Biomedical subjects
Publications and source records attributed to Barry L Williams.
Tsong et al. characterize the role of mating type genes in Candida albicans and identify a new regulator of mating type and several mating type target genes. Comparison with Saccharomyces cerevisiae provides an in-depth view into the evolution of a well-characterized genetic regulatory circuit.
One of the most pervasive challenges in molecular phylogenetics is the incongruence between phylogenies obtained using different data sets, such as individual genes. To systematically investigate the degree of incongruence, and potential methods for resolving it, we screened the genome sequences of eight yeast species and selected 106 widely distributed orthologous genes for phylogenetic analyses, singly and by concatenation. Our results suggest that data sets consisting of single or a small number of concatenated genes have a significant probability of supporting conflicting topologies. By contrast, analyses of the entire data set of concatenated genes yielded a single, fully resolved species tree with maximum support. Comparable results were obtained with a concatenation of a minimum of 20 genes; substantially more genes than commonly used but a small fraction of any genome. These results have important implications for resolving branches of the tree of life.
The production of genetically engineered pigs as xenotransplant donors aims to solve the severe shortage of organs for transplantation in humans. The first barrier to successful xenotransplantation is hyperacute rejection (HAR). HAR is a rapid and massive humoral immune response directed against the pig carbohydrate Galalpha 1,3-Gal epitope, which is synthesized by alpha 1,3-galactosyltransferase (alpha1,3-GT). The Galalpha 1,3-Gal antigen also contributes to subsequent acute vascular rejection events. Genetic modifications of donor pigs transgenic for human complement regulatory proteins or different glycosyltransferases to downregulate Galalpha 1,3-Gal expression have been shown to significantly delay xenograft rejection. However, the complete removal of the Galalpha 1,3-Gal antigen is the most attractive option. In this study, the 5' end of the alpha 1,3-GT gene was efficiently targeted with a nonisogenic DNA construct containing predominantly intron sequences and a Kozak translation initiation site to initiate translation of the neomycin resistance reporter gene. We developed two novel polymerase chain reaction screening methods to detect and confirm the targeted G418-resistant clones. This is the first study to use Southern blot analysis to demonstrate the disruption of the alpha 1,3-GT gene in somatic HT-transgenic pig cells before they were used for nuclear transfer. Transgenic male pigs were produced that possess an alpha 1,3-GT knockout allele and express a randomly inserted human alpha 1,2-fucosylosyltransferase (HT) transgene. The generation of homozygous alpha 1,3-GT knockout pigs with the HT-transgenic background is underway and will be unique. This approach intends to combine the alpha 1,3-GT knockout genotype with a ubiquitously expressed fucosyltransferase transgene producing the universally tolerated H antigen. This approach may prove to be more effective than the null phenotype alone in overcoming HAR and delayed xenograft rejection.
Similar phenotypic changes have evolved independently in many animal taxa. It is unknown whether independent changes involve the same or different developmental and genetic mechanisms. Myriad pigment patterns in the genus Drosophila offer numerous opportunities to address this question. Previous studies identified regulatory and structural genes involved in the development and diversification of pigmentation in selected species. Here, we examine Drosophila americana and Drosophila novamexicana, interfertile species that have evolved dramatic pigmentation differences during the few million years since their divergence. Interspecific genetic analysis was used to investigate the contribution of five specific candidate genes and other genomic regions to phenotypic divergence by testing for associations between molecular markers and pigmentation. At least four distinct genomic regions contributed to pigmentation differences, one of which included the ebony gene. Ebony protein was expressed at higher levels in the more yellow D. novamexicana than the heavily melanized D. americana. Because Ebony promotes yellow pigment formation and suppresses melanization, the expression difference and genetic association suggest that evolution at the ebony locus contributed to pigmentation divergence between D. americana and D. novamexicana. Surprisingly, no genetic association with the yellow locus was detected in this study, and Yellow expression was identical in the two species. Evolution at the yellow locus underlies pigmentation divergence among other Drosophila species; thus, similar pigment patterns have evolved through regulatory changes in different genes in different lineages. These findings bear upon understanding classic models of melanism and mimicry.
