KIF17, a recently characterized member of the kinesin superfamily proteins, has been proposed to bind in vitro to a protein complex containing mLin10 (Mint1/X11) and the NR2B subunit of the NMDA receptors (NMDARs). In the mammalian brain, NMDARs play an important role in synaptic plasticity, learning, and memory. Here we present, for the first time, the dynamic properties of KIF17 and provide evidence of its function in the transport of NR2B in living mammalian neurons. KIF17 vesicles enter and move specifically along dendrites in a processive way, at an average speed of 0.76 microm/sec. These vesicles are effectively associated with extrasynaptic NR2B, and thus they transport and deliver NR2B subunits in dendrites. However, KIF17 does not seem to enter directly into postsynaptic regions. Cellular knockdown or functional blockade of KIF17 significantly impairs NR2B expression and its synaptic localization. Interestingly, the decrease in the number of synaptic NR2B subunits is followed by a parallel increase in the number of NR2A subunits at synapses. In contrast, upregulation of the expression level of NR2B, after treatment with the NMDAR antagonist D(-)-2-amino-5-phosphonopentanoic acid, simultaneously increases the expression level of KIF17. These observations concerning the downregulation or upregulation of KIF17 and NR2B reveal the probable existence of a shared regulation process between the motor and its cargo. Taken together, these results illustrate the complex mechanisms underlying the active transport and regulation of NR2B by the molecular motor KIF17 in living hippocampal neurons.

Establishment and maintenance of cell structures and functions are highly dependent on the efficient regulation of intracellular transport in which proteins of the kinesin superfamily (KIFs) are very important. In this regard, how KIFs regulate the release of their cargo is a critical process that remains to be elucidated. To address this specific question, we have investigated the mechanism behind the regulation of the KIF17-Mint1 interaction. Here we report that the tail region of the molecular motor KIF17 is regulated by phosphorylation. Using direct visualization of protein-protein interaction by FRET and various in vitro and in vivo approaches we have demonstrated that CaMKII-dependent phosphorylation of KIF17 on Ser 1029 disrupts the KIF17-Mint1 association and results in the release of the transported cargo from its microtubule-based transport.

No abstract

Neuronal differentiation and function require extensive stabilization of the microtubule cytoskeleton. Neurons contain a large proportion of microtubules that resist the cold and depolymerizing drugs and exhibit slow subunit turnover. The origin of this stabilization is unclear. Here we have examined the role of STOP, a calmodulin-regulated protein previously isolated from cold-stable brain microtubules. We find that neuronal cells express increasing levels of STOP and of STOP variants during differentiation. These STOP proteins are associated with a large proportion of microtubules in neuronal cells, and are concentrated on cold-stable, drug-resistant, and long-lived polymers. STOP inhibition abolishes microtubule cold and drug stability in established neurites and impairs neurite formation. Thus, STOP proteins are responsible for microtubule stabilization in neurons, and are apparently required for normal neurite formation.