Understanding Hereditary Sensory and Autonomic Neuropathy type IV through a novel knock-in mouse model

in pain sensation. Indeed, a functional NGF-TrkA system is an essential requisite for the generation and maintenance of long-lasting thermal and mechanical hyperalgesia in adult mammals. Mutations in the gene encoding for TrkA are responsible for a rare condition, named Hereditary Sensory and Autonomic Neuropathy type IV (HSAN IV), characterized by the loss of response to noxious stimuli, sweating defects and cognitive impairment. However, to date, there is no available mouse model to properly understand how the NGF-TrkA system can lead to pathological phenotypes that are distinctive of HSAN IV. Since the diversity of HSAN IV TrkA-related mutations determines variable degrees of clinical phenotype and intellectual disabilities in affected individuals, we have decided to deeply investigate the missense Arg649Trp (R649W) mutation, located in the intracellular tyrosine kinase domain of TrkA receptor and known to induce a diminished kinase activity and reduced phosphorylation after NGF stimulation in transfected cells. First, by in vitro biochemical and biophysical analyses, I showed that the pathological R649W mutation leads to kinase-inactive TrkA, reducing the constitutive ubiquitination and also affecting the membrane dynamics and trafficking. Then, after the generation of the knock-in mouse line carrying the HSAN IV TrkAR649W mutation, I demonstrated that TrkAR649W/m mice displayed a lower response to thermal and chemical noxious stimuli, correlating with reduced skin innervation and altered expression of nociceptive markers in Dorsal Root Ganglia (DRGs). By performing a sweat assay, I also found that the pathological TrkAR649W mutation causes sweating deficits in HSAN IV TrkAR649W/m mice compared to TrkAh/m controls. Moreover, the R649W mutation decreased anxiety-like behavior and compromised cognitive abilities, by impairing spatial-working and social memory. In addition, the results obtained in this thesis uncovered unexplored roles of TrkA in thermoregulation and sociability. By exploiting suitable control animal models such as HSAN V NGFR100W/m and TrkA+/- mice, I demonstrated that HSAN IV TrkAR649W/m mice mimic the clinical phenotype of HSAN IV patients and they can be considered a suitable experimental platform to explain the clinical aspect of HSAN IV disease, also offering promising new routes for testing future therapies.