despite affecting the loading function, glycosylation did not have significant effect on presentation of a high affinity binding epitope to HLA-A*0201 specific CTLs. Finally, I show that co-operation from all domains of calreticulin is essential in order to generate a fully functional calreticulin. Interestingly, proline-rich (P) -domain of calreticulin downregulated expression of a number of cellular proteins including MHC class I HC, despite restoring the cytosolic calcium levels in calreticulindeficient cells. The effect of P-domain on class I expression was at the level of transcription.

No description available.

This work examines the ecological, social and genetic factors that predispose amphibians to infection. In the last 30 years many amphibian populations have declined due to infectious disease, although few researchers have studied the factors involved in mediating amphibian infection. My research is designed to explore some of these factors. I first examined the effects of crowding, kin composition (the relatedness of individuals in a group), and habitat complexity on the growth and survival of Xenopus laevis tadpoles exposed to the bacterial pathogen Aeromonas hydrophila. In tadpoles, stress, and in particular corticosterone, a hormone associated with stress, is known to inhibit growth. Crowding, kin composition, and habitat complexity have all been linked to tadpole growth. As corticosterone exposure is also linked to reduced immune function, I examined how these ecological factors influence tadpoles' disease resistance. Tadpoles exposed to the bacterium were significantly smaller and more likely to die than control tadpoles. Tadpoles reared only with siblings (pure sibship groups) were larger, less variable in size, and had lower mortality rates than tadpoles reared in mixed sibship groups. The size difference between pure and mixed sibship groups was greatest when they were exposed to the pathogen. Habitat complexity reduced size variation within tanks but did not affect mean tadpole size. Mixed kinship composition and high tadpole density can increase competition, reduce growth, and increase disease susceptibility. These results indicate that growth was inhibited by pathogen exposure but kin association may ameliorate this effect. The Major Histocompatibility Complex (MHC) is an integral part of the vertebrate adaptive immune system. To determine the importance of the MHC in conferring disease resistance in amphibians, I challenged X. laevis tadpoles, bearing different combinations of four MHC haplotypes (f, g, j, and r), with A. hydrophila in two experiments. Exposure to A. hydrophila affected the growth and survival of these tadpoles and that the MHC moderated these effects. Tadpoles with two MHC haplotypes (r and g) were susceptible to this pathogen and tadpoles with the other two haplotypes (f and j) were resistant. Heterozygous tadpoles with both susceptible and resistant haplotypes were always intermediate to either homozygotes in size and survival. These results demonstrate that MHC genotype plays a major role in determining the impact of bacterial pathogens on the growth and survival of X. laevis tadpoles. To test whether the effect of exposure to pathogens differs according to the similarity of the hosts I challenged tadpoles with natural levels of the microorganisms associated with different MHC genotypes by exposing the tadpoles to water preconditioned by adults of different MHC genotypes. If the pathogens are adapted to the MHC genotype of their hosts, tadpoles exposed to water from adults with which they shared MHC haplotypes would be more susceptible than those exposed to water from MHC-dissimilar adults. Alternatively, if the hosts are adapted to their pathogens tadpoles may be more resistant to pathogens from MHC-similar frogs than those from MHC-dissimilar frogs. I found that tadpoles exposed to water from MHC-dissimilar animals developed faster, but without increased growth, and were more likely to die than those exposed to water from MHC-similar animals. Furthermore, there was an optimal difference between the tadpoles’ and the donors’ MHC where tadpoles were sufficiently different to the donor to defend against its locally adapted pathogens, and sufficiently similar to not be exposed to especially virulent foreign pathogens. Finally, I present an inventory of bacteria found in the gut and skin (nonsystemic sites) and heart, muscle, and abdominal cavity (systemic sites) of captive frogs. I found several species of bacteria previously identified as amphibian pathogens and many bacteria in systemic sites that have not been considered pathogenic to amphibians. None of the frogs tested positive for the amphibian chytrid fungus, Batrachochytrium dendrobatidis. I discuss the potential importance of these species of bacteria as amphibian pathogens and as protective probiotics, using New Zealand frogs as a case study. In its sum, this work describes some of the factors that can affect amphibians’ ability to resist disease. I show that the genetic constitution of an individual, specifically in terms of the MHC, affects the impact of a disease, and so too does its social and ecological conditions, including the level of crowding, the kinship of its groupmates and the specific microbial challenges of its immediate environment. I also show that many of the factors linked to tadpole growth and development that are well described in other amphibians also affect Xenopus tadpoles.

Dissertation (Ph.D.)--Auburn University, 2007. / Abstract. Vita. Includes bibliographic references.

Thesis (M.S.)--Virginia Polytechnic Institute and State University, 1993. / Vita. Abstract. Includes bibliographical references (leaves 54-58). Also available via the Internet.

Thesis (M.S.)--Georgia State University, 2006. / Title from title screen. Susanna Greer, committee chair; Delon Barfus,Yuan Liu, committee members. Electronic text (72 p. : ill.) : digital, PDF file. Description based on contents viewed Aug. 20, 2007. Includes bibliographical references (p. 69-72).

No description available.

The major histocompatibility complex of the rat (RT1 complex) encodes two sets of class II molecules referred to as RT1 B and RT1 D. The RT1 Bα gene was isolated from a Sprague-Dawley (RT1b) rat genomic library using a rat RT1 Bα chain cDNA as a hybridization probe. The coding and the majority of the intron DNA sequence was determined. The structure of the RT1 Bα gene is equivalent to that of H-2 and HLA a chain genes. Comparison of the nucleotide and predicted amino acid sequences of the RT1 Bα gene to those of the H-2 and HLA genes revealed a high degree of overall sequence conservation. However, two regions of the first external domain (a1), residues 19-23 and 45-78, exhibit marked sequence variation. Two blocks of conserved nucleotide sequence were identified in the 5' promoter region of the RT1 Bα gene that have been described in all MHC class II genes sequenced to date. These conserved sequences may be involved in the co-ordinate regulation of expression of class II genes. The cloned RT1 Bα gene was efficiently transcribed when transfected into mouse L cells. / Medicine, Faculty of / Medical Genetics, Department of / Graduate

published_or_final_version / Microbiology / Doctoral / Doctor of Philosophy