Master of Science (MS), Wright State University, 2007, Biological Sciences

Reproductive isolation results when members of the same species cannot reproduce due to either prezygotic or postzygotic mechanisms, therefore restricting gene flow between populations. A leading model describing reproductive isolation developed by J.B. Haldane (1922) states, “When offspring from two different animal species have one sex that is rare, absent, or sterile, that sex is the heterozygous (XO) or heterogametic (XY) sex”. Haldane's rule is illustrated among various animal taxa regardless of which sex is heterogametic. To date, Caenorhabditis mating tests are the only example of Haldane's rule that is caused by sexual transformation and not gender specific lethality or sterility. Crosses between C. briggsae strain AF16 males and C. remanei strain EM464 females resulted in all female F1 hybrids. The F1 hybrids were all phenotypically female even though some hybrids were genetically XO (Baird et al., 1992, Baird, 2002). The absence of male hybrids resulted from sex reversal rather than male specific lethality. This suggested that perhaps dysgenic interactions within the sex determination pathway of Caenorhabditis may serve as the mechanism for reproductive isolation. Utilizing the previously identified C. briggsae tra-2 gene as a candidate gene for sexual transformation, haplotype crosses were set up between C. remanei males and C. briggsae females from temperate and tropical strains. Results indicated that the sex reversal phenotype did not map to both the tropical and temperate clades. AF16, a tropical strain, was the only strain that exhibited the sex reversal phenotype. The tra-2 gene was also analyzed for selection pressure by comparing nonsynonymous and synonymous substitutions in the available C. briggsae tra-2 sequences. Data showed no evidence of natural selection acting on the gene region, indicating that perhaps the selective pressures have relaxed since the time of speciation.

Committee: Scott Baird (Advisor) Subjects: Biology, Genetics

Master of Science (MS), Wright State University, 2007, Biological Sciences

Sharma, Monita. M.S., Department of Biological Sciences, Wright State University, 2007. Molecular Characterization of Chironomid species and their use as bio-indicators. Of all major aquatic invertebrate groups, members of family Chironomidae are most abundant and show a wide range of habitat preferences. The importance of correct identification of Chironomids has been realized in many bioassessment studies mainly because of their worldwide distribution, substrate specificities and predictable responses to various pollutants in the water sources. This study establishes that the sequence data from the Intergenic Spacer Regions (ITS) of ribosomal DNA could be used as molecular markers to distinguish between different Chironomidae species and also to identify them. The need to use molecular approaches, to identify various Chironomidae species, comes from the fact that the rate of misidentifications is fairly high when morphological features are used. A difference of six nucleotides in the sequence data of Chironomus tentans from North America and Europe suggest a low intraspecific variation. A detailed analysis of the ITS-1 and ITS-2 sequence data from seven new species of Chironomids (Thienemanniella xena, Xylatopus par, Tribelos fuscicorne, Robackia demeijerei, Tribelos jucundum, Polypedilum aviceps and Chironomus tentans) along with 15 species obtained from Genbank considered in this study shows a high amount of interspecific variations and also that the European species tend to cluster close to each other when compared to North American ones. The high bootstrap values and short intercluster branches, depicted in the phylogram, might suggest presence of various clusters and rapid divergence of species, respectively within the genus Chironomus. Such phylogenetic analysis could also provide more information on the genetic relatedness among different species.

