Feline Lindhout

Quantitative mapping of transcriptome and proteome dynamics during polarization of human iPSC-derived neurons 2 49 proteins, including AnkG and Trim46, cooperate to form a stable AIS-microtubule structure in the proximal axon (Freal et al. 2019). It has been shown that AnkG can act as a scaffold to recruit Trim46-positive microtubules and subsequently direct AIS protein trafficking to the proximal axon (Freal et al. 2019). The possible function of AIS proteins in the distal axon remains elusive. Similar to their function in stabilizing microtubules in the proximal axon following proper AIS formation, redistribution of AIS proteins in the distal axon may help to locally stabilize microtubules and drive axon outgrowth. AnkG in distal axons may provide additional support to the formation of Trim46-positive parallel microtubules during outgrowth. Then, as Trim46 moves proximally, the axonal microtubule network close to the cell body is remodeled to the characteristic uniform plus-end out orientation. This idea is consistent with the observed shift of the unidirectional parallel microtubule organization in the distal parts of the axon first, followed by proximal reorganization. Finally, AnkG and Trim46 together may drive AIS assembly at the proximal axon, as previously described in dissociated rodent neurons (Freal et al. 2019). It remains unknown if the observed intermediate step of distal AIS protein accumulation is unique to human neurons. Axons in humans grow significantly longer compared to rodents, thus additional regulatory mechanisms enhancing axon outgrowth might be at play in humans. Alternatively, it is possible that these changes have not been observed in rodent neurons because of their relatively faster development. We also observed differences in microtubule dynamics, as MT+TIP growth speeds are higher than those found in rodent neurons in vitro and in vivo , hinting at species-specific regulation of microtubule dynamics in human neurons (Stepanova et al. 2003; Kleele et al. 2014; Yau et al. 2016). Future studies are required to examine the potential human-specific attributes of neuronal polarity and axon outgrowth. In summary, our quantitative map of neuronal transcriptome and proteome dynamics provides a rich resource for future analysis of early neurodevelopmental processes in human iPSC-derived neurons. We investigated early development in human neurons and uncovered an intermediate axon developmental step, thereby illustrating the potential of this model system to study neurobiological processes in human cells in stage 3 neurons. This study also provides a framework and excellent starting point for further studies that aim to complement our understanding of neuronal polarization in human cells. ACKNOWLEDGEMENTS We thank Dr. Didier Trono for the lentiviral vector, and Nicky Scheefhals for critically reading the manuscript. This work was supported by the Netherlands Organization for Scientific Research (NWO-ALW-VICI, 865.10.010, CCH, the Netherlands Organization for Health Research and Development (ZonMW-TOP, 912.16.058, CCH), the European Research Council (ERC) (ERC-consolidator, 617050, CCH), and the research program of the Foundation for Fundamental Research on Matter (FOM, #16NEPH05, CJW).