Active transport by cytoplasmic dynein sustains dendritic MAP2 localization in developing neurons

MAP2 has been widely used as a marker of neuronal dendrites because of its extensive restriction in the somatodendritic region. Despite that, how the precise localization of such a soluble protein is established and maintained against diffusion has been elusive and long remained a mystery in neuroscience. In this study, using GFP-tagged MAP2 expressed in cultured hippocampal neurons, we discovered a crucial protein region responsible for the localization of MAP2, the serine/proline-rich (S/P) region. Our pulse-chase live-cell imaging revealed the slow but steady migration of MAP2 toward distal dendrites, which was not observed in the presence of a Dynein inhibitor. By integrating these experimental parameters into a mathematical model, we demonstrated that an intermittent active transport mechanism by Dynein is sufficient to explain the observed MAP2 dynamics. In addition, our experiments using a Dynein inhibitor as well as Dynein knockdown, showed that impaired Dynein function leads to mislocalization of MAP2 in the axon. Furthermore, we verified that cytoplasmic Dynein binds to MAP2 through the S/P region in heterologous cells. Finally, we show that the failure of this Dynein-mediated transport mechanism results in the ectopic stabilization of axonal microtubules, thereby disrupting axonal identity. Thus, we propose that the cytoplasmic Dynein recruits and transports free MAP2 toward distal dendrites, thereby maintaining the precise dendritic localization of MAP2. Our findings shed light on the previously unknown mechanism behind MAP2 localization and provide a new direction for soluble protein trafficking research in the field of cell biology of neurons.