Committee: Dan Krane (Advisor) Subjects: Biology, Genetics

Master of Science (MS), Wright State University, 2006, Biological Sciences

Hampton, Rachael M. M.S., Department of Biological Sciences, Wright State University, 2006. Analysis of clade structure and gene flow in Caenorhabditis briggsae. In this study multilocus DNA sequence variation was examined in geographic isolates of C. briggsae to determine clade structure and to test for evidence of gene flow between clades. Sequence data were obtained for a total of seventeen genes scattered throughout the genome for five C. briggsae strains. Phylogenetic reconstructions at thirteen of seventeen loci strongly supported the presence of two separate clades with AF16 and VT847 comprising clade 1 and HK104, HK105, and PB800 comprising clade 2. The remaining four loci were not informative for generating this distinction due to a lack of polymorphism in DNA sequence. Comparisons were also extended to include eleven additional strains for nine of these loci. Clade structure was also strongly supported by phylogenetic reconstructions at seven of these loci. Clade 1 comprised strains AF16, VT847, PS1185, and PS1186. Clade 2 comprised strains HK104, HK105, PB800, PB826, PB857, PB858, PB859, JU279, JU348, JU383, JU405, JU439, and JU441.The lack of support for clade structure at Cb-her-1 and Cb-glp-1 was likely due to gene flow. At Cb-her-1 shared polymorphisms were observed indicating the presence of gene flow. Thus, it appears that outcrossing does occur to some extent between the C. briggsae clades. To assist in identifying the genes contributing to reduce fitness in AF16::HK104 F2 hybrids, sequenced loci were also tested for cosegregation with developmental delay through bulk segregant analysis. Multiple pools of delayed AF16::HK104 F2 hybrids were collected then genotyped at thirteen loci. Of those tested, eleven consistently showed random segregation of both alleles. Genes Cb-egl-5 and Cb-glp-1 showed nonrandom segregation and both loci were skewed towards AF16. This indicated that the region linked to Cb-egl-5 and Cb-glp-1 on chromosome three is associa (open full item for complete abstract)

Committee: Scott Baird (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2001, Allied Health Sciences : Genetic Counseling

PURPOSE: To assess the impact of race upon coping strategies and to identify support resources available to low-income, urban African American and Caucasian parents of children with developmental disabilities.METHODS: Conducted telephone interviews with 43 parents of children with developmental disabilities who were seen in genetics clinic. RESULTS: Of the 15 coping mechanisms assessed, Caucasians reported significantly greater use of only two, Active coping (p=.0476) and Acceptance (p=.0500). Social support utilization was similar for all, with both groups reporting family support almost daily. Formal supports such as health professionals were utilized less often, despite their availability. CONCLUSIONS: Coping strategies and support resources are not affected by race in this community. Use of formal support resources may be facilitated by genetics professionals.

Committee: Leah Hoechstetter (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2001, Allied Health Sciences : Genetic Counseling

Background: Breast cancer occurs at higher rates in women over the age of 55. Young-adult women, however, often do not realize they may also be at increased risk if they have a strong family history of breast cancers. Breast cancer genetics education, when incorporated into a general education program about breast cancer, would inform college-age women about hereditary breast cancer risks and options for risk reduction, to ultimately promote early detection or prevention of breast cancer. The aim of this study was to identify the optimal breast cancer genetics education method for college-age women. Methods: A convenience sample of 35 college women was recruited for focus group inclusion. Participants completed demographics and knowledge assessment questionnaires and viewed portions of four common breast cancer genetics education tools (CD-ROM, video, brochure, and lecture). During the focus groups, women discussed positive and negative aspects of each tool, ideas for disseminating the information, and the importance of the topic. Results: Focus group participants preferred tools which were convenient, self-paced, personal, and reflective of real situations. Aspects considered negative included the inability to ask questions or repeat information and situations not relevant to the life of a college female. Participants ranked the CD-ROM as most preferred, followed by the lecture, the brochure, and the video. They suggested combining tools and widely disseminating breast cancer information in conjunction with other health-related events. Participants expressed that global breast cancer education is important, but only women with a family history are likely to seek information about inherited breast cancer. Conclusion: In order to be received by college populations, breast cancer and genetics information must be delivered in a relevant way. An educational intervention based on these findings might be comprised of a general breast cancer lecture delivered by a young br (open full item for complete abstract)

Committee: Dr. Margaret Miller (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2001, Allied Health Sciences : Genetic Counseling

