Differential Proteomics Reveals Multiple Components in Retrogradely Transported Axoplasm After Nerve Injury

Perlson, Eran; Medzihradszky, Katalin F.; Darula, Zsuzsanna; Munno, David W.; Syed, Naweed I.; Burlingame, Alma L.; Fainzilber, Mike

doi:10.1074/mcp.m400004-mcp200

Cited by 55 publications

(26 citation statements)

References 41 publications

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“…Forty proteins that were retrogradely transported have been identified by two-dimensional PAGE and mass spectrometry in regenerating axons of the snail Lymnea stagnalis. Several of them comprise posttranslationally modified proteins and calpain cleavage products of a 51-kDa intermediate filament protein (254,255). Similar analyses made on purified axons of injury-conditioned dorsal root ganglion neurons led to the identification of cytoskeletal proteins and a large number of other proteins, including heat shock proteins, endoplasmic reticulum proteins, enzymes, and proteins associated with neurodegenerative diseases (330).…”

Section: Vertebrate Axonsmentioning

confidence: 82%

“…) end of the proximal stump of axotomized neurons (174,177) demonstrates the participation of local protein synthesis in axon regeneration. Its physiological significance has been recently elucidated by investigations of regenerating axons in various model systems (75,101,133,134,146,215,254,255,302,330,339; for reviews, see Refs. 23,331).…”

Section: B Nerve Terminalsmentioning

confidence: 99%

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Local Gene Expression in Axons and Nerve Endings: The Glia-Neuron Unit

Giuditta¹,

Chun²,

Eyman³

et al. 2008

Physiological Reviews

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Neurons have complex and often extensively elongated processes. This unique cell morphology raises the problem of how remote neuronal territories are replenished with proteins. For a long time, axonal and presynaptic proteins were thought to be exclusively synthesized in the cell body, which delivered them to peripheral sites by axoplasmic transport. Despite this early belief, protein has been shown to be synthesized in axons and nerve terminals, substantially alleviating the trophic burden of the perikaryon. This observation raised the question of the cellular origin of the peripheral RNAs involved in protein synthesis. The synthesis of these RNAs was initially attributed to the neuron soma almost by default. However, experimental data and theoretical considerations support the alternative view that axonal and presynaptic RNAs are also transcribed in the flanking glial cells and transferred to the axon domain of mature neurons. Altogether, these data suggest that axons and nerve terminals are served by a distinct gene expression system largely independent of the neuron cell body. Such a local system would allow the neuron periphery to respond promptly to environmental stimuli. This view has the theoretical merit of extending to axons and nerve terminals the marginalized concept of a glial supply of RNA (and protein) to the neuron cell body. Most long-term plastic changes requiring de novo gene expression occur in these domains, notably in presynaptic endings, despite their intrinsic lack of transcriptional capacity. This review enlightens novel perspectives on the biology and pathobiology of the neuron by critically reviewing these issues.

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Section: Vertebrate Axonsmentioning

confidence: 82%

Section: B Nerve Terminalsmentioning

confidence: 99%

Local Gene Expression in Axons and Nerve Endings: The Glia-Neuron Unit

Giuditta¹,

Chun²,

Eyman³

et al. 2008

Physiological Reviews

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“…Nerve injury studies must also contend with variations in inflammatory and other responses in injured versus non-injured tissues. Previous studies of axonally transported complexes using proteomics used either affinitytargeted investigations of purified organelles (28 -30) or gel-based differential screens (15,31). Here, we sought to obtain a more comprehensive view of axon transport ensemble changes induced by nerve injury by interrogating ligature axoplasm preparations with iTRAQ quantification and LC-MS/ MS.…”

Section: Fig 3 Pca Analysis For Itraq Datamentioning

confidence: 99%

“…This prominent role for post-transcriptional processes suggests that comprehensive characterization of retrograde signaling will require proteomics approaches. In a previous study we used two-dimensional PAGE and mass spectrometry to analyze retrogradely concentrated axoplasm from injured mollusc nerve, identifying a vesicular ensemble blocked by the lesion and an up-regulated ensemble highly enriched in calpain cleavage products of an intermediate filament (14,15). Follow-up studies in rodent sciatic nerve showed that the mammalian intermediate filament vimentin is produced by local translation of axonal mRNA upon axonal injury and then undergoes calpain-mediated proteolysis, generating a cleavage product that interacts with importins bound to dynein and enables protected retrograde transport of phosphorylated forms of the mitogen-activated protein kinases Erk1 and Erk2 (16,17).…”

mentioning

confidence: 99%

Axonal Transport Proteomics Reveals Mobilization of Translation Machinery to the Lesion Site in Injured Sciatic Nerve

