In vivo time-lapse imaging revealed that EB1 comets emerge from approximately 45% of dendritic Golgi outposts ( Figure 1). To confirm that these Golgi outposts can nucleate MTs, the authors used an in vitro assay where purified Golgi outposts were collected and incubated with purified α- and β-tubulin dimers and GTP. Indeed, MTs formed on Golgi outposts

that contained γ-tubulin and CP309 (the Drosophila homolog of AKAP450) but did not form on any Golgi outpost that lacked γ-tubulin. To determine the necessity of γ-tubulin, a function-blocking γ-tubulin antibody was incubated with the purified Golgi outposts prior to addition of γ-tubulin and GTP; as expected, no MTs could be nucleated. To resolve the importance of γ-tubulin and CP309/AKAP450 in vivo, Ori-McKenney et al. (2012) made use CB-839 chemical structure of two Drosophila mutants where these genes are inactivated. In these mutants, Golgi outposts are still localized to branch points and throughout the dendritic arbor, but mutant neurons show a striking decrease in the number of EB1 comets nucleating http://www.selleckchem.com/products/lgk-974.html from the outposts located in the terminal

branches. To establish the role of Golgi-associated acentrosomal MT nucleation during dendritic arborization, the authors used the two mutants mentioned above and performed Scholl analysis which allows a quantitative assessment of the effect on dendritic branching as a function of distance from the cell body. Remarkably, the primary and secondary branches formed properly but a drastic reduction in the number Sodium butyrate of terminal branches occurred in γ-tubulin and CP309 mutant neurons, leading to a significant simplification of dendritic arborization. To understand why terminal branches were more specifically affected in these mutants, the authors compared distal branches with or without EB1 comet formation in vivo and determined that EB1 comet formation correlated with branch growth or stability, whereas the lack of comet formation correlated with a high probability of branch retraction.

This suggested that Golgi outpost-associated acentrosomal MT nucleation is critical for terminal branch stabilization. The authors confirmed that in the γ-tubulin and CP309 mutant neurons, significantly fewer EB1 comets entered terminal branches and the majority of terminal branches retracted. These results provide compelling evidence defining the critical role of Golgi outpost-associated acentrosomal MT nucleation during dendritic morphogenesis. The authors propose a likely scenario where all three modes of acentrosomal MT nucleation are involved in proper formation and maintenance of dendritic morphology. This work highlights potential differences between axonal and dendritic morphogenesis because Golgi outposts are not present from the axon (Horton et al., 2005). These results also beg the question of whether or not acentrosomal MT nucleation plays an important role in dendritic morphogenesis of vertebrate neurons.