Friendships are an important component of community integration. As young adults with Down syndrome become more active in the community, their social relationships gain importance. To assess these relationships, structured interviews were conducted with 14 young adults with Down syndrome, between the ages of 18 and 30. The participants named an average of 3.5 friends, most of whom had disabilities. The most frequent places where individuals met their friends were at school, sporting activities, and social clubs. These were also the most common places young adults saw their friends. When asked to define friendship, typical responses included "going out together" and "funny." Definitions of boy/girlfriend relationships were similar to those of friendship. In both instances, the definitions failed to express the emotional depth expected from these types of relationships. Ten of the 14 young adults in this study reported having a boyfriend or girlfriend. Of the 14, 11 want to get married and 9 would like to become parents. However, none of the adults were married or had children. The young adults in this study rely on their parents, rather than their peers, for emotional support. In addition, the young adults depend on their parents for housing and transportation. Twelve of the participants lived with their parents and often participate in social activities with their families. This high level of parental involvement may prevent increased independence, which may, in turn, cause problems as parents age. Young adults need to understand that peer relationships can satisfy emotional needs. Parents and the community can work together to teach adults social skills and model reciprocal relationships. If individuals can maintain friendships long term, they may have a stronger support network when they transition away from parents.

Committee: Carol Christianson (Advisor) Subjects: Biology, Genetics

PhD, University of Cincinnati, 2001, Medicine : Molecular Genetics, Biochemistry and Microbiology

Juvenile rheumatoid arthritis is the most common childhood rheumatic disease. Previous studies have shown that null alleles of the T cell receptor Vβ (TCRBV) gene segment TCRBV6S1 are associated with some subsets of JRA, namely polyarticular JRA. Linkage disequilibrium has been identified between the null alleles of TCRBV6S1 and alleles of the neighboring TCRBV genes TCRBV5S5P, TCRBV1S1 and TCRBV13S5, and this genetic association may therefore be due to a linked susceptibility locus. In Caucasian individuals, only three of the possible 12 haplotypes were represented, and the two TCRBV6S1 null alleles are present on distinct haplotypes. However, in additional ethnic groups significant deviations in the patterns of linkage disequilibrium were observed. This needs to be taken into account in any genetic studies involving TCR polymorphisms. TCRBV repertoire analysis was undertaken in an effort to establish a functional role for TCRBV6S1 null allele haplotypes in the pathogenesis of JRA. TCRBV1S1 is the only member of this haplotype that is readily expressed in the peripheral repertoire; analysis of synovial and peripheral blood TCRBV1S1 repertoires revealed an allelic bias wherein TCRBV1S1A2 is under-represented in heterozygotes. Additionally, individuals who are carriers of TCRBV1S1A2 have lower levels of TCRBV1S1 mRNA and reduced numbers of Vβ1 + T cells. Although we have not definitively established a role for TCRBV1S1A2 in JRA, reduced numbers of Vβ1 + T cells may nonetheless be relevant to disease pathogenesis. The transmission disequilibrium test (TDT) was used to test whether TCRBV6S1 null alleles are preferentially transmitted to JRA probands. Unexpectedly, the TDT demonstrates under-transmission of TCRBV6S1 null alleles to polyarticular JRA patients, suggesting these alleles may be protective. In fact, the two null alleles do not appear to contribute equally to disease protection in all JRA subtypes. Although this data is preliminary, it suggests that caution m (open full item for complete abstract)

Committee: Edmund Choi (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2001, Allied Health Sciences : Genetic Counseling

Purpose: It seems quite feasible that adult long term survivors of childhood cancer can have numerous medical concerns when considering a pregnancy. The aims of this study were to determine whether survivors of childhood cancer, who attend a long-term survivors pediatric oncology clinic, (1) recall their diagnosis accurately, and (2) have reproductive concerns that differ from reproductive concerns of their friends. Patients and Methods: Subjects were all individuals over the age of 18 years who attended a local long term survivors oncology clinic. They were interviewed about reproductive concerns and were asked to contact a friend of similar age and gender to serve as a control. Controls were also interviewed and asked similar questions. Results: Of the 25 long-term survivors of childhood cancer interviewed (mean age 26.3 years; 17 males, 8 females), 24 accurately recalled their diagnoses. The comparison of cancer survivors with their friends (study controls) showed no statistically significant differences in worries regarding reproductive issues. Furthermore, there was no statistically significant difference in perceived risk of future cancer in themselves or of cancer in their offspring when responses from the two groups were compared. Discussion: The results of this study suggest that cancer survivors who attend annual multidisciplinary follow-up clinics have concerns and worries about reproductive issues similar to those of their peers. These findings highlight the need to include an appropriate comparison group when examining these issues for long term survivors.

