Tau Is Enriched on Dynamic Microtubules in the Distal Region of Growing Axons

Abstract

It is widely held that tau determines the stability of microtubules in growing axons, although direct evidence supporting this hypothesis is lacking. Previous studies have shown that the microtubule polymer in the distal axon and growth cone is the most dynamic of growing axons; it turns over more rapidly and is more sensitive to microtubule depolymerizing drugs than the polymer situated proximally. We reasoned that if the stability of axonal microtubules is directly related to their content of tau, then the polymer in the distal axon should have less tau than the polymer in the proximal axon. We tested this proposition by measuring the relative tau content of microtubules along growing axons of cultured sympathetic neurons immunostained for tau and tubulin. Our results show that the tau content of microtubules varies along the axon, but in the opposite way predicted. Specifically, the relative tau content of microtubules increases progressively along the axon to reach a peak near the growth cone that is severalfold greater than that observed proximally. Thus, tau is most enriched on the most dynamic polymer of the axon. We also show that the gradient in tau content of microtubules does not generate corresponding gradients in the extent of tubulin assembly or in the sensitivity of axonal microtubules to nocodazole. On the basis of these findings, we propose that tau in growing axons has functions other than promoting microtubule assembly and stability and that key sites for these functions are the distal axon and growth cone.Key words: microtubule-associated proteins; cytoskeleton; axon growth; quantitative digital image analysis; cultured sympathetic neurons Tau is a developmentally regulated microtubule-associated protein (MAP). Tau is encoded by a single gene, but because of alternative splicing and phosphorylation, it shows multiple isoforms (for review, see Wiche et al., 1991). During neuronal differentiation tau undergoes a transition from immature to mature forms that involves a dramatic increase in the number of isoforms (for review, see Schoenfeld and Obar, 1994). A role for tau in axon growth was initially suggested by studies that demonstrated a temporal correlation between the expression of tau, microtubule (MT) assembly, and axon extension (Drubin et al., 1985). More recently, studies that have altered tau expression either upward or downward have reinforced the view that tau participates in axon growth. Specifically, suppressing tau expression can prevent axon growth, whereas overexpressing tau can promote the elaboration of neurite-like processes that contain arrays of parallel MTs in cells that normally do not elaborate such processes (for review, see Hirokawa, 1994).Although the importance of tau in axon growth is well established, its specific functions are unknown. Because tau binds MTs, its functions presumably involve, at least in part, binding to MTs. On the basis of its ability to promote MT assembly and stability in vitro, tau has been proposed to promote MT assembly and stabilization in ...

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