Amyotrophic Lateral Sclerosis
​​Amyotrophic lateral sclerosis (ALS) is a devastating adult-onset neurodegenerative disorder. In ALS, motor neurons in the motor cortex, brainstem and spinal cord degenerate, resulting in motor problems, muscle weakness and paralysis. These motor impairments are progressive and ALS is usually fatal within 3 to 5 years after detection of the first symptoms. In approximately 10% of ALS patients the disease runs in the family. Here, neurodegeneration is caused by mutations in a heterogeneous set of genes. While the cause of the disease in sporadic cases is mostly unknown, the majority patients present with similar pathological aggregates of the RNA binding protein TDP-43 and related proteins. Since this pathology unites around 98% of ALS patients, understanding the causes and mechanisms of the misregulation and aggregation of RNA binding proteins will undoubtedly give us invaluable clues on disease pathogenesis, and will allow us to develop novel therapeutic approaches, which hopefully can interfere with this process and prevent neuronal death.

We and others have investigated the major genetic cause of the disease, being tandem repeat expansions in the C9orf72 gene, for its potential involvement in the pathology of RNA binding proteins in ALS.
Nucleocytoplasmic Transport
​​We and others identified repetitive peptides generated via non-canonical translation of the C9orf72 repeat expansions as major pathological species. Specifically, arginine-rich dipeptide repeats or DPRs, i.e., GR and PR, were found to be highly toxic in yeast and fly models of the disease. We set out to use high-throughput genetic screens to uncover the molecular determinants of DPR toxicity. We identified nucleocytoplasmic transport factors as key genetic modifiers, and found that arginine-rich DPRs indeed induce nucleocytoplasmic transport defects. Under normal conditions TDP-43 shuttles between the nucleus and cytoplasm through the nuclear pore complex. We now postulate that arginine-rich DPRs exactly perturb the function of the nuclear pore and related transport factors, and hence, could explain the initial cytoplasmic mislocalization of TDP-43 and other RNA binding proteins in C9orf72 carriers. Given that this pathology occurs also in sporadic cases, nucleocytoplasmic transport defects might be more broadly implicated in ALS.​​

(Jovicic et al., 2015 Nat. Neurosci.; Boeynaems et al., 2016 Sci. Rep.; Boeynaems et al., 2016 Acta Neuropath.)  
Protein Phase Separation
In recent years, several of these key ALS-related RNA binding proteins were found to possess the ability to undergo a liquid-liquid phase transition. This process provides a framework for the biogenesis of several membraneless organelles, including stress granules. Interestingly, due to similarity in protein content, these dynamic stress granules had been proposed as stepping stones towards pathological aggregate formation in ALS/FTLD. We found that arginine-rich DPRs can undergo liquid-liquid phase transitions. Moreover, they preferentially interacted with RNA binding proteins linked to ALS or stress granule metabolism, and promoted their insolubilization. Indeed, when expressed in cells arginine-rich DPRs did perturb stress granule dynamics and protein content. These findings argue that arginine-rich DPRs can indeed directly affect the phase separation of several ALS-related RNA binding proteins, and promote their aggregation in C9orf72 cases. These findings provide a better understanding of the pathogenic pathways initiated by the repeat expansion, and give us insight into the molecular underpinnings of protein phase separation in health and disease.

(Boeynaems et al., 2017 Mol. Cell ; Boeynaems et al., 2018 Trends Cell Biol.)