Thesis (M.M.S.)--Memorial University of Newfoundland, 2005. / Bibliography: leaves 76-80.

Thesis (M.Sc.)--Memorial University of Newfoundland, 2002. / Restricted until October 2003. Bibliography: leaves 114-129.

Thesis advisor: David Burgess / This doctoral thesis addresses two central topics divided into separate chapters. In Chapter 1: The cortical response to RhoA is regulated during mitosis, experimental findings using sea urchin embryos are presented that demonstrate that the small GTPase RhoA participates in positive signaling for cell division and that this activity is negatively regulated prior to anaphase. In a second series of experiments, myosin phosphatase is shown to be a central negative regulator of myosin activity during the cell cycle through metaphase of mitosis and experimental findings support the conclusion that myosin phosphatase opposes RhoA signaling until anaphase onset. These experiments also reveal that myosin activation alone is insufficient to stimulate cortical contractions during S phase and during metaphase arrest following activation of the spindle checkpoint. In Chapter 2: Annotation of cytoskeletal and motility proteins in the sea urchin genome assembly, as part of a collaborative project, homologs of cytoskeletal genes and gene families were derived and annotated from the sea urchin genome assembly. In addition, phylogenetic analysis of multiple gene families is presented based on these findings. / Thesis (PhD) — Boston College, 2008. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.

Thesis advisor: David R. Burgess / Polarity established by the first cleavages in sea urchin embryos was investigated in this thesis revealing precocious embryonic polarity. Studies of embryonic polarity have focused on protostomes such as <italics>C. elegans</italics>, and those on deuterostomes have focused on later developmental stages. I find asymmetries in the sea urchin membrane cell cortex as early as the first division after fertilization as a result of new membrane addition in the cleavage furrow. Membrane domains and the polarity determinants Par6, aPKC, and Cdc42 are polarized to the apical, or free, cell surface, while the cell-cell contact site remains distinct. Using immunofluorescence, fluorescence recovery after photobleaching (FRAP), and specific inhibitor treatments, myosin filaments were identified as the major regulator of membrane cortex polarity. However, membrane domains and cortical polarity determinants are differentially regulated with respect to blastomere dissociation. These asymmetries are required for proper spindle alignment and cleavage plane determination and are responsible for polarized fluid phase endocytosis. The work in this thesis and future studies addressing the connection between the membrane cortex and myosin filaments has and will lead to a greater understanding of the maintenance of embryonic polarity in cleavage stage sea urchin embryos. / Thesis (PhD) — Boston College, 2009. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.

Thesis (M.A.)--Memorial University of Newfoundland, 2005. / Bibliography: leaves 144-152.

Many organisms that lack vision rely on chemical signals to glean information from their environment. Little is known, however, about the ability of sea urchins to detect and respond to such signals. This lack of understanding is especially surprising given the ecological impact of urchins in their respective communities. Regardless of geography, urchins exert strong top down control of plants, algae, and sedentary invertebrates, and these effects are especially evident when urchins, or urchin predators, are removed from an ecosystem. Facultative omnivorous species such as Lytechinus variegatus may greatly alter the abundances of other invertebrates in seagrass communities by preying on juvenile and adult bivalves as well as gastropod egg masses. These potential food resources, however, are patchily distributed within seagrass beds. To find such resources before other organisms can exploit them may require acute abilities to detect signals emanating from these patches. Experiments performed in this study demonstrated a consistent ability of L. variegatus to detect and orient to chemicals emanating from potential food resources over a distance of 1 m. Unlike what has been found in some other marine organisms, turbulent flow conditions did not negatively affect the ability of L. variegatus to find the source of this chemical cue. In fact, only the slowest flows hindered this ability; the bluff shape of the urchin formed a relatively large boundary layer at slow flows, preventing the delivery of chemical signals to the sensors. The relatively high success rates of L. variegatus in turbulent flows may allow it to effectively forage in areas where other organisms cannot. Thus, turbulence may provide a selective advantage for this animal, based on its comparative ability to detect and respond to signals in its environment.