Michaelevski

Medzihradszky

Lynn

et al. 2010

Molecular & Cellular Proteomics

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Peripheral nerve injuries elicit a cascade of axonal responses that are required for a successful regenerative response. The lesioned axons must signal retrogradely to their cell bodies to activate intrinsic neurite outgrowth mechanisms (1-3) and then overcome physical barriers and inhibitory cues in the extracellular environment to achieve functional regeneration (4, 5). The first indications of a breach in axonal integrity upon injury are most likely abnormal generation of action potentials and/or waves of calcium propagating from the lesion site toward the intact portions of the cell (6, 7). At a later stage, signals carried by motor-driven transport systems start to affect the cell body. This phase includes both an interruption of the normal supply of retrogradely transported molecules such as trophic factor signals (8) and arrival of new signals elicited at the injury site (3, 9). The latter interact with a variety of dynein-associated carriers including importins (10, 11) and kinase family scaffolds (12, 13).In mammalian neurons, the distances between axonal lesion sites and the nucleus can reach many centimeters (up to 1 m in humans); hence, retrograde signaling events within the first few hours after injury must be independent of new transcription in the cell body. Proteolysis, local protein synthesis, and post-translational signaling modifications such as phosphorylation have all been implicated in generation of the retrograde signaling ensemble (1). This prominent role for post-transcriptional processes suggests that comprehensive characterization of retrograde signaling will require proteomics approaches. In a previous study we used two-dimensional PAGE and mass spectrometry to analyze retrogradely concentrated axoplasm from injured mollusc nerve, identifying a vesicular ensemble blocked by the lesion and an up-regulated ensemble highly enriched in calpain cleavage products of an intermediate filament (14,15). Follow-up studies in rodent sciatic nerve showed that the mammalian intermediate filament vimentin is produced by local translation of axonal mRNA upon axonal injury and then undergoes calpain-mediated proteolysis, generating a cleavage product that interacts with importins bound to dynein and enables protected retrograde transport of phosphorylated forms of the mitogen-activated protein kinases Erk1 and Erk2 (16,17). Here, we extend our efforts to determine the components of the retrograde injury signaling ensemble in lesioned nerve by using LC-MS/MS coupled with iTRAQ TM1 labeling to directly analyze mammalian axoplasm samples after nerve injury. The analyses reveal extensive changes in both anterograde and Research

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“…However, other proteins that might be carried by importin-dynein complexes were not identified in those investigations. Retrograde transport of ~30 proteins was demonstrated in cultured frog nerves by Edbladh et al (Edbladh et al, 1994), and of >100 proteins in lesioned nerves of the mollusc Lymnia (Perlson et al, 2004), although the identities and possible roles of those proteins in axonal regeneration are largely unknown.…”

Section: Retrograde Signallingmentioning

confidence: 99%

Enhancement of axonal regeneration by in vitro conditioning and its inhibition by cyclopentenone prostaglandins

Tonge

Chan

Zhu

et al. 2008

Journal of Cell Science

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Axonal regeneration is enhanced by the prior `conditioning' of peripheral nerve lesions. Here we show that Xenopus dorsal root ganglia (DRG) with attached peripheral nerves (PN-DRG) can be conditioned in vitro, thereafter showing enhanced neurotrophin-induced axonal growth similar to preparations conditioned by axotomy in vivo. Actinomycin D inhibits axonal outgrowth from freshly dissected PN-DRG, but not from conditioned preparations. Synthesis of mRNAs that encode proteins necessary for axonal elongation might therefore occur during the conditioning period, a suggestion that was confirmed by oligonucleotide microarray analysis. Culturing PN-DRG in a compartmentalized system showed that inhibition of protein synthesis (but not RNA synthesis) in the distal nerve impaired the conditioning response, suggesting that changes in gene expression in cultured DRG depend on the synthesis and retrograde transport of protein(s) in peripheral nerves. The culture system was also used to demonstrate retrograde axonal transport of several proteins, including thioredoxin (Trx). Cyclopentenone prostaglandins, which react with Trx, blocked the in vitro conditioning effect, whereas inhibition of other signalling pathways thought to be involved in axonal regeneration did not. This suggests that Trx and/or other targets of these electrophilic prostaglandins regulate axonal regeneration. Consistent with this hypothesis, morpholino-induced suppression of Trx expression in dissociated DRG neurons was associated with reduced neurite outgrowth.

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Differential Proteomics Reveals Multiple Components in Retrogradely Transported Axoplasm After Nerve Injury

Cited by 55 publications

References 41 publications

Local Gene Expression in Axons and Nerve Endings: The Glia-Neuron Unit

Local Gene Expression in Axons and Nerve Endings: The Glia-Neuron Unit

Axonal Transport Proteomics Reveals Mobilization of Translation Machinery to the Lesion Site in Injured Sciatic Nerve

Enhancement of axonal regeneration by in vitro conditioning and its inhibition by cyclopentenone prostaglandins

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