Molecular Evolution of Indian Hedgehog Following the Teleost Genome Duplication

Presenter : Ryan Loker

Mentor : Ingo Braasch

Major : Biology

Poster 26

Indian Hedgehog(IHH) is a signaling molecule that acts in several important developmental processes in vertebrates including gut, vascular, and skeletal development. Duplicated(paralogous) copies of the ihh gene have been retained in the teleost fish lineage, ihha and ihhb, following the teleost genome duplication(TGD). The mechanisms of evolution following a whole genome duplication are not completely understood, but one possible reason that ihh and many other duplicates remain in the teleost genomes is
that each paralog carries out essential sub-functions of the ancestral pre-duplication gene, following the Duplication-Degeneration-Complementation(DDC) model. Investigation of this mechanism requires characterization of the regulatory elements controlling expression of ihh genes in teleosts as well as the ancestral state, which can be represented using spotted gar(Lepisosteus oculatus), a member of a sister group to teleosts with an unduplicated genome. Here, we use a comparative genomics approach to compare the ge- nomes of several teleosts and gar in order to identify ancestral conserved non-coding elements(CNE) representing possible regulatory elements, and test their functionality in vivo using transgenic reporter constructs in zebrafish(Danio rerio). In addition to the insight of the evolutionary mechanisms, characterizing enhancers of ihha and ihhb will provide an opportunity to further characterize the role of these genes in developmental pathways, which are commonly conserved throughout vertebrates including humans.

Tracing the Endocardial Cell Lineage of Developing Heart Valves Using MADMMatter Pools?

Presenter : Amy Jones

Mentor : Kryn Stankunas

Major : Biology, Human Physiology

Poster 15

Heart valve development is a complex, multi-step process. During development, endocardial cushions form at specific locations in
the early heart tube. These cushions are populated by endocardial cells that delaminate in a process known as epithelial-mesenchymal transformation (EMT). In the mouse, EMT takes place between embryonic day 9.5-10.5. It is a vital process to understand since the heart valves are derived from this cardiac cushion tissue. Current models of cushion EMT suggest that a large number of endocardial cells undergo EMT. An alternative hypothesis is that only a few individual cells initially populate the cushions that then proliferate to expand cushion mesenchyme.To better understand how endocardial-derived cells contribute to heart valves, I am using a novel mouse genetic system termed MADM (mosaic analysis with double markers). MADM uses the Cre-lox system to permanently label specified cells by fluorescent protein expression. For my studies, I use the Tie2 promoter to direct Cre expression and instruct the MADM system to specifically trace the endocardial cell lineage. The low frequency by which labeled cells are generated allows me to exquisitely moni- tor contributions of clonally-related endocardial cells to developing valves. By gaining a complete understanding of the contribution of endothelial-lineage cells to developing valves, we can identify when embryonic heart malformations originate, supporting the development of therapeutics to prevent defective valves from progressing to a diseased state.

AICAR Administration Promotes a Cytoprotective and Pro-Angiogenic Stimulation in an Ex Vivo Model of Placental Ischemia

Presenter : Sarah Johnson

Mentor : Jeffrey Gilbert

Major : Biology/Human Physiology

Poster 28

The pregnancy-specific hypertension known as preeclampsia (PE) is widely observed worldwide and is recognized as a leading contributor to sickness and death of a mother and her baby. This pervasive condition is yet to be fully characterized, as is an effective therapy of symptom relief outside of inducing early delivery. We have recently reported treatment with an adenosine-mimetic, AICA-riboside (AICAR), in an experimental model of PE reduces the onset of PE-like characteristics; however, the underlying mechanisms are poorly understood. Therefore, our hypothesis was AICAR would initiate cytoprotective and pro-angiogenic stimulation in cultured placental tissue explants, and this effect would be mediated by the adenosine (ENT1/2) transporter. Tissues were cultured at 37°C for 12 hours in physiologic normoxic (8% O2) or hypoxic (1.5% O2) conditions, and treated with AICAR (2mM) and an adenosine transporter blocker (dipyridamole, DPM) (100μM). Explants treated with AICAR exhibited a decreased (P<0.05) secreted sFlt-1 in both O2 conditions, and DPM blocked this effect. The energy regulatory protein AMPKα phosphorylation was elevated in the tissues treated with AICAR, but was not statistically significant (0.1>P>0.05). Through modeling placental ischemia ex vivo, we have demonstrated AICAR decreas- es placental secretion of sFlt-1, mediated by adenosine transport activity. In concert with our previous in vivo studies with AICAR, this study further supports a placental specific mechanism of AICAR’s actions in vivo.

