Paradoxical relationships between active transport and global protein distributions in neurons

Abstract

Neural function depends on continual synthesis and targeted trafficking of intracellular components, including ion channel proteins. Many kinds of ion channels are trafficked over long distances to specific cellular compartments. This raises the question of whether cargo is directed with high specificity during transit or whether cargo is distributed widely and sequestered at specific sites. We addressed this question by experimentally measuring transport and expression densities of Kv4.2, a voltagegated transient potassium channel that exhibits a specific dendritic expression that increases with distance from the soma and little or no functional expression in axons. In over 500 h of quantitative live imaging, we found substantially higher densities of actively transported Kv4.2 subunits in axons as opposed to dendrites. This paradoxical relationship between functional expression and traffic density supports a model-commonly known as the sushi belt model-in which trafficking specificity is relatively low and active sequestration occurs in compartments where cargo is expressed. In further support of this model, we find that kinetics of active transport differs qualitatively between axons and dendrites, with axons exhibiting strong superdiffusivity, whereas dendritic transport resembles a weakly directed random walk, promoting mixing and opportunity for sequestration. Finally, we use our data to constrain a compartmental reaction-diffusion model that can recapitulate the known Kv4.2 density profile. Together, our results show how nontrivial expression patterns can be maintained over long distances with a relatively simple trafficking mechanism and how the hallmarks of a global trafficking mechanism can be revealed in the kinetics and density of cargo.

Transport between im/mobile fractions shapes the speed and profile of cargo distribution in neurons

“…Experimental and computational methods are described in full elsewhere 5 , 23 and are summarized briefly here.…”

“…After 24 h, cells were transferred to neurobasal media containing 2% GlutaMAX and 2% Gibco B-27 supplement, and half the media were replaced every 3 to 4 days for 9 to 13 days until terminal experiments. 23 …”

“…The larger pinhole at 4 Airy units, although noisier, produced a strong signal from a z -plane depth that encompassed the diameter of large dendrites, which was suitable for primary cultures on cover glass. 5 , 23 …”

“…Kv4.2 conducts A-type, voltage-gated transient potassium current abundant in dendrites and has a hypothesized role in dendritic integration. 21 , 22 Kv4.2 is firstly compelling since its transport is not completely understood, 5 , 23 yet its unique subcellular distribution is critical for maintaining cellular excitability. 21 , 22 Further, experiments showing Kv4.2 regulation within dendritic spines suggests a potential dynamism between mobile and immobile states.…”

“… 20 , 26 , 27 , 28 Lastly, in a study of active Kv4.2 transport, our group has identified vesicles containing Kv4.2 that remain static for over hour-long recordings, suggesting the possibility of a prolonged immobile state. 23 …”