Exploration of developmental brain abnormalities in mouse models of Duchenne muscular dystrophy

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PROJECT SUMMARY Duchenne Muscular Dystrophy (DMD) results from mutations in the DMD gene, which generates the protein dystrophin. Although the gene produces various sized isoforms, only the largest isoform, Dp427, performs a critical function in skeletal muscle by linking the extracellular matrix (ECM) to the cytoskeleton. DMD mutations can also cause neurological dysfunction, but the cell and molecular basis of these changes are poorly under- stood. Interestingly, the severity of cognitive deficits seen in DMD worsens with successive loss of the smaller isoforms that cannot connect the ECM and cytoskeleton, implying additional functions these shorter dystrophins. DMD patients have reduced total brain and gray matter volume, with mutations that affect transcription of the mid-size isoform Dp140 being most strongly linked to this reduction16. I propose here to explore the role of dys- trophin in the developing ventricular/subventricular zone (V-SVZ), the major neural stem cell niche (NSC) in the adult mammalian brain. A key output of the V-SVZ during postnatal brain development is oligodendrocyte pro- genitor cells (OPCs), which go on to myelinate the forebrain. The timing and proper execution of myelination plays a critical role in many of the same neurological processes that are affected in DMD. Ependymal cells (ECs) are specialized multi-ciliated cells in the V-SVZ that line the ventricles of the brain that surround NSCs and regulate NSC quiescence and activation. My sponsor’s lab recently reported that dystroglycan, the binding part- ner of dystrophin, modulates notch signaling in V-SVZ NSCs to regulate both NSC fate decisions and the devel- opment of ECs24. Dystroglycan and dystrophin were also both found to influence postnatal OPC development, including delaying white matter tract myelination. Dysregulated notch signaling has been reported in muscle stem cells in animal models of DMD41, however, whether dystrophin regulates notch in NSCs remains unknown. In my first aim, I will investigate how dystrophin isoforms regulate early postnatal V-SVZ niche formation by examining EC development and organization into pinwheels. In the second aim, I will explore how dystrophin isoforms regulate V-SVZ NSC function and the production of neuronal and glial progenitors. Throughout I will examine dystrophin’s ability to regulate notch signaling in V-SVZ NSCs and test whether dystrophin-deficient cell phenotypes can be rescued by modulating the notch pathway. I will use small dystrophin constructs and DMD mouse models (mdx, mdx4cv, mdx3cv) in combination with notch activity reporter mice. Intriguingly, small dystrophins have been reported to translocate to the nucleus in muscle cells, indicating the potential for novel functional roles for small dystrophins in the nucleus of NSCs, which will be assessed by modification of se- quences needed for nuclear import/export. Lastly, as a complementary approach, I will use neonatal ventricle electroporation strategies to prevent or rescue dystrophin expression in the developing V-SVZ and use V-SVZ cell cultures that model NSC and EC development. Together, my studies will investigate dystrophin’s role in the formation and function of a crucial stem cell niche that generates neural progenitors for the postnatal brain.