Myosin X Was Involved In NLT Cell Migration And Cell Polarity

Myo X, an unconventional myosin expressed only in vertebrates, could interacts both with actin and microtubule, implying a relationship between Myo X and cell migration. Our group has proved that Myo X influences cell adhesion and migration. And in this situation, we continue to focus our work on the function of Myo X in cell-cell adhesion. N-Cadherin is a transmembrane glycoprotein, and present knowledge suggests that N-Cadherin is closely related to cytoskeletal microtubule system. By over expressing N-Cadherin, migratory ability and adhesion efficiency of low E-Cadherin expressed cells were significantly enhanced, indicating the promoting function of N-Cadherin in cellular migration and invasion. Accordingly, both Myo X and N-Cadherin were proved to affect cell motility by interact with cytoskeleton, thus we supposed that Myo X, as a cytoskeletal protein, is possibly participate in N-Cadherin induced cell migration.Firstly, we construct miRNA to knock-down endogenous Myo X in NLT. For the next step, we performed the Boyden's chamber assay, manifest differences occurred between Scramble RNA and Myo X miRNA, that fewer MyoΧmiRNA cells were observed in the lower chamber. Then we used the collagen gel assay to analyze cell migration in 3-D matrix, and MyoΧmiRNA shorten the cell migratory distance and made cell lose orientation. In the Wound healing assay, with Golgi complex stained to display cell polarity and10% NCS induction, MyoΧmiRNA cells were randomly oriented compared to Scramble miRNA transfected cells. By immunofluorescence staining, we found co-localization of Myo X and N-Cadherin in NLT cells. Interestingly, N-Cadherin was proved simultaneously less expressed by immunofluorescence and Western blot assay. In conclusion, our research demonstrate that knock-down Myo X in NLT cells could influence the migratory ability and orientation, also down regulate N-Cadherin expression in NLT cells which lay a good foundation for investigating the role of Myo X in GnRH nerve cells migrating process by ex vivo explants culture and GnRH transgenic mice.