Investigating the mechanism of MT motor protein recruitment to the NE in muscle and its role in myonuclear positioning

Myotubes are syncitial cells that make up muscle fibres, a primary component of muscle tissue. Myotubes are formed by the fusion of myoblasts in a process termed myogenesis. In healthy muscle tissue, upon myoblast fusion, the multiple nuclei of the myotube move to be roughly equidistant from one another and then proceed to the cell periphery. Nuclear positioning that deviates from this is observed in muscle disorders such as Emery-Dreifuss muscular dystrophy.
Nesprin-12 is a short isoform of the ubiquitously expressed Nesprin-1 and is specifically expressed in muscle tissues. Nesprins are nuclear envelope (NE) proteins that form part of the LINC complex, a bridge between the cytoskeleton and nuclear matrix. Nesprin-12, along with a MT organising centre (MTOC) protein called PCM1, have been implicated in the recruitment of motor proteins, kinesin-1 and dynein/dynactin, to the NE and anchoring them there during myogenesis. The recruitment of motor proteins to the nuclear envelope is thought to assist nuclear repositioning by facilitating nuclear movement along the microtubules. Kinesin-1 has been shown to be recruited to the NE by Nesprin-12. Specifically, kinesin light chains1 and 2 (KLC1/2), bind via a conserved 'LEWD' motif in Nesprin-12. In contrast, Nesprin-12 interaction with dynein/dynactin is yet to be investigated and the precise role of PCM1 is also unknown.
The aim of this project is to determine how motor proteins are recruited to the NE in myogenesis through the characterization of interactions between the motor proteins, Nesprin-12 and PCM1 and whether these interactions are affected by the presence of disease-associated mutations. The aims will be addressed using both structural-based and cell biology methods. It is expected that the project will provide insight into the mechanisms of nuclear repositioning by the recruitment of motor proteins to the NE and build on the current model of myogenesis.