Activity-Dependent Regulation of Distinct Transport and Cytoskeletal Remodeling Functions of the Dendritic Kinesin KIF21B

Abstract

Summary The dendritic arbor is subject to continual activity-dependent remodeling, requiring a balance between directed cargo trafficking and dynamic restructuring of the underlying microtubule tracks. How cytoskeletal components are able to dynamically regulate these processes to maintain this balance remains largely unknown. By combining single molecule assays and live imaging in rat hippocampal neurons, we have identified the kinesin-4 KIF21B as a molecular regulator of activity-dependent trafficking and microtubule dynamicity in dendrites. We find that KIF21B contributes to the retrograde trafficking of BDNF/TrkB complexes, and also regulates microtubule dynamics through a separable, non-motor microtubule-binding domain. Neuronal activity enhances the motility of KIF21B at the expense of its role in cytoskeletal remodeling, the first example of a kinesin whose function is directly tuned to neuronal activity state. These studies suggest a model in which KIF21B navigates the complex cytoskeletal environment of dendrites by compartmentalizing functions in an activity-dependent manner.

Kinesin-4 KIF21B is a potent microtubule pausing factor

“…Analysis of fluorescence intensity of single KIF21B-MD-CC1-GFP molecules in comparison to monomeric GFP and dimeric EB3-GFP indicated that they were dimers, as expected (Figure 2A, Supplementary file 2). This conclusion was confirmed by two-step photobleaching profiles (Figure 2B) and was in agreement with the published data obtained in HeLa cell lysates with a similar construct (Ghiretti et al, 2016).

10.7554/eLife.24746.006Figure 2.Dimeric motor domain of KIF21B slows down MT polymerization in vitro.( A ) Histograms of fluorescence intensities at the initial moment of observation of single molecules of the indicated proteins immobilized on coverslips (symbols) and the corresponding fits with lognormal distributions (lines).

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“…Unlike its paralogue KIF21A, which is largely diffuse when expressed in similar conditions (van der Vaart et al, 2013), KIF21B bound to MTs and accumulated at their ends at the cell periphery (Figure 1A). Live cell imaging showed that KIF21B processively moves along MTs with an average speed of 0.63 ± 0.22 µm/s (mean±SD) (Figure 1B); this velocity is three times faster than that recently described for HaloTag-labeled KIF21B in neurons (Ghiretti et al, 2016). In internal cell regions, where no clear accumulation of the motor at growing MT ends was observed, the expression of KIF21B led to a ~1.5 fold reduction in the MT growth rate measured with the MT plus-end marker EB3-TagRFP-T (Stepanova et al, 2003; van der Vaart et al, 2013) (Figure 1C).…”

“…KIF21B-MD-CC1 also bound to and walked along MTs with a velocity of 1.23 ± 0.27 µm/s (mean±SD) (Figure 1E), but did not specifically accumulate at MT plus ends (Figure 1F,G). The observed velocity was again ~3 times faster than that observed in neurons (Ghiretti et al, 2016), which might be due to the fact that in neuronal cells kinesins are slowed down by specific MAPs. Expression of KIF21B-MD-CC1 reduced MT growth rate by ~1.6 fold (Figure 1H,I), which is similar to what we previously observed with a comparable deletion mutant of KIF21A (van der Vaart et al, 2013).…”

“…This work showed that mice lacking KIF21B are viable, but display defects in learning and memory, which are likely to be due to several dendritic phenotypes, such as reduced complexity of the dendritic arbor and diminished density of dendritic spines that correlate with defects in synaptic transmission. An even more recent paper showed that KIF21B contributes to activity-dependent regulation of some aspects of retrograde trafficking of brain-derived neurotrophic factor-TrkB complexes in cultured neurons (Ghiretti et al, 2016). Both papers showed that KIF21B can affect MT plus-end dynamics, although the results were complex: while both studies reported an increase in MT growth processivity upon KIF21B loss, MT grew slower in Kif21b knockout neurons, but faster in neurons depleted of KIF21B by RNA interference (Ghiretti et al, 2016; Muhia et al, 2016).…”

“…An even more recent paper showed that KIF21B contributes to activity-dependent regulation of some aspects of retrograde trafficking of brain-derived neurotrophic factor-TrkB complexes in cultured neurons (Ghiretti et al, 2016). Both papers showed that KIF21B can affect MT plus-end dynamics, although the results were complex: while both studies reported an increase in MT growth processivity upon KIF21B loss, MT grew slower in Kif21b knockout neurons, but faster in neurons depleted of KIF21B by RNA interference (Ghiretti et al, 2016; Muhia et al, 2016). In vitro reconstitution work suggested that KIF21B increases MT growth rate and catastrophe frequency, although, surprisingly, the purified protein mostly associated with depolymerizing MT plus ends in these experiments (Ghiretti et al, 2016).…”