Spatiotemporal Patterns of Inversion Allele Frequencies in Threespine Stickleback

Presenter :Erika Jackson

Mentor : William Cresko

Major : Biology

Poster 21

Chromosomal inversions have been linked to complex traits that facilitate adaptation in new environments in a small number of stud- ies. However, the generality of this pattern is still unclear. Studying the frequency of chromosomal inversions in threespine stickleback (Gasterosteus aculeatus) could provide an important case study to help us understand the role of chromosomal inversions in adaptive evolution. We focused on Alaskan and Oregon marine and freshwater stickleback populations to determine inversion frequencies between distant locations as well as between salinities in different bodies of water. We predicted that inversion allele frequency diver- gence between ocean and freshwater populations would occur if the inverted region contained genes important for adaptation to the alternative environments. We genotyped a large number of individuals for an inversion on Linkage Group XXI using polymerase chain reaction (PCR) designed to indicate alternate forms of the inversion. Our results showed that the inversion allele frequencies are highly divergent between Alaskan oceanic and freshwater populations. In addition, while Alaskan populations are geographically distant from Oregon populations, we found a similar pattern of divergence between Oregon populations in the two habitats. Our study provides ad- ditional evidence that chromosomal inversions may play an important role in adaptation to novel environments.

The Indigenous Siberian Health and Adaptation Project: Seasonal Variation in Autoimmune Thyroid Disorders among the Yakut (Sakha) of Siberia

Presenter : Vimal Balu

Mentor : Josh Snodgrass

Major : Biology, Anthropology

Northern populations physiologically adapt to extreme cold by upregularing basal metabolic rate. The thyroid appears central to this adaptation. Thyroid hormones regulate metabolic responses to chronic cold among indigenous Arctic populations. However, it is un- clear whether this adaptation among circumpolar groups predisposes them to autoimmune thyroid disorders (AITDs). This study ad- dresses this question by examining correlates of seasonal variation in anti-thyroid peroxidase antibody (TPOAb) concentrations among the Yakut of Siberia. Anthropometric and biomarker data were obtained on two occasions (Summer 2009 and Winter 2011) on a sample of Yakut men (n=52) and women (n=88) (≥18 years old). TPOAb levels are higher in summer than winter in both men (P<0.01) and women (P<0.05). Women have significantly higher TPOAb levels than men (P=0.05), and are more likely to have an AITD (28% of women versus 4% of men; TPOAb >30 IU/mL). TPOAb concentrations are associated with several anthropometric dimensions among men (negative trends with weight [P=0.08]). Finally, changes in TPOAb showed a positive trend association with change in thyroid stimulating hormone (TSH; P=0.06) among men, but among women change in TPOAb was negatively associated with change in HDL cholesterol and showed a positive trend with change in triglycerides (P=0.1) and hemoglobin (P=0.08). This study documented impor- tant sex differences in AITD risk among the Yakut, and an unexpected drop in TPOAb levels between summer and winter.

Do Distinct Types of Progenitors Contribute to the Diversity of Enteric Neurons and Glia?

