Generation and analysis of mouse models to define the roles of the ALS-associated factor Zfp106/ZNF106 in neurodegeneration

Amyotrophic lateral sclerosis (ALS) and related motor neuron degenerative diseases affect more than 30,000 people in the US, and 90% of patients die within five years of diagnosis. Although ALS and ALS-like disease manifestation varies, all forms are characterized by progressive loss of motor neurons. These disorders have a strong genetic component, and although more than 50 genes involved in ALS have been identified, much still remains to be learned about the genes and mechanisms that contribute to ALS susceptibility and protection. Aberrant RNA metabolism and processing plays an important mechanistic role in neurodegeneration, and mutations in genes encoding RNA-binding proteins underscore several of the more common causes of ALS. The general idea that altered RNA metabolism and processing leads to neurodegenerative disease suggests that additional genes encoding RNA-binding or RNA processing factors are likely to be involved in the genetics of ALS and related neurodegenerative disorders. The ZNF106 gene encodes an RNA binding protein and maps to human chromosome 15q15.1, the site of an incompletely described rare form of juvenile onset ALS. Genetic inactivation of Zfp106, the mouse ortholog of ZNF106, results in an early onset, profound ALS-like neurodegenerative phenotype with a corresponding loss of motor neurons, suggesting that this factor plays a neuroprotective role in vivo and that its loss-of-function could be involved in ALS in humans. Zfp106 is expressed in the nucleolus and nuclear speckles in both motor neurons and skeletal muscle and its function is essential for maintenance of neuromuscular junctions in vivo. Zfp106 binds selectively to G-quadruplex-forming RNA molecules and strongly associates biochemically with other RNA binding proteins, including several with strong causative roles in ALS, further suggesting the possibility that Zfp106/ZNF106 might play a role in ALS. Consistent with this notion, unpublished work has identified two unrelated patients with a rare, homozygous variant in ZNF106 that is associated with adolescent onset ALS-like neurodegenerative disease. Introduction of the human disease-associated ZNF106 variant into mouse Zfp106 also results in profound juvenile onset ALS-like neurodegeneration similar to the disease observed in the patients and in Zfp106-null mice. This proposal will test the hypotheses that mutations in human ZNF106 cause or contribute to ALS-like neurodegeneration and that Zfp106 plays a protective role in maintaining neuromuscular junctions through its RNA binding function and its autonomous functions in either motor neurons or skeletal muscle. Studies proposed in this application will define the functional consequences ALS0associated ZNF106 variants by characterizing the ALS-like neurodegenerative phenotype in mice harboring knock-in of the variant and by determining the effect of the variant in cell culture-based and in vitro assays of ZNF106/Zfp106 protein function. These studies will also determine the cell autonomous requirement for Zfp106 in motor neurons and skeletal muscle using conditional genetic approaches in mice.