Golgi compaction facilitates microtubule nucleation to drive adult vertebrate peripheral neuron regeneration

Background

Regeneration of adult vertebrate neurons following a peripheral neuronal injury (PNI) encompasses substantial axon extension to reestablish function. Existing clinical treatments rely on surgical repair, which is inadequate in attaining a consequential recovery. Comprehensive analysis of molecular mechanism supporting peripheral neuronal regeneration is both timely and needed to fast-track therapeutic interventions.

Although neurons lack a conventional centrosome for microtubule organization, this study introduces the Golgi apparatus as a key player. The authors reveal that peripheral neuron injury is successively followed by the rapid fragmentation and compaction of the Golgi which facilitates microtubule nucleation, indispensable for the recruitment of AKAP9 and γ-tubulin that drive axon elongation and regeneration process.

Key findings of the study

(a) Golgi revealed as key microtubule organizer in neuron regeneration. Rat dorsal root ganglion (DRG) neurons were obtained post peripheral neuronal injury, cultured and assessed by high-resolution STED microscopy in live and fixed samples. The initiation of axon regeneration was observed 24-30 hours post-injury, overlapping with Golgi compaction. Further examination looking at various morphological hallmarks of axonal regeneration revealed that cells with retraction bulbs (indicative of acute injury) had fragmented Golgi, while cells with multiple extensions or dominant axons had compacted Golgi. Microtubule comets emerging from Golgi to axons were observed at 36 hours, after which somatic microtubules appeared around 48 hours post injury. These observations were made in both in vivo rat models as well as human DRG neurons.

(b) Golgi mediated microtubule nucleation. Golgi compaction post injury caused these organelles to become fundamental sites for  microtubule nucleation to be recruit factors AKAP9 and γ-tubulin. AKAP9 was found to associated with Golgi from 2 h to 7 days post injury and started showing a prominent association with the Golgi from 16h onwards. In contrast, γ-tubulin association with the Golgi peaks at 24 and 48 h post injury.

(c) Golgi integrity is indispensable for microtubule nucleation and axon regeneration. Brefeldin A (BFA) was added 23 hours following peripheral neuron injury to disrupt the compacted Golgi. The authors observed a significant reduction in AKAP9 and γ-tubulin recruitment to the Golgi, resulting in decreased microtubule polymerization and impaired axon regeneration. In addition, Gatastatin G2 induced γ-tubulin inhibition hindered axon growth, reiterating that an integral Golgi structure is vital for microtubule nucleation and axon growth.

(d) AKAP9 association with the Golgi is vital for γ-tubulin localization. AKAP9 displacement from the Golgi using CRISPR knockouts disrupted γ-tubulin localization and weakened microtubule nucleation, underlining the importance of sequentially recruiting AKAP9 and γ-tubulin to the Golgi for effective axon regeneration.

Significance of the study

The authors have successfully highlighted sequence of events for axon regeneration following peripheral nerve injury, conserved across species following peripheral nerve injury, conserved across species. This study beautifully demonstrates Golgi fragmentation as a consequence of acute peripheral nerve injury, which afterwards is compacted as a key step in the regeneration, recruiting the scaffolding protein AKAP9, which in turn recruits γ-tubulin to the Golgi initiating microtubule nucleation (Figure 1).  These findings have noteworthy clinical relevance as the mechanism is conserved between rats and humans. The authors suggest that therapeutic strategies directed at enhancing axon regeneration through Golgi compaction could improve existing clinical outcomes. Finally, understanding how molecular and cellular mechanisms underlying Golgi structure and function are linked to neuronal regeneration paves the way for new research approaches with the potential to mitigate neurodegenerative disorders.

What do I like most about the preprint?

What to me stands out most about this study is that the Golgi apparatus’s role in peripheral nerve regeneration is conserved across species, from rats to humans, emphasizing its potential clinical relevance. The temporal dynamics of the Golgi structure and its relevance in axon regulation are exceptionally compelling. Furthermore, targeting of the Golgi dynamics in therapeutic interventions opens new avenues for improving outcomes when treating neurodegenerative disease. Altogether, the combination of a comprehensive molecular mechanism and practical implications makes this study a noteworthy contributor to our understanding of nerve regeneration and repair.

 Open questions:

(1) Does Golgi-mediated protein trafficking contribute to peripheral neuronal regeneration?

(2) Are there similarities or differences in sensory vs. motor peripheral neurons to post-injury and Golgi dynamics?

(3) As discussed in the study extracellular cues possibly impact Golgi compaction post-injury, could integrin-mediated signaling have a possible role?

Tags: axon regeneration, golgi

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