Development of the cerebellum is exquisitely dependent on the Sonic Hedgehog (SHH) pathway that drives the expansion of granule cell progenitors (GCPs) and other cells. Impairment of this pathway leads to cerebellar hypoplasia. On the opposite, its untimely/excessive activation results in medulloblastoma development, supporting the oncogenic role of the pathway. SHH signal transduction largely occurs in the primary cilium (PC), an antenna-like structure enriched in receptors and other signaling molecules. This is organized by the basal body in non-mitotic cells. After PC disassembly, the same organelle organizes the centrosome, before mitotic onset. Dysfunctions of this dynamic organelle affect SHH pathway and are involved in genetic diseases known as ciliopathies, in CNS developmental diseases and cancer.
Nbs1 is a member of the Mre11/Rad50/Nbs1 complex, which is essential for genome integrity and the DNA damage response. Mutations in the Nbs1 gene cause the Nijmegen Breakage syndrome, a DDR-defective syndrome characterized by immunodeficiency, microcephaly and cancer predisposition.
Taking advantage of a CNS restricted Nbs1 knock-out mouse model, we recently demonstrated that Nbs1-KO abrogates tumorigenesis in a spontaneous model of SHH-MB and impairs the growth of SHH-MB allografts. Inducible Nbs1-KO inhibits the SHH-pathway in developing GCPs, in vivo and in vitro. Hints from the literature and our preliminary data suggest that Nbs1 may exert previously unrecognized functions on the PC, thus affecting SHH-dependent cerebellar development and tumorigenesis.
In this project we will develop new animal and cellular models to specifically address this hypothesis and we will integrate these studies with dynamic biochemical and imaging analysis of PC structure and function.
We believe that disclosing a novel role for Nbs1 on PC and SHH pathway may provide new insights into different genetic diseases affecting cerebellar development and cancer.
The characterization of the emerging role of NBS1 on the regulation of ciliogenesis and SHH pathway has several important implications, and candidates to reach results that will be important and innovative for multiple fields related to human health. Few examples are provided below.
1. Identification of the molecular mechanisms through which DDR proteins, such as NBS1 and MRE11, regulate PC structure and PC-dependent signal transduction is per se an innovative biological issue, since it may provide evidence for previously undescribed molecular functions of these molecules, in addition to their well-established role in the DDR.
2. The main focus of the application deals with SHH-dependent tumorigenesis and will shed light on the role of NBS1 in this regard. A better understanding of SHH signaling pathway and its interaction with previously unpredicted players, such as the MRN complex, may lead to identification of new anticancer strategies. Although clinical-grade MRN inhibitors are not yet available, the possibility to target the MRN complex for cancer therapy has already been approached in several labs, including ours (11).
3. Primary cilium aberrations, SHH and DDR genetic defects are at the base of a huge number of inherited syndromes, known as ciliopathies and DDR-defective syndromes, including the Nijmegen Breakage Syndrome, due to NBS1 mutations. Importantly, most causes of microcephaly are genetically determined and appear to be often due to proteins located and function at the centrosome, the alter ego of the PC, in different phases of the cell cycle. Improving our understanding of the molecular interactions between the DDR and the PC/centrosome world may provide new hints for a better comprehension of these genetic disorders, that are mostly orphan of effective therapeutic intervention.
4. An additional outcome of our work work will be the development and characterisation of mouse and cellular models that might be relevant frothier studies related to the human diseases described above.
For these reasons, we are confident that our proposed investigation will have a strong impact on several fields of biology and human health.
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