Fortuna, Tyler
(2023)
Identifying the molecular pathways perturbed by pathogenic mutations in GEMIN5.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
The heterogeneity of patient symptoms derived from neurological diseases hampers diagnosis and management. Many neurological patients go through a diagnostic odyssey with very little hope of pinpointing the main source of their neurological defects. Recent developments in next generation sequencing technologies have helped identify these genetic defects that have previously been unrecognized to cause neurological diseases. Using this approach, we identified biallelic mutations in the GEMIN5 gene among individuals exhibiting overlapping neurological symptoms of developmental delay, hypotonia, ataxia, and cerebellar abnormalities. Across these patients, we discovered autosomal recessive variants of GEMIN5 in 39 individuals from 29 unrelated patient families. To date, mutations in GEMIN5 have not been clinically linked to any neurological disorders/diseases.
GEMIN5 is an RNA-binding protein that is essential for assembly of the survival motor neuron (SMN) protein complex and facilitates the formation of small nuclear ribonucleoproteins (snRNPs), the building blocks of spliceosomes. Here, we describe several studies to investigate the molecular mechanisms behind mutant GEMIN5, which may be the driving force behind the neurological defects observed in the identified patients. To understand the consequences of biallelic variants of GEMIN5, we generated induced pluripotent stem cells (iPSCs) harboring GEMIN5 mutations and differentiated them into neuronal cells. We found that mutations in GEMIN5 perturb the subcellular distribution, stability, and expression of GEMIN5 protein and its interacting partners, suggesting a loss-of-function (LOF) mechanism. Furthermore, we investigated the deleterious effects of loss of GEMIN5 protein in vivo and discovered that a reduction in GEMIN5 protein results in developmental defects, motor dysfunction, and a reduced lifespan in Drosophila and mice. In addition, we explored molecular determinants of GEMIN5-mediated disease. Using a Drosophila model of GEMIN5-mediated disease, we identified survival motor neuron (SMN) as a genetic suppressor of GEMIN5-mediated neurotoxicity. We discovered that an increase in SMN expression by either SMN gene therapy replacement or the antisense oligonucleotide (ASO) Nusinsersen, significantly upregulated the endogenous levels of GEMIN5 in mammalian cells and mutant GEMIN5 derived iPSC neurons. Collectively, these studies provide evidence that pathogenic variants in GEMIN5 result in a neurodevelopmental ataxia syndrome and indicate the importance of identifying modifiers of GEMIN5 toxicity which may be beneficial in identifying future therapeutic targets for this disease.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
8 December 2023 |
| Date Type: |
Publication |
| Defense Date: |
1 December 2022 |
| Approval Date: |
8 December 2023 |
| Submission Date: |
14 December 2022 |
| Access Restriction: |
1 year -- Restrict access to University of Pittsburgh for a period of 1 year. |
| Number of Pages: |
261 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
School of Medicine > Pediatrics |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
GEMIN5
Neurodegeneration
SMA
Gene Therapy
iPSC Neurons |
| Date Deposited: |
08 Dec 2023 16:21 |
| Last Modified: |
08 Dec 2023 16:21 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/44022 |
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