<p>We propose to establish a free-living, fresh water flatworm species from the superphylum Lophotrochozoa, Schmidtea mediterranea, to be a genetic model system. S. mediterranea has been vigorously investigated as a powerful system to study adult stem cells and organ regeneration. Its sexual biotype has also been established as a system to understand the inductive mode of germ cell formation that is broadly shared by a lot of species including mammals. However, little is known about the sexual reproduction and genetics in this organism, which limited the availability of genetic approaches. As the sexual planarian is found scattered but with relative abundance in Sardinia, its natural history also presents us the opportunity to study inbreeding?s effect on genetic variability and species survival. Therefore, to study sexual reproduction and genetic inheritance in S. mediterranea will provide us unique opportunities to understand whole body regeneration, inductive germ cell formation, and inbreeding. In this dissertation, progresses in the establishment of the foundation for genetic analysis in S. mediterranea were presented. Though a simultaneous hermaphrodite, the anatomical and genotyping studies concluded that S. mediterranea cross-fertilize. One worm (line S2) was inbred for 10 generations by taking one progeny from each generation and crossing this individual to its regenerated clones. Whole genome sequencing of four different generations in this inbreeding pedigree revealed ~300Mb of the genome maintained their heterozygosity. Further sequencing analysis of the male and female gametes found these regions had low recombination rates, and maintained as two haplotypes (J-/V- haplotypes). Failure of gametes of the same haplotype to form progeny is unlikely due to embryonic lethality as the arrested embryos were significantly less than hatchlings. Additional analysis of two lines (D5D/D5I) with 90% of these regions homozygous as the J-haplotype suggested failure in fertilization between gametes of the same haplotype. Hence, we propose that haplotype incompatibility is the driving mechanisms to maintain genome heterozygosity in the planarian genome. Understanding of the genetic strategies in S. mediterranea will help the development of genetic approaches to study regeneration and germ cell specification. Our findings also suggest S. mediterranea can be a model system to study the evolution of sex and gamete incompatibility.

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Thesis (Ph. D.)--University of Texas at Austin, 2006. / Vita. Includes bibliographical references.

Branched tubular organs, such as the lung and vascular system fulfill the respiratory needs of most animals. Optimal tissue function relies on the uniform sizes and shapes of the constituting branches in each organ. The Drosophila tracheal airways provide a recognized genetic model system for identification and characterization of tube size regulators. We found that the programmed secretion and assembly of the apical extracellular matrix (ECM) is required for the expansion of the trachea and salivary glands (SG) tubes. We have characterized Vermiform (Verm) and Serpentine (Serp), two chitin-binding proteins with predicted polysaccharide deacetylase domains (ChLDs). Verm and Serp mutants show overelongated tubes, suggesting that luminal ECM modification restricts tracheal tube elongation. The luminal deposition of ChLDs, but not other secreted components, depends on paracellular septate junction integrity (SJs) in the tracheal epithelium. Deletion of the deacetylase domain renders Serp-GFP intracellular, arguing that the deacetylase domain harbors uncharacterized secretion signals. To explore this possibility we transferred the deacetylase domain from Serp to Gasp, another tracheal luminal protein, which requires the Emp24 adaptor for ER exit. The Gasp-Deac-GFP chimera was normally secreted in emp24 mutants indicating that the deacetylase domain contains potential ER-exit signals. To identify such signals we characterized conserved sequence motifs in the Serp deacetylase domain. Mutations of the N-glycosylation sites gradually reduced Serp-GFP luminal deposition suggesting that increased glycosylation enhances apical Serp secretion. By contrast, substitutions in three conserved amino acid stretches completely blocked the ER-exit of Serp-GFP. The mutated proteins were N-glycosylated suggesting that the motifs may be involved in a subsequent protein-folding step or facilitate ER exit through interactions with unidentified specific adaptors. / At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.