Ph.D. Theses
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Browsing Ph.D. Theses by Author "Battaloğlu, Esra."
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Item From mutations to disease mechanism in Rett Syndrome, breast cancer, and congenital hypothyroidism(Thesis (Ph.D.)-Bogazici University. Institute for Graduate Studies in Science and Engineering, 2008., 2008.) Barış, İbrahim.; Battaloğlu, Esra.Epidemiological studies provide the correlative data to understand the etiology of human inherited diseases and develop efficient genetic testing assays. Additionally, the accumulated data of genetic and epigenetic findings, expression profiling, and proteomics allows disease diagnosis, to understand the molecular mechanisms leading to the disease pathogenesis, and to develop efficient therapeutic approaches. In the framework of this thesis, we have investigated genetic and epigenetic changes and performed genotypephenotype correlations to unravel the molecular mechanisms that lead to three different diseases, Rett Syndrome, breast cancer, and congenital hypothyroidism. The genetic basis of Rett Syndrome (RTT) was investigated in a total of 71 RTT patients. A heterogeneous spectrum of disease-causing MECP2 mutations was identified in 68.2 per cent of a clinically well defined group of cases whereas in only 12.5 per cent of the patients referred for differential diagnosis suggesting that this gene does not represent a major cause of the disease among patients with Rett-like features. For the first time, we have identified gene duplications as causative mutations in female atypical RTT cases. Consistent with the animal models, our results support the possibility that duplication of MECP2 that leads to increased expression might underlie some cases of X-linked delayedonset neurobehavioral disorders including Rett Syndrome. Our results showed that exon rearrangements that could not be detected by standard techniques contribute to 19.3 per cent of these MECP2 mutations, and should be considered in especially RTT variants in order to determine the actual significance of the gene in the etiology of RTT. Genotype/phenotype correlation was performed based on comparison of severity score of patients with the type and location of the mutation and the XCI pattern. The results did not reveal a statistically significant correlation, but, the patients with exon deletions were found to be more severely affected than patients with all other types of mutations and patients with exon duplications to present with severe eye contact problems. Additionally, we have developed and validate a novel multiplexed amplification refractory mutation system (ARMS) assay for identification of seven common mutations that accounts for almost 65 per cent of all MECP2 gene mutations. The validation studies revealed that our novel assay is an efficient, reliable, and cost-effective screen for molecular genetic testing of patients with RTT. Furthermore, we tested the effect of DNA concentration on reliablity and reproducibility of SYBR green dye-based Real Time PCR analysis to detect the MECP2 exon rearrangments. The results suggested that Real Time PCR analysis is reliable for determination of the exon copy number if the DNA amount is in the range of 1-50 ng. To our knowledge, there are no known reports investigating the role of methylation of hHR23A and hHR23B genes in the tumor tissues. We have characterized the 5' flanking region of the hHR23A and hHR23B genes using web-based analysis and investigated the involvement of methylation status of putative promoter region of hHR23A and hHR23B genes in breast carcinogenesis. The observations of the hypermethylation of hHR23A gene and the presence of methylated conserved motifs and transcription binding sites in hHR23B gene among the analyzed tumor tissues suggested the involvement of methylation of hHR23 genes in the breast carcinogenesis. Investigation of epigenetic changes in tumor samples of breast cancer patients was a pioneering work since available literature implicates its presence only in cell lines. Since our CH patient was the first case with Bamforth Syndrome and suffered the plasma cholinesterase deficiency, the genetic mechanisms leading to congenital hypothyroidism and prolonged paralysis after mivacurium were investigated. In contrast to other reported two patients with TTF2 gene mutation, the presence of thyroid tissue in our patient suggested further phenotypic heterogeneity associated with human TTF-2 mutations. The functional study with a collaborative work also helped to understand the genetic mechanisms and provided original evidence that implicated differential effects of TTF-2 mutations on downstream target genes required for normal human thyroid organogenesis.Item Genetic and molecular analyses of Turkish patients with pelizaeus - merzbacher disease(Thesis (Ph.D.)