Investigation of Microtubule Stability in Fibroblast Cultures of Spinal Muscular Atrophy Patients

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Thesis Type: Postgraduate

Institution Of The Thesis: Hacettepe University, Tıp Fakültesi (Türkçe), Temel Tıp Bilimleri Bölümü, Turkey

Approval Date: 2020

Thesis Language: Turkish

Student: Pelin Zobaroğlu

Supervisor: Gamze Bora

Open Archive Collection: AVESIS Open Access Collection

Abstract:

Spinal muscular atrophy (SMA) is an autosomal recessive inherited neurodegenerative disease, which mostly affects infants and children, characterized by motor neuron loss and muscle atrophy. Mutations in the survival of motor neuron 1 (SMN1) gene, located at the 5q13 chromosomal region, are the causes of SMA disease. SMN protein is expressed in all cells however, its deficiency primarily affects motor neurons. Absence of SMN causes defects in neuronal morphology and function which is associated with structural and functional defects in cytoskeleton elements. Microtubules, one of the basic elements of the cytoskeleton, are important for the establishment of axon and dendrite morphology in specialized cells such as neurons, and intracellular transport. In in vivo and in vitro SMA models, SMN deficiency leads to alterations in architecture and polymerization of microtubules, however underlying molecular mechanisms have not been elucidated. Microtubules are hollow cylindrical polymers, composed of alpha (α) and beta (β) tubulin proteins. Microtubule polymerization and depolymerization occurs by tubulin assembly and disassembly, respectively. Microtubule dynamics and stability are regulated by post-translational modifications of tubulin proteins and microtubule-associated proteins (MAPs). Various post-translational modifications (acetylation, detyrosination, delta2-tubulin, etc.) occur at the amino and carboxyl-terminal of the α and β tubulin proteins. Tubulin post-translational modifications play a role in the interaction of microtubules with other regulatory proteins and cytoskeletal elements. In our department, to explain the molecular mechanisms of SMA disease, basic studies have been performed. Previously, we showed significant alterations in the expression of some microtubule related proteins (MAP1B, MAP2 and EB3) and tubulin post-translational modifications (acetylation and detyrosination) in an in vitro and in vivo SMA models, indicating reduced microtubule stability. However, there is no study in literature investigating microtubule structure and stability in patient cells. Therefore, in this thesis, we aim to investigate microtubule stability in fibroblast cultures of SMA patients. The structure of the microtubule network and stability will be analyzed with commercially available fibroblast cultures of two patients from different clinical severities (Type I and Type II) and two healthy controls. With this thesis, microtubule stability will be studied for the first time in SMA patient fibroblast cultures. Detection of alterations in microtubule stability will provide a basis for further studies on microtubule functions (intracellular transport, organelle position, etc.) and small molecules for restoring microtubule stability.