@phdthesis{vanDiepen2017, author = {Laura-Kristina van Diepen}, title = {Untersuchungen zur Aufkl{\"a}rung der molekularen Ursachen erblich bedingter Erkrankungen des zentralen Nervensystems}, journal = {Investigations into the molecular causes of hereditary disorders affecting the central nervous system}, url = {https://nbn-resolving.org/urn:nbn:de:gbv:9-opus-21831}, pages = {203}, year = {2017}, abstract = {Intellectual disability (ID) is a cognitive impairment disorder characterized by an intelligence quotient (IQ) below 70 and is one of the most common forms of cognitive handicaps resulting from genetic aberrations [1]. Hereditary forms of ID are genetically heterogeneous but a considerable number involve genes participating in the build-up and maintenance of the glycocalyx [2]. The glycocalyx is composed of carbohydrates that are part of the cell surface exposed lipids and proteins (including proteoglycans) or part of the extracellular matrix. We recently demonstrated that mutations in ST3GAL3 (NM\_006279.2), which encodes the Golgi enzyme β-galactoside-α2,3-sialyltransferase-III, lead to varying clinical manifestations. Two independent mutations (p.Ala13Asp and p.Asp370Tyr) identified in Iranian pedigrees were associated with mild forms of non-syndromic autosomal recessive intellectual disability (NSARID)[3]. However, a third point mutation (p.Ala320Pro) identified in a Palestinian family caused West Syndrome (WS), an age-dependent epileptic encephalopathic syndrome associated with developmental arrest or, as in the case of our patients [3], regression[4]. In humans ST3GAL3 forms, among others, the sialyl Lewis a (sLea) epitope on proteins. Ectopic expression of full-length-c-MYC fusion proteins with the respective mutations in LMTK- cells showed severe functional impairment and subcellular mislocalization of all mutant gene products [5]. To further elucidate the molecular and cellular mechanisms causing West Syndrome due to the lack of ST3GAL3 function, we successfully generated induced pluripotent stem cell lines from fibroblasts obtained from a patient with West Syndrome, carrying a mutation in exon 12 (c.958G>C, p.Ala320Pro) of ST3GAL3, and a healthy sibling using lentiviral reprogramming. Since ST3GAL3 showed highest expression in the frontal cortex, which is also in accordance with the source of epileptic seizures, a differentiation protocol to cortical neurons was established and successfully accomplished for both cell lines. One of its advantages is mimicking the in vivo neurogenesis in vitro, enabling us to investigate temporal processes. IPSCs and cortical neurons derived thereof were analysed by lectin blots, mRNA sequencing, adherence assays, and FACS. While no significant difference was observed at stem cell or fibroblast level between patient and control cells, patient-derived cortical neurons displayed an additional band (70 kDa) in the lectin blot staining, enhanced adherence to a poly-L-ornithine/laminin coated surface and decreased levels of neurons expressing T-box transcription factor brain 1 (2.28 fold). Thus, our results indicate that ST3GAL3 function is important for the normal development and functioning of the brain.}, language = {de} }