Committee: Dr. Robert Noll (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2001, Allied Health Sciences : Genetic Counseling

Predictive genetic tests for complex genetic diseases will lead to earlier diagnosis and improved prevention strategies. Personalized genetic risk information could therefore be of considerable benefit to young adults, even though genetic tests for complex diseases may only provide an estimation of disease risk rather than a precise diagnosis. This study assessed interest in genetic testing for a complex genetic disease, osteoporosis, and a single-gene disorder, hemochromatosis, in a group of college women. these diseases are both late-onset and preventable, but genetic tests for these conditions differ in their potential predictive abilities. We hypothesized that interest in a genetic test for hemochromatosis would be higher than interest in a theoretical test for osteoporosis susceptibility, given the higher predictive value of the hemochromatosis test, and that health behaviors would predict interest in testing. Participants were 181 undergraduate women recruited through introductory psychology classes at the University of Cincinnati. They were randomly assigned to receive a questionnaire containing either osteoporosis or hemochromatosis. After assessment of prior disease knowledge and health behaviors, the clinical features of each disease and the limits of a genetic test for each were described. Interest in genetic testing and disease specific health beliefs were then assessed. We compared interest in testing between the osteoporosis and hemochromatosis groups and analyzed variables for association with genetic test acceptance. Sixty-three percent of the total population was interested in genetic testing. There was a trend toward higher interest in the osteoporosis group (68%) than in the hemochromatosis (58%), though no significant difference was found. Significant predictors of test acceptance were disease familiarity, perceived disease severity, perceived risk, and perceived benefits of testing. Health behaviors were not related to interest in testing. These (open full item for complete abstract)

Committee: Dr. Richard Wenstrup (Advisor) Subjects: Biology, Genetics

PhD, University of Cincinnati, 2007, Medicine : Environmental Health

Association studies offer great promise in dissecting the genetic basic of human complex diseases. The rapid expansion of genomic information and the cost-effective genotyping technologies have enabled us to systematically interrogate the role of human genetic variation in common diseases by genome-wide association (GWA) mapping. However, the scale and complexity of such studies will raise significant challenges in study design and data analysis. In this dissertation, we investigated several statistical problems that relevant to population-based association studies and the fine-scale mapping of genetic variants that influence susceptibility to complex diseases. First, we developed a variance-based effect size estimator for the locus-specific genetic effect. Comparing to the traditional measures, the proposed estimator is less sensitive to the risk allele frequency and the population prevalence of the disease. We demonstrated the sample size requirement would be considerable large to obtain an accurate estimate on moderate genetic effect and the sample size will increase exponentially with increased demand for precision. We next compared the power of different association test statistics. We observed that the genotype based single-locus tests is generally more powerful than the multi-locus or haplotype based statistics, especially for risk alleles far from additive; and the power of genotype based tests can be uniformly improved by applying the ordered restriction on genotypic risks. Finally, we tested different GWA strategies and explored the factors that may influence the power of GWA studies by extensive simulations using empirical genotype data from the HapMap ENCODE Project. Our results indicate that current commercial genome-wide typing products are capable of capturing most of the common risk variants; however, their power in detecting rare risk variants or variants within recombination hot spots is not satisfactory. We also showed that the properties of the r (open full item for complete abstract)

Committee: Ranajit Chakraborty (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2007, Allied Health Sciences : Genetic Counseling