-Bogazici University. Institute for Graduate Studies in Science and Engineering, 2007., 2007.) Bilir, Birdal.; Battaloğlu, Esra.Pelizaeus-Merzbacher disease (PMD) is a rare inherited leukodystrophy with Xlinked recessive segregation. About 80 per cent of patients with PMD have been associated with mutations of the PLP1 gene on Xq21.3-Xq22 which encodes two proteins, PLP1 and DM20, expressed abundantly in oligodendrocytes. Mutations of GJA12 gene on 1q41-42 are responsible for some of the PMD cases with autosomal recessive inheritance. In the framework of this study, the molecular basis of PMD was investigated in a cohort of 21 Turkish families with PMD. In total, pathogenic mutations were identified in 57 per cent of the families, 19 per cent of which were due to PLP1 duplications, and nine and 29 per cent were due to mutations in the PLP1 and GJA12 genes, respectively. The distribution of the mutations identified in our cohort of patients was different from those reported in the literature, which may result due to the high frequency of consanguinity and autosomal recessive cases in our population. Absence of mutations in PLP1 or GJA12 genes in 43 per cent of the cases analyzed suggests presence of further genetic heterogeneity in PMD. In vitro immunocytochemical analyses of two PLP1 mutations identified in our cohort revealed that accumulation of mutant proteins in the endoplasmic reticulum, leading to UPR activation and subsequent apoptosis were observed for the mutant proteins. However, one of the mutations showed a different pattern of localization for DM20 isoform. Since patients present similar clinical features, the results implicate that PLP1 and DM20 may have different roles in myelin.|Keywords: Pelizaeus-Merzbacher Disease, Dysmyelination, Proteolipid Protein 1 (PLP1) Gene, Gap Junction Protein Alpha12 (GJA12) Gene, Unfolded Protein Response (UPR)Item Identification on new loci, genes, and mutations responsible for hereditary spastic paraplegia(Thesis (Ph.D.)-Bogazici University. Institute for Graduate Studies in Science and Engineering, 2018., 2018.) Özeş, Burçak.; Battaloğlu, Esra.Hereditary Spastic Paraplegia (HSP), characterized by lower limb spasticity and progressive weakness, is a group of inherited neurodegenerative disorders. 47 of the 63 genes responsible for HSP, have been associated with autosomal recessive (AR) HSP. In this study, whole exome sequencing analysis was performed for one patient from each 27 ARHSP families to identify causative genes. When WES data was not informative, homozygosity mapping was performed by using WES and/or whole genome SNP genotyping data. After segregation analyses of candidate variants, the causative variants were identified in fifteen families. SPG11 gene was causative in four families. Single families had mutations in CYP7B1 (SPG5A), SPG7, SPG15, and ALS2 genes. SACS and CYP27A1 genes that are associated with Charlevoix-Saguenay Spastic Ataxia and CTX, respectively were determined as the causative genes in two families providing differential diagnosis to these families. KIF1C (SPG58) (Caballero-Oteyza et al., 2014) and PLA2G6 genes (Ozes et al., 2017) identified in two families were reported as novel HSP genes. SAMHD1, ATAD1, and SEMA3C were identified as strong HSP gene candidates. Immortalized B-lymphocytes derived from family members were analyzed to unravel the involvement of candidate HSP genes on disease pathogenesis. ATAD1, SAMHD1 and CYP27A1 protein levels were shown to be reduced in immortalized cell lines of patients. This study contributed to understanding of the genetic heterogeneity of HSP by identifying five novel HSP genes, one of which locates to SPG27. We also underlined the importance of genetic analysis for differential diagnosis, and the necessity of primary genetic screening of SPG11 in AR-HSP-TCC and CYP7B1 in pure AR-HSP cases highlighting the effect of correct genotype-phenotype correlation.Item Investigation of novel recessive