Browsing by Subject "Spectrin"

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    Murine models of spinocerebellar ataxia type 5
    (2009-06) Armbrust, Karen Rose
    Spinocerebellar ataxia type 5 (SCA5) is a slowly progressive neurodegenerative disease of the cerebellum caused by mutations in the SPTBN2 gene, which encodes the protein β-III spectrin. To characterize how β-III spectrin with the American SCA5 mutation causes Purkinje cell degeneration and cerebellar dysfunction, I developed the first transgenic murine models of SCA5 and identified brain proteins that potentially interact with the region of β-III spectrin where the American SCA5 mutation occurs. Behavioral studies with a conditional model that drives expression of untagged β-III spectrin and a second 3xFLAG-tagged SCA5 model show that overexpressing mutant β- III spectrin in murine cerebellar Purkinje cells causes cerebellar dysfunction. Further studies with the conditional tet-regulated mice show that untagged mutant β-III spectrin alters the localization of the glutamate transporter EAAT4 and the metabotropic glutamate receptor mGluR1α and produces a concomitant deficit in mGluR1 function. Histologic analysis of the 3xFLAG-tagged SCA5 murine model shows that the American SCA5 mutation also alters the Purkinje cell distribution of the mutant β-III spectrin protein itself. Additionally, I identified a number of brain proteins that are novel β-III spectrin interaction candidates, including the dynactin subunit p150Glued. I show that the American and French SCA5 mutations alter the interaction strength of β-III spectrin with p150Glued and α-II spectrin respectively.
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    Role of spectrin mutations in spinocerebellar ataxia type five (SCA5)
    (2009-08) Lorenzo Vila, Damaris Nadia
    Spinocerebellar ataxia type 5 (SCA5) is a dominant neurodegenerative disorder caused by mutations in the SPBTN2 gene encoding the cytoskeletal protein beta-III spectrin. To get insight into the biology of the disease and the normal function of beta-III spectrin, and to estimate the frequency of SCA5 mutations among ataxia patients, I used a forward human genetic approach to identify novel SPTBN2 mutations. Screening of the SPTBN2 gene in a cohort of families with dominant ataxia of unknown etiology and a large group of ataxia samples identified seventeen novel variants not found in the general population. Putative mutations were identified in the areas comprising the second calponin homology domain, spectrin repeat two to four, and the ninth spectrin repeat of beta-III spectrin. To investigate the downstream effects of the American and German SCA5 mutations in neurons, I established a series of transgenic Drosophila models that express human beta-III-spectrin or fly beta-spectrin proteins containing SCA5 mutations. Through genetic and functional analyses I show that expression of mutant spectrin in the eye causes a progressive neurodegenerative phenotype and expression in larval neurons results in posterior paralysis, reduced synaptic terminal growth, and axonal transport deficits. These phenotypes are genetically enhanced by both dynein and dynactin loss-of-function mutations. I have additionally used the SCA5 fly models to conduct modifier screens and identify genes and biological pathways that may contribute to SCA5 pathogenesis. These studies revealed genetic interactors implicated in a wide range of biological functions including intracellular transport, synapse formation and function, protein homeostasis, and transcription regulation.