Detection of the genetic effects of recent habitat fragmentation in natural populations can be a difficult task, especially for high gene flow species. Previous analyses of mitochondrial DNA data from across the current range of Speyeria idalia indicated that the species exhibited high levels of gene flow among populations, with the exception of an isolated population in the eastern portion of its range. However, some populations are found on isolated habitat patches, which were recently separated from one another by large expanses of uninhabitable terrain, in the form of row crop agriculture. The goal of this study was to compare levels of genetic differentiation and diversity among populations found in relatively continuous habitat to populations in both recently and historically isolated habitat. Four microsatellite loci were used to genotype over 300 individuals from five populations in continuous habitat, five populations in recently fragmented habitat, and one historically isolated population. Results from the historically isolated population were concordant with previous analyses and suggest significant differentiation. Also, microsatellite data were consistent with the genetic effects of habitat fragmentation for the recently isolated populations, in the form of increased differentiation and decreased genetic diversity when compared to nonfragmented populations. These results suggest that given the appropriate control populations, microsatellite markers can be used to detect the effects of recent habitat fragmentation in natural populations, even at a large geographical scale in high gene flow species.
Research in pig-to-primate xenotransplantation aims to solve the increasing shortage of organs for human allotransplantation and develop new cell- and tissue-based therapies. Progress towards its clinical application has been hampered by the presence of xenoreactive natural antibodies that bind to the foreign cell surface and activate complement, causing humoral graft rejection. Genetic engineering of donor cells and animals to express human complement inhibitors such as hCD59 significantly prolonged graft survival. Strategies to decrease the deposition of natural antibodies were also developed. Expression of human alpha1,2-fucosyltransferase (H transferase, HT) in pigs modifies the cell-surface carbohydrate phenotype resulting in reduced Galalpha1,3-Gal expression and decreased antibody binding. We have developed transgenic pigs that coexpress hCD59 and HT in various cells and tissues to address both natural antibody binding and complement activation. Functional studies with peripheral blood mononuclear cells and aortic endothelial cells isolated from the double transgenic pigs showed that coexpression of hCD59 and HT markedly increased their resistance to human serum-mediated lysis. This resistance was greater than with cells transgenic for either hCD59 or HT alone. Moreover, transgene expression was enhanced and protection maintained in pig endothelial cells that were exposed for 24 h to pro-inflammatory cytokines. These studies suggest that engineering donor pigs to express multiple molecules that address different humoral components of xenograft rejection represents an important step toward enhancing xenograft survival and improving the prospect of clinical xenotransplantation.
Some species of parasites occur on a wide range of hosts while others are restricted to one or a few host species. The host specificity of a parasite species is determined, in part, by its ability to disperse between host species. Dispersal limitations can be studied by exploring the genetic structure of parasite populations both within a single species of host and across multiple host species. In this study we examined the genetic structure in the mitochondrial cytochrome oxidase I (COI) gene of two genera of lice (Insecta: Phthiraptera) occurring on multiple sympatric species of doves in southern North and Central America. One genus, Columbicola, is generally less host-specific than the other, Physconelloides. For both genera we identified substantial genetic differentiation between populations of conspecific lice on different host species, generally 10-20% sequence divergence. This level of divergence is in the range of that often observed between species of these two genera. We used nested clade analysis to explore fine scale genetic structure within species of these feather lice. We found that species of Physconelloides exhibited more genetic structure, both among hosts and among geographical localities, than did species of Columbicola. In many cases, single haplotypes within species of Columbicola are distributed on multiple host species. Thus, the population genetic structure of species of Physconelloides reveals evidence of geographical differentiation on top of high host species specificity. Underlying differences in dispersal biology probably explain the differences in population genetic structure that we observed between Columbicola and Physconelloides.