Anatomy of the axon: Dissecting the role of microtubule plus-end tracking proteins in axons of hippocampal neurons

The question that drove most of the work presented in this thesis is: what are the roles of recently identified microtubule plus-end tracking proteins (+TIPs) in neurodevelopment? The majority of results concerns the development and maintenance of axons of primary hippocampal neurons.
Chapter 1, 3 and 4 prominently feature a number of recently identified +TIPs. To summarize, Chapter 1 reviews the state of the neuronal +TIP field as of early 2016. In Chapter 3, we show that a recently identified SxIP protein regulates axon maturation via a complex autoinhibitory mechanism. It does not track growing microtubule (MT) plus-ends, but rather recruits end-binding proteins (EBs) to actin in neurons. We further characterize this protein in Chapter 4, suggesting that its SxIP motif contributes to binding selectivity, and describe our findings concerning a number of other +TIPs that were screened over the course of this thesis.
Chapter 2 describes how the retrograde cytoplasmic dynein motor and its regulators NDEL1 and LIS1 are responsible for transporting mis-sorted dendritic cargoes out of the axon initial segment (AIS). NDEL1 is immobilized at the AIS cortex via a direct interaction with the master AIS scaffolding protein Ankyrin-G. On the other hand, levels of LIS1 enrichment vary greatly between neurons, the protein exhibits fast turnover times and LIS1 depletion results in reduced levels of NDEL1 bound to Ankyrin-G. We propose that a cortical association between immobilized NDEL1 and the LIS1 may ‘prime’ the quickly diffusing LIS1, perhaps via phosphorylation or a conformational change, resulting in a local pool of primed LIS1 that can intercept and activate dynein.
Although both new functions and new working mechanisms have emerged for neuronal +TIPs, it is clear that the body of knowledge about this class of proteins is far from complete. Prominent neuronal proteins maintain their status as candidate +TIPs, and data hint that essentially unstudied +TIPs may have key functions in establishing neuronal polarity. The challenge remains in dissecting the contribution of EB binding for each of these proteins. This becomes important especially now that roles for +TIPs away from the MT plus-end are starting to challenge the status quo.