Autosomal dominant nonsyndromic hearing loss (ADNSHL) is characterized by postlingual, progressive hearing impairment. This study sought to identify the gene responsible for hereditary nonsyndromic sensorineural hearing loss in a family with multiple generations affected by hearing impairment presenting in the second decade of life. The Affymetrix GeneChip was used to identify three linkage intervals on chromosomes 4, 10, and 16. The observed hearing loss in this family is not likely due to previously identified deafness-causing genes as no such genes have been reported in the identified intervals. Since preliminary candidate gene sequencing within the regions did not identify any pathogenic mutations, haplotype mapping was employed to further refine the intervals. The intervals on chromosomes 4 and 16 were excluded and the interval on chromosome 10 was narrowed to a 0.4Mb region at 10q22-q23. Future work will employ candidate gene analysis to identify the gene responsible for this family's hearing impairment.

Committee: Dr. John Greinwald (Advisor) Subjects: Biology, Genetics

PhD, University of Cincinnati, 2006, Medicine : Molecular Genetics, Biochemistry, and Microbiology

The morphogenetic Hedgehog (Hh) signal transduction pathway is evolutionarily conserved and required for proper animal growth, maintenance and development. Disruption of the Hh pathway in humans leads to a variety of disease states that range from developmental disorders to cancer in children and adults. Its importance to human health is exemplified by the growing number of cancers shown to have elevated Hh signaling activity. As a means of developing and identifying targets for therapeutic intervention of aberrant Hh signaling, I investigated the fundamental mechanisms underlying Hh signal transduction. To this end, I studied the Hh pathway in the fruit fly Drosophila melanogaster, taking advantage of the rich genetic and biochemical tools available for this model organism. A large multi-protein cytosolic Hh signaling complex (HSC) functions to properly interpret the Hh signal. The Ser/Thr protein kinase Fused (Fu) is an integral member of the HSC. Fu appears necessary to ensure that the HSC remain in an ‘off” state in the absence of Hh, but is also required for full HSC activity in the presence of Hh. Here I describe work characterizing the role of Fu in HSC function. I found that Fu is engaged in a number of protein interactions with the other components of the HSC, including with itself. Fu is directly bound to the kinesin-related protein Costal2. Targeted disruption of their interaction leads to a total loss of Hh activation. Furthermore, Fu appears capable of forming self-associated monomers. By promoting self-association in vivo, we are able to rescue a fu loss of function phenotype in Drosophila. Finally we discover that the HSC is in direct contact with Smoothened, the G-protein coupled receptor-like transmembrane protein. By examining the consequences of identifying and disrupting protein complex interactions, we are now able to derive a mechanistic model of Hh signal transduction.

Committee: David Robbins (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2006, Allied Health Sciences : Genetic Counseling

Currently, there are an estimated six hundred genetic advocacy / support organizations in existence throughout the United States and abroad. Groups consist of individuals and family members affected by a genetic condition. Genetic counselors are in a pivotal position for referring to and educating patients about groups, as well as collaborating professionally. Collaborations between counselors and groups remain largely anecdotal today, despite years of nationally recognized importance. Utilizing an internet-based instrument, we surveyed 141 genetic counselors in this crosssectional analysis to identify genetic counselors' 1) experience with; 2) understanding of roles and functions; 3) ways that counselors currently collaborate with groups; and 4) recommendations for and/ or barriers to future professional collaborations with groups. Novel roles, self-identified responsibilities and definitions of advocacy and support groups were identified. In comparing the area of counseling with collaboration, cancer counselors were found to participate with groups more than prenatal counselors.

