TDP-43, FUS, SOD1 Localization and Aggregation Assay

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that forms cytoplasmic aggregates through proteins including TDP-43 and SOD1 in cells affected in the central nervous system (CNS). Protein aggregation in affected motor neurons is a central feature of ALS. The analysis of protein mislocalization, aggregation formation, and proteolysis defect in ALS may provide the basis for understanding the common pathological events of ALS. At Creative Biolabs, we have the expertise to optimize each stage in protein localization and aggregation assay to ensure you get your desired outcome, achieving the highest levels of efficiency throughout.

The Molecular Makeup and Formation of Cellular Aggregates in ALS

In ALS, several different proteins have been implicated in neurodegeneration, including SOD1, TDP-43, and FUS. The dysregulation of protein localization is one of the hallmarks of ALS. The formation of aggregates is considered to be involved in the pathogenesis of ALS.

• TDP-43

TDP-43 is an aggregation-prone protein under strict autoregulation and belongs to a family of RNA-binding proteins. It forms a homodimer even in the absence of RNA. It has a range of physiological functions involving transcription and translation control, including splicing and regulation of non-coding RNA. The accumulation of aggregated and post-translationally modify TDP-43 is considered a biochemical signature of ALS and may play a significant role in driving pathogenesis. TDP-43 is a shuttling protein, that moves from the nucleus to the cytoplasm. Impairment of nucleo-cytoplasmic transport of TDP-43 may contribute to ALS. Normally, TDP-43 is localized mainly in the nucleus.

• SOD1

The mutations in the gene encoding copper-zinc SOD1, account for 20% of fALS cases. Its canonical role is as a free radical scavenger, but there is emerging evidence that it may also act as a transcription factor and an RNA binding protein. ALS-associated mutations, as well as aberrant posttranslational modifications to the WT protein, destabilize SOD1 and cause it to misfold.

• FUS

FUS is another RNA-binding protein that is usually found in the nucleus but has an increased tendency to aggregate due to ALS-related mutations, with a small portion deposited in cytoplasmic inclusion bodies in both sALS and fALS cases. Most of the ALS-related FUS mutations reported so far are located in the NLS of this protein, resulting in impaired nuclear transport of the FUS. FUS mediates a wide range of cellular processes, including DNA repair, splicing, transcription, and miRNA processing. The protein shuttles between the nucleus and cytoplasm and are involved in mRNA transport.

Fig.1 Overview of TDP 43 and FUS toxicity through aberrant localization and aggregation. (Tsubota, et al., 2016)

Molecular Mechanisms Underlying Protein Aggregation in ALS

• Low Complexity Domains in ALS Proteins with Aggregation Prone Properties

RNA-binding proteins (RBPs) such as FUS and TDP-43 contain domains similar to those of yeast prions. Aggregation of FUS and TDP-43 has been shown to rely on regions resembling prion domains and mutations in TDP-43 associated with ALS occur mainly in its prion-like region.

• Stress Granule (SG) Formation and ALS Protein Aggregation

ALS-related mutations in TDP-43, FUS, and ATXN2 were associated with SGs.

• Protein Sequestering

FUS and TDP-43 aggregation may play a toxic role by sequestrating multiple binding partners or even interactions that are critical to neuronal function.

• Dysfunction of Protein Degradation Pathways

Components of the protein degradation pathway have become important regulators of protein aggregation and toxicity in ALS.

Fig.2 Cellular mechanisms linked to protein aggregation in ALS. (Blokhuis, et al., 2013)

Creative Biolabs is a leading international biotechnology company, we believe in the transformative power of science and technology. Further understanding of the aggregation of TDP-43, FUS, and SOD1 in ALS is important, as they will provide insights into the pathogenesis and degeneration of motor neurons, and provide ideas and opportunities for the development of new therapeutic strategies. Please feel free to contact us for more details about TDP-43, FUS, SOD1 localization, and aggregation assay services.

References

1. Tsubota, A.; et al. Mislocalization, aggregation formation and defect in proteolysis in ALS. AIMS Molecular Science. 2016, 3(2): 246-268.
2. Blokhuis, A.M.; et al. Protein aggregation in amyotrophic lateral sclerosis. Acta neuropathologica. 2013, 125(6): 777-794.