No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
The Mammalian-Specific Protein Armcx1 Regulates Mitochondrial Transport during Axon Regeneration
... Depletion of either MIRO1 or MIRO2 strikingly decreased the number of TOM + MDVs, a phenotype that was even more apparent by their simultaneous depletion (Fig. 4e,f and Extended Data Fig. 4g). In addition, the TOMM20 + -MDV proteome included the MIRO1/2 binding partners CENP-F and ARMCXs (Fig. 2a), which are reported to modulate MIRO1/2-dependent mitochondrial motility and thus mitochondrial segregation, axon regeneration as well as neuronal maturation and survival [34][35][36][37][38] . Providing support for a role in MDV formation, the short interfering RNA (siRNA)-mediated knockdown of CENP-F or double knockdown of ARMCX1 and ARMCX3 decreased the number of TOM + MDVs, whereas single knockdown of either ARMCX1 or ARMCX3 had no significant effect (Fig. 4g,h and Extended Data Fig. 4h,i). ...
... Finally, these insights will provide a framework to dissect the functional contribution of MDV-mediated QC in neurodegenerative disorders using various loss-of-function and disease models of DRP1, MIRO1/2 and ARMCXs, where the interpretation of mutant phenotypes similar to those seen following the loss of MIROs must now be expanded from roles in fission, motility or mitophagy to include their core functions in MDV formation 32,33,37,38,[72][73][74][75] . Together, this study provides an excellent base for future analyses of MDVs to solve broader mechanistic questions of cargo selection, integration of MDVs into the endolysosomal pathway and differential regulation of MDV scission versus organelle division, including the roles of organellar contact sites. ...
... Unprocessed blots and numerical source data, including exact P values and test statistics, are provided.and CENP-F in the MDV proteome[33][34][35][36][37] ; second, by the dependency of MDV formation on these components or microtubules itself; and third, through the direct observation that TOM + -MDV biogenesis occurs at the tip of thin mitochondrial protrusions via scission events driven by DRP1, phosphatidic acid enrichment and the receptors MID49, MID51 and MFF. ...
Mitochondrial-derived vesicles (MDVs) are implicated in diverse physiological processes—for example, mitochondrial quality control—and are linked to various neurodegenerative diseases. However, their specific cargo composition and complex molecular biogenesis are still unknown. Here we report the proteome and lipidome of steady-state TOMM20⁺ MDVs. We identified 107 high-confidence MDV cargoes, which include all β-barrel proteins and the TOM import complex. MDV cargoes are delivered as fully assembled complexes to lysosomes, thus representing a selective mitochondrial quality control mechanism for multi-subunit complexes, including the TOM machinery. Moreover, we define key biogenesis steps of phosphatidic acid-enriched MDVs starting with the MIRO1/2-dependent formation of thin membrane protrusions pulled along microtubule filaments, followed by MID49/MID51/MFF-dependent recruitment of the dynamin family GTPase DRP1 and finally DRP1-dependent scission. In summary, we define the function of MDVs in mitochondrial quality control and present a mechanistic model for global GTPase-driven MDV biogenesis.
... Studies into mitochondrial positioning and axon regeneration have identified a number of key regulatory molecules, including Armcx1 and syntaphilin. Armcx1 was identified after comparative gene expression profiling of non-regenerative and highly regenerative RGCs [133]. Overexpression of Armcx1, a mitochondria resident protein, resulted in increased mobility of mitochondria, which in turn promoted neurite outgrowth and survival not only in cultured embryonic cortical neurons but also in adult RGCs after optic nerve crush [133]. ...
... Armcx1 was identified after comparative gene expression profiling of non-regenerative and highly regenerative RGCs [133]. Overexpression of Armcx1, a mitochondria resident protein, resulted in increased mobility of mitochondria, which in turn promoted neurite outgrowth and survival not only in cultured embryonic cortical neurons but also in adult RGCs after optic nerve crush [133]. Interestingly, when Armcx1 is knocked out in a highly regenerative genetic model, both axon regeneration and cell survival are inhibited [133]. ...
... Overexpression of Armcx1, a mitochondria resident protein, resulted in increased mobility of mitochondria, which in turn promoted neurite outgrowth and survival not only in cultured embryonic cortical neurons but also in adult RGCs after optic nerve crush [133]. Interestingly, when Armcx1 is knocked out in a highly regenerative genetic model, both axon regeneration and cell survival are inhibited [133]. In a different study, mitochondria were found to be less mobile with maturation which was attributed to increased expression of mitochondrial anchor protein-syntaphilin and correlated with reduced regenerative capacity [118]. ...