Committee: Nancy Warren (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2004, Medicine : Molecular Genetics, Biochemistry, and Microbiology

Stroke is the third leading cause of death after cardiovascular disease and cancer. Stroke is the leading cause of disability in industrialized nations. Spontaneous intracerebral hemorrhage (ICH) accounts for approximately 15% of all cases of stroke and is the subtype with the highest mortality rate (40-50%). One of the major unanswered questions is whether ICH has a genetic component and if so, what alleles contribute to susceptibility. The study of ICH is complex because different pathophysiologic mechanisms can lead to ICH. Through a population-based case ascertainment method, we have recruited a subset of cases of ICH for direct interview and genetic sampling and matched them to two population-based controls by age, race and gender. In our study of this cohort of subjects, we have found that the risk factors for ICH vary by location with hypertension being of greater importance among non-lobar ICH cases and Apolipoprotein E4 alleles being of greater importance among lobar ICH cases. We have further discovered that apolipoprotein E4 alleles are a significant risk factor for lobar ICH in individuals older than 70 years of age but does not appear to be a risk factor for individuals less than 70 years of age. This is consistent with its proposed influence through cerebral amyloid angiopathy which occurs only rarely in persons less than 70 years of age. We found that Apolipoprotein E4 is associated with lobar ICH and that haplotypes which included polymorphisms of the 5' untranslated region were associated with lobar ICH as well. In further analysis, we explored the importance of genes related to hypertension to ICH and found that polymorphisms of the endothelial nitric oxide synthase gene, which is the primary regulator of cerebrovascular tone, as well as of amiloride sensitive epithelial sodium channel genes are associated with non-lobar ICH but the associations were not as strong as that seen for Apo E4. These findings should be considered in the evaluation of the (open full item for complete abstract)

Committee: Dr. Anil Menon (Advisor) Subjects: Biology, Genetics

MS, University of Cincinnati, 2002, Allied Health Sciences : Genetic Counseling

Neurofibromatosis type 1 (NF1) is an autosomal dominant condition whose features include a number of cutaneous, systemic, cognitive, and behavioral problems. The degree to which NF1 impacts an adolescent's daily life depends on the clinical features of the condition as well as on social and emotional influences, including their perception of the condition's severity. The purpose of this study was to examine the relationship between adolescent and parental perception of disease severity and the clinical severity of NF1. The Perception of Severity of Chronic Illness (PSCI) questionnaire was administered to 56 parents and 47 adolescents with NF1. Scores assessing the clinical severity of the adolescent's condition were assigned. Correlation coefficients were used to evaluate the relationship between the clinical features of NF1 and the families' perceptions. Parental perceptions were significantly correlated with the degree of systemic (r = 0.3116, p<0.05), cognitive (r = 0.4911, p<0.0001), and behavioral (r = 0.3341, p<0.05) impairment of the adolescent. Adolescent perception was correlated only with the degree of cognitive impairment (r = 0.5429, p<0.0001). Parental and adolescent perceptions were closely correlated (r = 0.6724, p<0.0001); however, adolescents consistently viewed the condition's impact as being less than the parents' perceptions. The results of this study have implications for the clinical care and counseling of families of adolescents with NF1.

Committee: Dr. Elizabeth Schorry (Advisor) Subjects: Biology, Genetics

Master of Science, University of Toledo, 2007, Biology (Ecology)

DNA sequencing has led to the resolution of many cryptic taxa, which are especially prevalent in the North American darter fishes (Family Percidae). The Greenside Darter Etheostoma blennioidescommonly occurs in the lower Great Lakes region, where two putative subspecies, the eastern “Allegheny” type E. b. blennioidesand the western “Prairie” type E. b. pholidotum, overlap. The objective of this study was to test the systematic identity and genetic divergence distinguishing the two subspecies in areas of sympatry and allopatry in comparison to other subspecies and close relatives. DNA sequences from the mtDNA cytochrome bgene and control region and the nuclear S7 intron 1 comprising a total of 1,497 bp were compared from 294 individuals across 18 locations, including the Lake Erie basin and the Allegheny, Meramec, Obey, Ohio, Rockcastle, Susquehanna, and Wabash River systems. Results showed pronounced divergences among taxa ( θ ST= 0.92 – 0.97; p-distance = 0.025 – 0.039) presently designated as E. b. blennioides, E. b. newmanii, and E. b. pholidotum, as well as identification of a fourth clade in the Meramec River. Most traditional morphological characters were significantly different ( P= 0.0001) in distinguishing between E. b. blennioidesand E. b. pholidotum, including scale counts and degree of ventral squamation. However, the range of variation within these characters overlapped, obscuring accurate assignment of individuals to the taxa. The four significantly divergent taxa of the Greenside Darter complex should be evaluated further for potential elevation to species status.