Presenter: Charlotte Taylor

Mentor: Judith Eisen

Oral Presentation

Major: Biology 

The enteric nervous system (ENS), the largest component of the peripheral nervous system, provides intrinsic innervation of the intestinal tract and modulates gut function. The ENS forms a complex network composed of different neuronal and glial subtypes. Whether these different subtypes arise from distinct progenitors is currently unknown. Developing zebrafish embryos are transparent and genetic manipulations can be used to label progenitor cells and their progeny, thus zebrafish is an excellent model in which to address this question. ENS progenitors express several marker genes, including phox2b, sox10, and ret. Using the zebrafish model, we investigated whether expression of these genes designates distinct ENS progenitor populations. Our co-expression analysis identified three different progenitor subpopulations that express the following marker combinations: phox2b/sox10/ret, phox2b/ret, and phox2b. Our next goal is to test the hypothesis that these subpopulations give rise to distinct neuronal and glial cell types during ENS development. We will use the Cre/loxP lineage tracing system to track progeny of identified progenitor subpopulations. Currently, we are generating BAC constructs that drive expression of Cre recombinase under the control of enteric progenitor specific promoters. We will inject these BAC constructs into a red fluorescent reporter line to permanently label all Cre expressing cells and their progeny and then follow the fate of of these cells in living embryos during ENS development. These results will provide a comprehensive lineage analysis of ENS precursors in vivo and thus offer new insights into ENS development and the developmental potential of individual ENS progenitors.

Saturating the Neurospora Genome for Mutants Defective in Methylation

Presenter: Calvin Summers

Mentor: Eric Selker

Poster: 31

Major: Biology

Cytosine methylation, a fundamental form of epigenetic regulation, is found in many eukaryotes and plays a significant role in cancer and other diseases. Using the genetically tractable model organism Neurospora crassa, the Selker laboratory has identified genes that when mutated, cause the strains to be defective in methylation (dim). The process of DNA methylation in Neurospora has been shown to be dependent on DCDC, a five member complex that directs the histone methyltransferase DIM-5 to trimethylate Lysine 9 on histone H3 (H3K9me3). This mark is recognized by HP1, which directs DIM-2 to methylate DNA. Another silencing complex, HCHC, employs HDA-1, CDP-2, HP1, and CHAP to deacetylate lysines on the histone. While we know a good deal about DNA methylation, it is still unclear whether we have identified all genes involved in the process. Thus this research focuses on our search for dim mutants, and asks whether our current model for DNA methylation is saturated. Using a selection for reactivation of resistance genes silenced by DNA methylation we aimed to answer this question. Interestingly, we predominantly identified known dim genes, including dim-5, dim-7, dim-8, dim-9, chap, cdp-2, and hda-1, suggesting our model may be saturated. We localized mutations in these known dim genes by DNA sequencing. The dim mutant collection generated should be a useful resource for further investigation into the roles of these genes and their protein products in DNA methylation. Understanding epigenetic pathways in Neurospora is an auspicious first step in elucidating mechanisms of complex genetic regulation in our own cells.

Effects of the Aquatic Contaminant Perchlorate on Expression of NIS Clade Genes in Divergent Populations of Threespine Stickleback

Presenter: Amanda Redmond

Mentor: William Cresko

Poster: 28

Major: Biology

Perchlorate is a known endocrine disruptor and a wide spread environmental contaminant that causes hypothyroidism in humans. Our previous work demonstrated that perchlorate results in a masculinizing effect in anadromous threespine stickleback (Gasterosteus aculeatus), but it is not known whether ecotypes of stickleback respond differently to this contaminant. Perchlorate has a known effect of suppressing thyroid hormone synthesis by competitively inhibiting the sodium-iodide symporter (slc5a5). The large slc5 gene family is associated with thyroid function and vitamin/mineral transport in all vertebrates. Using bioinformatic approaches we have identified that
there are several closely related genes to slc5a5 in stickleback including slc5a6a, slc5a6b, slc5a8a, and slc5a8b.
We investigate the effect of perchlorate on developmental gene expression in three genetically diverged populations of stickleback from Rabbit Slough (oceanic), Boot Lake (freshwater), and Riverbend (freshwater) ecosystems. We have developed RNA anti-sense probes for the NIS genes that we will be using for gene expression analysis using in situ hybridization to document the spatio-temporal expression of slc5a6a, slc5a6b, slc5a8a, and slc5a8b genes in each ecotype in response to perchlorate exposure. Because evolutionarily diverged populations of stickleback living in different habitats have different osmoregulatory and developmental nutrient needs, we hypothesize that a single chemical that affects solute transporters may have variable developmental effects among populations. In a preliminary study, we have mapped the temporal and spatial distribution of these genes in Rabbit slough (oceanic) fishes, and have found that they are expressed starting at 8-10 days in numerous tissues including the thyroid and the gonad. Our goal next is to identify gene expression patterns of the slc5 genes in the thyroid and gonad at 8, 14, and 30 days post fertilization (dpf) in the diverged populations, and to test for changes in spatial or temporal expression of the genes based on population, perchlorate treatement, and/or an interaction of both. We hypothesize that a gene by environment interaction (G-by-E) will lead to unique patterns of slc5a5 clade gene expression response to perchlorate in each of the three populations. Using cryosections of both control and 100ppm perchlorate treated fish we will determine the expression of these genes at 8, 14, and 30dpf. Our study will be relevant to understanding more about effects of toxin exposure between very genetically divergent populations of fish and other vertebrates, including humans.