causative genes and gene / allele frequency for CMT disease in Turkey(Thesis (Ph.D.) - Bogazici University. Institute for Graduate Studies in Science and Engineering, 2021., 2021.) Candayan, Ayşe.; Battaloğlu, Esra.Inherited peripheral neuropathies are a group of genetic disorders of the periph eral nervous system. The most common type is called Charcot-Marie-Tooth (CMT) disease that constitutes an interesting research focus due to its clinical and genetic heterogeneity. Mutations in more than 90 genes and loci are associated with CMT that presents with autosomal dominant (AD), autosomal recessive (AR), X-linked, or mitochondrial inheritance. Despite the advances in genetic testing, approximately 35% of all CMT cases worldwide remain without a molecular diagnosis. The diagnostic rate is even lower for ARCMT due to the presence of many individually rare genes. This diagnostic gap points to the presence of yet unidentified causative genes, as well as po tential non-Mendelian features of the disease. In order to identify novel genes/alleles causing ARCMT and determine the frequency for known genes in Turkey, we have analyzed 56 consanguineous families diagnosed with CMT who present with early on set polyneuropathy with additional symptoms. Through the screening of patients for GDAP1 mutations, and subsequent whole-exome sequencing and homozygosity map ping, we have identified 22 recurrent and 13 novel alleles in known CMT genes achieving a potential diagnosis rate of 62,5%. Moreover, we identified FXN as a candidate gene for a novel disease in the spectrum between CMT and Friedreich’s ataxia, ATP8B3 for ARCMT2, and SEPT11 for AR-cerebellar ataxia with axonal neuropathy. This study paints the genetic landscape of the Turkish ARCMT population, reports candidate genes that might enlighten new disease mechanisms, and can serve as a reference for diagnosis strategies specific to populations with similar genetic backgrounds.Item Molecular and cellular development of cortical projection neurons: specification, diversity, and directed differentiation from endogenous progenitors for functional circuit repair(Thesis (Ph.D.)-Bogazici University. Institute for Graduate Studies in Science and Engineering, 2016., 2016.) Özkan, Abdulkadir.; Battaloğlu, Esra.; Macklis, Jeffery D..Responsible for perception, integration of sensory information, cognitive function, motor control, and consciousness, the complex, yet highly organized, six-layered mammalian neocortex contains distinct classes of neurons. Specific subtypes of cortical projection neurons are selectively vulnerable in distinct neurodegenerative, developmental, and acquired diseases of the central nervous system (CNS), resulting in irreversible functional deficits. Evidence for the existence of progenitors in restricted regions of the adult brain, and integration of new neurons into preexisting neural circuitry, support the feasibility of cellular repair in the CNS. However, functional repair of diseased or injured neuronal circuitry requires detailed understanding of molecular controls over development of neuronal lineages, and manipulation of these controls in progenitors to direct the differentiation of functional neurons with appropriate identity, maturity and circuit connectivity. In this study, I target endogenous cortical progenitors present in postnatal and adult brain to direct their differentiation into corticofugal projection neurons. Application of a select combination of central and complementary transcriptional controls, Ngn2, VP16:Olig2 and Fezf2, in cultured cortical Sox6+/NG2+ progenitors directs acquisition of cardinal morphological, molecular, and electrophysiological features of corticofugal projection neurons. These findings demonstrate the feasibility of achieving subtype-specific differentiation of cortical projection neurons from a widely distributed in vivo neocortical progenitor population. Further, in the framework of this thesis, I describe the ongoing effort to identify key molecular controls over development, diversity and connectivity of corticostriatal projection neurons, which would serve as a solid step toward achieving a holistic view of the establishment of corticostriatal circuitry and its potential dysgenesis in disease.Item The effect of nNav1.5 gene expression in breast cancer metastasis(Thesis (Ph.D.)