| 1. |
- Niss, Frida, 1991-
(author)
-
Imaging the molecular pathways of neurodegeneration : New pathologies of SCA7
- 2022
-
Doctoral thesis (other academic/artistic)abstract
- Spinocerebellar Ataxia type 7 (SCA7) is a genetic neurodegenerative disease with lethal outcome that affects the cerebellum and retina of patients. This thesis focuses on characterising molecular pathological pathways that cause toxicity and cell death in SCA7. Using primarily an inducible cell model and patient fibroblasts I found that the three RNA binding proteins FUS, TDP-43 and TIA1 are co-sequestered into aggregates formed by the SCA7 causing protein, ATXN7. Consequently I investigated the cellular functions in which these proteins have important roles. I found that FUS’ ability to regulate mRNAs is altered due to mislocalisation, DNA damage is increased, and that stress granules (SGs) are induced in a SCA7 cell model and in patient fibroblasts. Surprisingly, I also found that ATXN7 was present within SGs, and that SGs exhibited an altered shape upon induction of mutant ATXN7. I also participated in developing a microscopy-based method for monitoring chromatin organisation in live cells called FRIC. FRIC is able to detect even subtle changes to peripheral chromatin organisation, and since ATXN7 is a subunit of the transcription regulational complex SAGA, we used FRIC to investigate the effect of mutant ATXN7 on peripheral chromatin organisation. While we found no evidence that mutant ATXN7 affected peripheral chromatin organisation, the inner nuclear membrane protein Samp1 was found to be important for normal chromatin organisation in the nuclear periphery. Finally, I characterised the effect of mutant ATXN7 expression on the nuclear lamina, nuclear pore complexes, and nucleocytoplasmic transport. I found that although key transport factors such as Ran and Importin ß intermittently co-localised with ATXN7 aggregates, there were no apparent defects in nucleocytoplasmic protein import or nuclear envelope integrity. In summation, my investigations resulted in new findings that may be built upon to find key targets for treating SCA7 patients.
|
|
| 2. |
- Niss, Frida, et al.
(author)
-
Key Modulators of the Stress Granule Response TIA1, TDP-43, and G3BP1 are Altered by Polyglutamine Expanded ATXN7
- 2022
-
In: Molecular Neurobiology. - : Springer Science and Business Media LLC. - 0893-7648 .- 1559-1182. ; 59:8, s. 5236-5251
-
Journal article (peer-reviewed)abstract
- Spinocerebellar ataxia type 7 (SCA7) and other polyglutamine (polyQ) diseases are caused by expansions of polyQ repeats in disease-specific proteins. Aggregation of the polyQ proteins resulting in various forms of cellular stress, that could induce the stress granule (SG) response, is believed to be a common pathological mechanism in these disorders. SGs can contribute to cell survival but have also been suggested to exacerbate disease pathology by seeding protein aggregation. In this study, we show that two SG-related proteins, TDP-43 and TIA1, are sequestered into the aggregates formed by polyQ-expanded ATXN7 in SCA7 cells. Interestingly, mutant ATXN7 also localises to induced SGs, and this association altered the shape of the SGs. In spite of this, neither the ability to induce nor to disassemble SGs, in response to arsenite stress induction or relief, was affected in SCA7 cells. Moreover, we could not observe any change in the number of ATXN7 aggregates per cell following SG induction, although a small, non-significant, increase in total aggregated ATXN7 material could be detected using filter trap. However, mutant ATXN7 expression in itself increased the speckling of the SG-nucleating protein G3BP1 and the SG response. Taken together, our results indicate that the SG response is induced, and although some key modulators of SGs show altered behaviour, the dynamics of SGs appear normal in the presence of mutant ATXN7.
|
|
| 3. |
- Niss, Frida, et al.
(author)
-
Monitoring of Chromatin Organization at the Nuclear Pore Complex, Inner Nuclear Membrane, and Nuclear Interior in Live Cells by Fluorescence Ratiometric Imaging of Chromatin (FRIC)
- 2022
-
In: The Nuclear Pore Complex. - New York : Humana Press Inc.. - 9781071623367 - 9781071623374 ; , s. 151-160
-
Book chapter (peer-reviewed)abstract
- The image analysis tool FRIC (Fluorescence Ratiometric Imaging of Chromatin) quantitatively monitors dynamic spatiotemporal distribution of euchromatin and total chromatin in live cells. A vector (pTandemH) assures stoichiometrically constant expression of the histone variants Histone 3.3 and Histone 2B, fused to EGFP and mCherry, respectively. Quantitative ratiometric (H3.3/H2B) imaging displayed a concentrated distribution of heterochromatin in the periphery of U2OS cell nuclei. As a proof of concept, peripheral heterochromatin responded to experimental manipulation of histone acetylation as well as expression of the mutant lamin A protein “progerin,” which causes Hutchinson-Gilford Progeria Syndrome. In summary FRIC is versatile, unbiased, robust, requires a minimum of experimental steps and is suitable for screening purposes.
|
|