The Study on Global AGEing and Adult Health (SAGE): Depression and Body Composition Among Aging Populations

Presenter: William Olson

Mentor: Josh Snodgrass

Poster: 26

Major: Biology

Previous research has documented complex associations between depression and body composition; in some studies, depression increases risk for being underweight, while in other studies it has been linked to obesity. However, the links between depression and body composition remain relatively unexplored among older adults and, additionally, no studies have systematically examined this relationship in non-Western countries. We used Wave 1 data from WHO’s Study on global AGEing and adult health (SAGE), a longitudinal study of nationally representative samples of older adults (>50 years old) in six middle income countries (China(nM=6024 nF=6803 n=12827), Ghana(nM=2237 nF=2050 n=4287), India(nM=3302 nF=3255 n=6557), Mexico(nM=869 nF=1330 n=2199), Russian Federation(nM=81 nF=267 n=348), and South Africa(nM=1551 nF=2103 n=3654)), to examine relationships among body mass index (BMI), waist circumference (WC), and depression (based on a symptom-based algorithm). Results indicate substantial differences in depression prevalence by sex and country, ranging from 1.6% (men in China) to 22.9% (women in Mexico). Variation by sex and country was also evident in prevalence of obesity (from 4.3% among Indian men to 52.7% in South African women) and underweight (0.5% in Mexican women to 40.3% in Indian men). Multiple regression analyses were used with weighted population samples to examine the relationship between body composition measures and depression classification while controlling for key covariates such as age, smoking, drinking, marital status, and income. Among older South African women, depression was positively associated with BMI (P=0.003). Otherwise, the lack of a significant relationship between depression and body composition variables in the individual SAGE countries suggests that depression is not a major driver of body composition among older adults.

Attentional and Neuropsychological Assessments Following Concussion in Adolescents and Young Adults

Presenter: Madison Murray

Mentor: Li-Shan Chou

Poster: 25

Major: Biology

Executive function has been defined as utilizing external stimuli to plan purposeful action and may be a sensitive way to detect concussion-related deficits. The adolescent brain has not reached full maturation and is undergoing rapid development of executive functions, so this age group may be affected by head trauma more than young adults. It was hypothesized that adolescents with concussion would perform worse on executive function tasks than young adults with concussion in relation to matched control groups. Two executive function tests, (Attentional Network Test and Task Switching Test) and one neuropsychological test (ImPACT), were each administered to 44 subjects who sustained a concussion (24 adolescents, 20 young adults) and 44 healthy control subjects matched by sex, height, and weight (24, adolescents, 20 young adults). Tests were administered within 72 hours of injury, and again at 1 week, 2 weeks, 1 month, and 2 months post-injury. Results indicated that conflict resolution ability deficits persisted in the adolescent group compared to their controls throughout the two months of testing (p=0.043). Such differences were not detected between young adult concussion and control groups. Differences between adolescent concussion and control groups on ImPACT variables were detected up to one-month post injury (p<0.05), but not between young adult concussion and control groups. Based on the results, adolescents appear to display greater deficits throughout the two months post-injury than young adults in relation to a control group on measures of attentional and neuropsychological function.