TIPsy tour guides: how microtubule plus-end tracking proteins (+TIPs) facilitate axon guidance

Abstract

The growth cone is a dynamic cytoskeletal vehicle, which drives the end of a developing axon. It serves to interpret and navigate through the complex landscape and guidance cues of the early nervous system. The growth cone's distinctive cytoskeletal organization offers a fascinating platform to study how extracellular cues can be translated into mechanical outgrowth and turning behaviors. While many studies of cell motility highlight the importance of actin networks in signaling, adhesion, and propulsion, both seminal and emerging works in the field have highlighted a unique and necessary role for microtubules (MTs) in growth cone navigation. Here, we focus on the role of singular pioneer MTs, which extend into the growth cone periphery and are regulated by a diverse family of microtubule plus-end tracking proteins (+TIPs). These +TIPs accumulate at the dynamic ends of MTs, where they are well-positioned to encounter and respond to key signaling events downstream of guidance receptors, catalyzing immediate changes in microtubule stability and actin cross-talk, that facilitate both axonal outgrowth and turning events.

Keywords: +TIPs; axon guidance; cytoskeleton; growth cone; microtubule dynamics.

Figure 1

The growth cone cytoskeleton. (A) The axon has to travel a complex landscape which presents a multitude of external chemotropic cues (e.g., red and green circles represent diffusible cues, light green represents substrate-bound cues). To properly interpret and navigate these signals, the axon is equipped with a dynamic cytoskeletal vehicle called the growth cone. (B) Structural organization of the growth cone cytoskeleton: Bundled, stable Microtubules (MTs) extend through the axon, entering the growth cone “wrist.” These terminate in the growth cone “central” domain, surrounded by a cage of F-actin arcs. F-actin bundles extend into the growth cone “fingers” (filopodia) to sample the environment. Between the filopodia there exist a series of cortical actin networks which create the lamellipodial veils. A subset of MTs can escape the central domain, to trace along F-actin structures and explore the growth cone periphery. The dynamic plus-ends of MTs are decorated by set of proteins called plus-end tracking proteins (+TIPs) (not shown). Concentration of +TIPs at the dynamic leading edge is particularly important as they come in close contact with signaling cascades that are triggered by external cues.

Figure 2

Phosphorylation dependent +TIPs spatial distribution along MT is key to axon outgrowth and steering. (A) Spatial distribution of CLASP and APC is key to the modulation of MT dynamic behavior and generation of the navigational response. Asymmetrically-distributed guidance signals differentially regulate +TIP localization. On the side of attractive guidance cues, CLASP and APC demonstrate MT plus-end binding and promote axon outgrowth. Faced with repellent cues, APC dissociates from MTs and CLASP shows lattice binding, inhibiting axon outgrowth. (B) Distribution of CLASP is modulated by phosphorylation by kinases. High levels of GSK3 activity lead to two sites of phosphorylation of CLASP, which then dissociates from MTs causing MT destabilization and axon outgrowth inhibition (upper panel). Moderate GSK3 activity promotes a single site of phosphorylation of CLASP, facilitating CLASP plus-end localization and axon growth (middle panel). With low levels of GSK3, CLASP remains unphosphorylated, and CLASP binds to the MT lattice, inhibiting axon growth by coupling MTs to F-actin retrograde flow. A similar phenotype is observed when CLASP is exposed to high levels of Abl activity, which leads to phosphorylation at a different site (lower panel). (C) Phosphorylation-dependent APC distribution. Low GSK3 activity allows APC to remain unphosphorylated, promoting its association with plus-ends, facilitating axon growth (upper panel). However, under high GSK3 activity, phosphorylated APC dissociates from MTs and causes MT destabilization of MTs and axon growth inhibition (lower panel).

Figure 3

Additional +TIPs in axon outgrowth and guidance. A brief synopsis of other presently-identified +TIPs that function in axon outgrowth and guidance, highlighting some of their known interactions: note that a number of these proteins demonstrate an ability to facilitate MT-F-actin coupling.