Messenger RNAs localized to distal projections of human stem cell derived neurons

Abstract

The identification of mRNAs in distal projections of model organisms has led to the discovery of multiple proteins that are locally synthesized for functional roles such as axon guidance, injury signaling and regeneration. The extent to which local protein synthesis is conserved in human neurons is unknown. Here we used compartmentalized microfluidic chambers to characterize the transcriptome of distal projections of human embryonic stem cells differentiated using a protocol which enriched for glutamatergic neurons (hESC-neurons). Using gene expression analysis, we identified mRNAs proportionally enriched in these projections, representing a functionally unique local transcriptome as compared to the human neuronal transcriptome inclusive of somata. Further, we found that the most abundant mRNAs within these hESC-neuron projections were functionally similar to the axonal transcriptome of rat cortical neurons. We confirmed the presence of two well characterized axonal mRNAs in model organisms, β-actin and GAP43, within hESC-neuron projections using multiplexed single molecule RNA-FISH. Additionally, we report the novel finding that oxytocin mRNA localized to these human projections and confirmed its localization using RNA-FISH. This new evaluation of mRNA within human projections provides an important resource for studying local mRNA translation and has the potential to reveal both conserved and unique translation dependent mechanisms.

Nestin in immature embryonic neurons regulates axon growth cone morphology and Semaphorin3a sensitivity

“…Nestin mRNA in neurons would be an indicator that nestin is actually being translated in early neurons. Several paper have described nestin mRNA in neurons in developmental, normal adult, and pathological/injury contexts (Crino & Eberwine 1996;Zhu et al 2011;Kuo et al 2005;Arner et al 2015;Perry et al 2012;Farzanehfar, Lu, et al 2017;Dey et al 2017;Farzanehfar, Horne, et al 2017;Bigler et al 2017), thus nestin expression is not excluded from neurons. Nestin's presence in subpopulations of adult neurons suggests that some neurons may never lose nestin expression, or may regain it later.…”

RACK1 is required for axon guidance and local translation at growth cone point contacts

“…Interestingly, we show that RACK1 is not required for ribosomal localization to growth cones (Figure 3), suggesting RACK1 does not transport ribosomes to growth cones, or there are compensatory mechanisms for when RACK1 is lost. Another possibility is that ribosomes are themselves locally translated in growth cones, which is not unexpected as mRNAs encoding ribosomal subunits have been observed in growth cones (34). Thus, this suggests that RACK1 does not regulate neural development through ribosomal transport.…”

CB1receptor-mediated inhibitory LTD triggers presynaptic remodeling via protein synthesis and ubiquitination

“…We have recently reported that protein synthesis is required for CB1-iLTD (Younts et al, 2016). also be a novel mechanism by which local translation is regulated (Bigler et al, 2017).…”