Overexpression of the dystrophins Dp40 and Dp40L170P modifies neurite outgrowth and the protein expression profile of PC12 cells

Abstract

Dp40 is ubiquitously expressed, including in the central nervous system. Dp40 mRNA and protein are detected in the early stages and postnatal stages of the mouse brain, respectively. In addition to being present in the nucleus, membrane, and cytoplasm, Dp40 is detected in neurites and postsynaptic spines in hippocampal neurons. Although Dp40 is expressed from the same promoter as Dp71, its role in the cognitive impairment present in Duchenne muscular dystrophy patients is still unknown. Here, we studied the effects of overexpression of Dp40 and Dp40L170P (a mutant of Dp40) during the neuronal differentiation process of PC12 Tet-On cells. We found that Dp40 overexpression increased the percentage of PC12 cells with neurites and neurite length, while Dp40L170P overexpression decreased them compared to Dp40 overexpression. Two-dimensional gel electrophoresis analysis carried out in nerve growth factor-differentiated PC12-Dp40L170P cells showed that the protein expression profile was modified compared to that of the control cells (PC12 Tet-On). The proteins with the highest upregulated expression were α-internexin and S100a6, which are involved in cytoskeletal structure. The expression of vesicle-associated membrane proteins increased in differentiated PC12-Dp40 cells, in contrast to PC12-Dp40L170P cells, while neurofilament light-chain was decreased in both differentiated cells. HspB1 was absent in undifferentiated cells and weakly detected in all differentiated cells. These results suggest that the subcellular distribution and expression of Dp40 has an important role in the neurite outgrowth of PC12 cells through the regulation of proteins involved in neurofilaments and exocytosis of synaptic vesicles, functions that might be affected in PC12-Dp40L170P.