All ETDs from UAB

Advisory Committee Chair

Michael Brenner

Advisory Committee Members

Anne B Theibert

Scott Wilson

Document Type


Date of Award


Degree Name by School

Master of Science in Biomedical Science (MSBMS) School of Engineering


Alexander disease (AxD) is a fatal astrogliopathy usually caused by mutations in the GFA gene, which encodes glial fibrillary acidic protein (GFAP). In this document, two unusual sequence differences in GFA that are associated with AxD are characterized. The first difference is the only reported case of an AxD-causing mutation occurring in a non-coding region of GFA, c.(619-3C>G), resulting in the largest known in-frame deletion in GFAP due to abnormal splicing in a small fraction of the GFA transcripts. The second difference is the farthest reported C-terminal point mutation in GFAP, g.1277A-->T, which produces a Q426L coding change. This mutation occurs in a region traditionally considered functionally unimportant. My work contributes genetic and functional evidence that both these sequence differences inhibit normal GFAP polymerization and cause Alexander disease. In the case of the non-coding c.(619-3C>G) mutation, I tested 525 samples from healthy, ethnically matched individuals for the presence of the mutation using a mutation-specific restriction enzyme. All 525 were negative, indicating that the sequence difference was not a polymorphism present in healthy individuals. I also verified the assumption of a lab-memberfs polymerization assay that co-transfection of equal amounts of plasmids encoding wild-type and exon-4 skipped GFAP would result in equal amounts of the respective GFAP proteins being produced. This confirmed the lab-memberfs conclusions that low amounts of exon-4 skipped GFAP are sufficient to alter polymerization. In the case of the far C-terminal sequence difference, g.1277A-->T, I transfected GFAP expression plasmids with the point mutation into a cytoplasmic intermediate filament-free cell line and examined GFAP polymerization using immunostaining. I found that the mutation caused GFAP to form amorphous perinuclear aggregates. These findings indicate that very low levels of abnormal GFAP transcripts are sufficient to induce Alexander disease, and recommend sequencing non-coding regions of GFAP for identifying disease-causing mutations. They also show that coding differences C-terminal to the arginine-glycine-aspartic acid (RDG) integrin binding site in GFAP can be disease causing and fatal.

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