Committee: Carol Stepien (Advisor) Subjects: Biology, Genetics

Doctor of Philosophy, The Ohio State University, 2008, Molecular, Cellular, and Developmental Biology

Organogenesis requires cells to coordinate their development. The similarities and differences between cells can be explained by looking at their gene expression profiles. At the center of these profiles, there are often transcription factors such as Pax and Sp-related factors that are critical for organ development.To understand the specific role of Pax factors in organogenesis, it is important to determine how they function with other transcription factors to regulate target genes.To better understand the different roles of EGL-38, I used an overexpression approach. I tested the effect of ectopic expression of EGL-38 and its mutants in animals. My results indicate that ectopic expression of EGL-38 and its male tail specific alleles (sy287 and gu22) in embryos induce embryonic lethality, whereas ectopic expression of its egg-laying defective allele (n578) does not have any effect. I hypothesized that these defects could be due to misexpression of different EGL-38 targets in response to the different alleles. Using gfp reporter transgenes, I have shown that indeed ectopic expression of EGL-38, induces ectopic expression of lin-48 (a known direct target).Pax factors have different targets in different cells because they are known to act in a combinatorial manner with other transcription factors. For example, lin-48 is a direct target of EGL-38 in the hindgut, but it is not expressed in other organs where EGL-38 is known to function. To identify EGL-38 cofactors important for lin-48 hindgut expression, I conducted a genetic screen. I have isolated two mutations (gu84 and gu85). I have shown that gu85 is a mutation in sptf-3, a Sp-related factor.I also characterized the role of sptf-3 in organogenesis. sptf-3(gu85) animals show developmental defects that are similar to these seen in Wnt mutants. Specifically, mutants exhibit the Bivulva phenotype similar to lin-17/frizzled and lin-18/Ryk mutants. I show that sptf-3(gu85) mutants are sensitive to the dose of lin-17. In (open full item for complete abstract)

Committee: Helen Chamberlin (Advisor) Subjects: Biology, Genetics; Biology, Genetics

Doctor of Philosophy, The Ohio State University, 2007, Integrated Biomedical Science

Improved treatment strategies in pediatric Hodgkin Lymphoma (HL) have resulted in a cure rate approaching 95%, yet the development of a second primary malignant neoplasm (SMN) is a risk for survivors. Oxidative stress has been linked to the development of cancer due to the damaging effects of reactive species on DNA, lipids, and proteins. The aim of this study was to assess the associations of antioxidant gene alleles and the risk of developing a SMN in HL patients. Polymorphisms were chosen from antioxidant-related genes including; SOD GPX, CAT, and NOS. Statistical analysis was completed using tests of association, haplotype, and multiple regression modeling. Out of the 36 SNPs that were included in the final analysis, 4 SNPs in the GPX1, GPX3, GPX4, and SOD2 genes, were potentially suggestive (p<0.05) of an association between genotype and the development of a SMN. Replication studies are necessary, though it is notable that polymorphisms within the GPX family may be associated with the development SMN in our cohort. X-chromosome inactivation (XCI) is an epigenetic process used to regulate gene dosage in mammalian females by silencing one X-chromosome. While the pattern of XCI is typically random in normal females, abnormalities of the X-chromosome may result in skewing due to disadvantaged cell growth. We describe a female patient with an X;1 translocation [46,X, t(X;1)(q28;q21)dn] and unusual pattern of XCI who was clinically diagnosed with Otopalatodigital syndrome (OPD) type 1. There was complete skewing of XCI in the patient, along with the atypical findings of an active normal X-chromosome and an inactive derivative X. OPD1 is characterized by skeletal abnormalities, craniofacial defects, and hearing loss. Mutations within the FLNA gene (Xq28) are known to cause OPD, though none were detected in our patient. Characterization of the translocation revealed that the patient's Xq28 breakpoint interrupts the DKC1 gene, located 400kB distal to FLNA. Analysis of t (open full item for complete abstract)