-Bogazici University. Institute for Graduate Studies in Science and Engineering, 2006., 2006.) Yamacı, Rezan Fahrioğlu.; Battaloğlu, Esra.Breast cancer is the most common cancer among women. Its metastasis is lethal that can not be detected at microscopic levels using current techniques. Thus, there is need for reliable early metastasis markers. Neonatal Nav1.5 (nNav1.5) is a voltage-gated sodium channel (VGSC) and one of the potential early markers for breast cancer metastasis. In this study, we determined that nNav1.5 expression was in parallel with breast cancer metastasis and estrogen receptor (ER) expression in a group of patients. To provide data for future drug development, we analyzed the expression pattern of nNav1.5 protein in normal human tissues. The protein was not expressed in skeletal and heart muscle, brain, small intestine, colon, stomach, esophagus, urinary bladder and prostate but expressed in breast at basal level. We also investigated the distribution of VGSC in these non-excitable human tissues. Except urinary bladder, VGSC protein was determined mostly in secretory cells in all of the tissues above that may indicate a role in secretion. Upon identification of VGSC upregulation in tumor regions of different cancers including, colon, stomach, urinary bladder, kidney and lung it is possible that VGSC expression could be a widespread mechanism in cancer metastasis. Within the scope of this thesis, we also investigated the possible role of estrogen on nNav1.5 upregulation and activity in metastatic breast cancer. Estrogen had no effect on proliferation of cells but slightly increased motility through nNav1.5 in highly metastatic cells that express the protein. In weakly metastatic cells that do not posses nNav1.5, estrogen decreased motility slightly. The quantity of nNav1.5 protein was not affected by estrogen but functionally available form on the plasma membrane was increased only in the highly metastatic cells. These results may suggest that estrogen increases motility capacity of breast cancer cells by regulating nNav1.5 activity.Item The myelination puzzle: do fibroblast growth factors and their receptors have regulatory roles in peripheral nerve myelination?(Thesis (Ph.D.)-Bogazici University. Institute for Graduate Studies in Science and Engineering, 2014., 2014.) Dağlıkoca, Emine Duygu.; Battaloğlu, Esra.Axon-Schwann cell interaction is the key step in myelination and critical in the pathogenesis of peripheral neuropathies. Molecules mediating this interaction to promote the complex mechanism of myelination have not been elucidated properly yet. However, recent studies emphasized the importance of tyrosine kinase receptor signaling in the process. Although expression of FGFs and their tyrosine kinase receptors FGFRs have been shown in PNS, little information is available about their regulation during the myelination process. In this study, we aimed to investigate whether FGF1, FGF2, FGF9 that were previously implicated in CNS development, involved in peripheral nerve myelination and regulate axon-Schwann cell interactions, through their high affinity receptors FGFR1-3. For this purpose, we used dorsal root ganglia (DRG) and the sciatic nerve from mice as in vivo models and Schwann cell-neuron co-culture developed from fetal mouse DRG tissue as an in vitro model. The expression patterns and localization of the molecules were investigated both in vivo and in vitro. Among the three FGFs analysed, FGF1 was chosen as a candidate for further investigation because of its high level of expression in all tested tissue types with an axonal localization. In contrast, FGFR1-3 were found to be expressed by Schwann cells. Protein expression levels of FGF1 and FGFR1-3 were examined through the developmental stage to adulthood from sciatic nerves by Western blotting. Both FGF1 and its receptors were found to be modulated at key time points of the myelination route. Immunolabeling studies showed that FGF1 expression in neurons and FGFR1-3 expression in Schwann cells continued throughout the process. When FGF1 was blocked in DRG culture, a reduction in the levels of myelin proteins and in the number of myelinated axonal segments was observed. Our findings provide evidence for the first time for the involvement of FGF1 in peripheral nerve myelination and suggest that FGF1 signaling through FGFR1-3 have regulatory roles at the onset of myelination, in myelin compaction and protection of the stability of mature structure.