Echinoids (sea urchins and sand dollars) undergo thyroid hormone (TH) regulated larval development and several species can endogenously synthesize these hormones. Although iodine is the essential component for TH synthesis, nothing is known about iodine uptake mechanisms in echinoids. This thesis primarily aimed to characterize integumental iodine uptake in larvae (echinoplutei) of the purple sea urchin Strongylocentrotus purpuratus. The two mechanisms considered were: iodine uptake via sodium dependent transport utilizing orthologs of vertebrate sodium iodide symporter (NIS) and apical iodide transporter (AIT) versus hydrogen peroxide dependent diffusion (PDD) of iodine. Pharmacology and radioiodine (125I) experiments characterized the effects of various compounds on echinoid iodine uptake. The results demonstrate that purple sea urchin echinoplutei acquire iodine from feeding on microalgae and through integumental transfer. Integumental transfer of iodine is inhibited by cyanide and is temperature dependent. Echinoplutei are not affected by perchlorate exposure demonstrating that NIS/AIT is not involved in S. purpuratus iodine uptake. NIS and AIT are both members of the Sodium Solute Carrier 5 (SSF5) transporter family and phylogenetic analysis of various vertebrate and invertebrate SLC5 members (obtained through BLAST searches) indicated that NIS-like transporters might be a vertebrate synapomorphy. Hydrogen peroxide exposure and oxidative stress induced an increase in iodine influx, whereas reducing agents and peroxidase inhibitors disrupted iodine uptake supporting a PDD-based iodine uptake model. In situ hybridization, immunohistochemistry and real time-PCR analysis demonstrate that the sea urchin dual oxidase 1 (Udx1) (an NADPH oxidase that produced hydrogen peroxide) is expressed throughout all stages of larval development in both S. purpuratus and Lytechinus variegatus specifically occurring in epithelial cells. These findings are the first data to demonstrate Udx1 presence and potential activity outside of fertilization and embryogenesis in echinoplutei. The expression patterns and pharmacological results make Udx1 an attractive candidate for involvement in integumental iodine transfer through PDD. These data provide the first evidence for PDD in an animal. The results also suggest that NIS/AIT may be a vertebrate synapomorphy and PDD of iodine across the integument may be widespread across organisms. Future characterization of iodine uptake mechanism in diverse taxa will address this issue. / Studies were funded by the Natural Sciences and Engineering REsearch Council (NSERC) to A.H. [grant number 400230], Equipment purchased with funding from the Caiadian Foundation for Innovation CFI and NSERC [grant number 400587] to A.H.

Decreases in ocean pH through ocean acidification has shown to have direct negative impacts on the early life stages of the Pacific purple sea urchin, Strongylocentrotus purpuratus. Research has suggested that multiple stressors could exacerbate, cancel, or even alleviate the impacts of ocean acidification on echinoderms. This study assessed the combined effects of changes in pCO2 concentrations (390, 800, 1500 ppm), salinities (28, 31, 34 ppt) and temperatures (12, 15, 18°C) on fertilization and larval development in S. purpuratus. Increased pCO2 was the predominant stressor, with additive and antagonistic effects from temperature changes, and no effect from salinity changes. Stressor combinations significantly decreased the rate of normal larval development by 28 – 68%, whereas fertilization and larval survival were unaffected. The strong impact on normal larval development likely indicates that later development stages could be detrimentally affected and could influence the population dynamics of S. purpuratus.

<p>Sea urchin larvae possess a beautiful, intricately patterned, calcium-carbonate skeleton. Formation of this complex structure results from two independent processes within the developing embryo: specification of the mesenchymal cells that produce the skeletal rods, and patterning inputs from the ectoderm, which secretes signals directing the growth and shape of the skeleton. To understand patterning of the skeleton therefore, the specification events behind these two processes must be understood separately, and then connected in order to understand how ectoderm signaling directs skeletal growth. While the former processes of mesenchyme specification and mineralization are under study elsewhere, the means by which ectodermal cues directing skeletal growth are activated and localized is not known. Using molecular genetics, including gene knock downs and mis-expression, as well as microsurgical manipulations of early cleavage embryos, I show how a previously undescribed territory within the ectoderm, the border ectoderm (BE) is specified with short range signaling inputs. Then, experiments show that the BE provides signals that initiate, and contribute to the propagation of skeletogenesis. From this dataset, and from biological experiments I outline a model for how the BE patterns and contributes to the directed growth of the skeleton. I also discuss challenges to this model that need to be addressed in future research. In principle, the mechanism proposed herein depends on the integration of information from both the primary and secondary embryonic axes. It requires both short-range signaling by Wnt5 from the endoderm to establish the BE fate, and TGFß signaling from the oral and aboral ectoderm which subdivides the BE into four territories. These findings demonstrate that the short-range signaling cascade that subdivides the embryo into first mesoderm and then endoderm also specifies ectodermal fates. Ultimately, this research paves the way for understanding how the larval skeleton is patterned during embryogenesis and may provide a paradigm for understanding other, more complex, developmental problems.</p> / Dissertation

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