Committee: Julie Gastier-Foster (Advisor) Subjects: Biology, Genetics

Doctor of Philosophy, The Ohio State University, 2007, Molecular, Cellular, and Developmental Biology

Gene amplification is a common mechanism for oncogene activation in cancer and has been used in the past as a tag for the identification of novel oncogenes. DNA amplification is common in head and neck squamous cell carcinoma (HNSCC) where numerous amplification events and potential oncogenes have already been reported. We applied restriction landmark genome scanning (RLGS) to study gene amplifications in HNSCC and to locate novel amplified and uncharacterized regions on primary tumor samples. Amplified sequences were identified from 33 different chromosomal regions including DNA amplifications of regions previously reported (e.g. 3q26.3-q29) and novel regions such (e.g. 8q13.1, 8q22.3, 9q32, 10q24.32, 14q32.32, 17q25.1 and 20q13.33). One enhanced RLGS fragment on 8q22.3, where YWHAZ (14-3-3zeta, KCIP-1) is located, is found in 30-40% HNSCC cases. YWHAZ mRNA is frequently upregulated in patients' tumor tissues. Data obtained from fluorescent in-situ hybridization (FISH) and immunohistochemistry on HNSCC tissue microarrays further confirm its low-level and frequent gene amplification and protein overexpression. Furthermore, YWHAZ siRNA significantly suppresses growth rate of HNSCC cell lines, and overexpression of YWHAZ in HaCaT immortalized human skin keratinocyte line promotes its growth, plus morphological change. Reduced YWHAZ level increases G1/G0-phase proportion, decreases S-phase proportion and DNA synthesis rate. Based on these evidences, we suggest that YWHAZ deserves further investigation into its role in HNSCC carcinogenesis.

Committee: Christoph Plass (Advisor) Subjects: Biology, Genetics

Doctor of Philosophy, The Ohio State University, 2007, Molecular Genetics

In order to improve treatment and facilitate timely detection in Head and Neck Squamous Cell Carcinoma (HNSCC), elucidation of early detection markers is crucial. DNA methylation markers are advantageous, because DNA methylation is an early event. Following an overview on methylation in HNSCC in Chapter 1, we describe a genomewide screen using Restriction Landmark Genomic Scanning (RLGS) in Chapter 2. This analysis found a set of potential tumor suppressor genes that are commonly methylated. Not all relevant candidates are detected by RLGS because of limitations of the assay. In Chapter 3, a candidate gene approach identified C/CAAT enhancer binding protein alpha (C/EBPα), a gene previously shown to exhibit tumor suppressor activity in acute myeloid leukemia (AML) and lung cancer, as a gene of interest. C/EBPα tumor suppressor activity in lung cancer has previously been shown to be downregulated by epigenetic mechanisms. More recently, this gene has been found to be downregulated in HNSCC. This prompted investigation into the involvement of epigenetics in downregulating C/EBPα in HNSCC. It was revealed that C/EBPα is downregulated in HNSCC by loss of heterozygosity (LOH) and upstream DNA methylation. Also, C/EBPα overexpression in a HNSCC cell line (SCC22B) revealed its ability to provide tumor suppressor activity in HNSCC in vitro and in vivo. Upstream methylation of C/EBPα correlates with decreased expression in HNSCC. In Chapter 4, investigation of previously unstudied AP2α binding sites within the upstream methylated region demonstrated that AP2α suppresses C/EBPα promoter activity and protein expression. Methylation analysis of the upstream C/EBPα sequence after AP2α downregulation revealed decreased methylation, suggesting that AP2α binding may preceed and facilitate methylation and stable silencing of the gene. Finally, in Chapter 5 we discuss the relevance of the findings in the preceeding chapters and the future direction of this body of work.

Committee: Christoph Plass (Advisor); Amanda Simcox (Other); Christopher Weghorst (Other); Harold Fisk (Other) Subjects: Biology, Genetics