Investigating the molecular mechanisms governing developmental axon growth has been a useful approach for identifying new strategies for boosting axon regeneration after injury, with the goal of treating debilitating conditions such as spinal cord injury and vision loss. The picture emerging is that various axonal organelles are important centers for organizing the molecular mechanisms and machinery required for growth cone development and axon extension, and these have recently been targeted to stimulate robust regeneration in the injured adult central nervous system (CNS). This review summarizes recent literature highlighting a central role for organelles such as recycling endosomes, the endoplasmic reticulum, mitochondria, lysosomes, autophagosomes and the proteasome in developmental axon growth, and describes how these organelles can be targeted to promote axon regeneration after injury to the adult CNS. This review also examines the connections between these organelles in developing and regenerating axons, and finally discusses the molecular mechanisms within the axon that are required for successful axon growth.
... Myc-tagged overexpressed ARMCX1 in HEK293 cells co-localises with the MitoTracker dye staining for mitochondria [3]. The first 40 residues are necessary for ARMCX1-HA-tagged to co-localise with MitoDsRed marker in mouse cortical neurons [21]. ...
... Indeed, SFB-ARMC10 and ARMC10-GFP were not observed in the nuclei of HEK293 and QSG-7701 cells, respectively [22;4]. Similarly ARMCX1 was not observed in the nuclei of mouse cortical neurons [21]. ARMCX3 has been clearly localised in the nucleus by several studies. ...
... MIRO1 in complex with TRAK2 interacts with ARMC10 [5], ARMCX3 [3] and ARMCX1 ( [21]; Table S1). The interaction, at least for ARMCX3, is lost at high calcium concentration (2mM) and a mutant MIRO1-myc-tagged lacking its two EF-Hands calcium binding motifs still co-immunoprecipitates with ARMCX3-GFP but this interaction becomes insensitive to intracellular calcium level, thus confirming a regulation by calcium ions [3]. ...
GPRASP (GPCR-associated sorting protein)/ARMCX (ARMadillo repeat-Containing proteins on the X chromosome) family is composed of 10 proteins, which genes are located on a small locus of the X chromosome except one. They possess at least two armadillo-like repeats on their carboxyl-terminal homologous sequence, but they can be subdivided on specific sequence features. Subfamily 1 (GPRASP1, GPRASP2, GPRASP3, ARMCX4 and ARMCX5) displays additional repeated motifs while a mitochondrial targeting transmembrane domain is present in subfamily 2 (ARMC10, ARMCX1, ARMCX2, ARMCX3 and ARMCX6). Although their roles are not yet fully understood, the recent identification of several interacting partners have shed new light on the processes in which GPRASP/ARMCX proteins are implicated. Among the interacting partners of proteins from subfamily 1, many are GPCRs. GPRASP1 binds trafficking proteins such as Beclin2 and the Dysbindin-HRS-Gas complex to participate in GPCR post-endocytic sorting. Moreover, in vitro as well as in vivo experiments indicate that GPRASP1 is a critical player in the adaptive responses related to chronic treatments with GPCR agonists. GPRASP2 seems to play a key role in the signalling of the hedgehog pathway in the primary cilium through a Smoothened-GPRASP2-Pifo complex. Identified small compound inhibitors of this complex could treat drug-resistant Smoothened derived cancer forms. Deletion of GPRASP2 in mice causes neurodevelopmental alteration and affects mGluR5 regulation, reflected by autism-like behaviour. Several members of subfamily 2, in complex with TRAK2 and MIRO, are involved in the trafficking of mitochondria in axons and on the regulation of their size and division, influencing the cell cycle. The essential role of GPRASP/ARMCX proteins in the cellular physiology is supported by human cases of deletions, causing male neonatal lethality by pulmonary delayed development, dysmorphic face and psychiatric and intellectual impacts in females.
... Precise regulation of mitochondrial transport during axon regeneration and axon extension has begun to be elucidated. Overexpression of armadillo repeat-containing X-linked 1 (Armcx1) in adult mice increases mitochondrial transport and enhances adult RGC survival and axon regeneration [106]. Phosphatase and tensin homolog (PTEN) deletion shows an elevated mammalian target of rapamycin (mTOR) activity. ...
... Phosphatase and tensin homolog (PTEN) deletion shows an elevated mammalian target of rapamycin (mTOR) activity. Armcx1 overexpression under PI3K activated conditions using PTEN deficient mice significantly increases RGC survival and axon regeneration, which depend on mitochondrial localization [106]. Impaired mitochondrial functions by OGD can be restored through the activation of the PI3K-Akt pathway and the inhibition of excessive Ca 2+ influx in NSCs [107]. ...
... In the CNS, genetic activation of the PI3K-Akt-mTOR pathway, through deficiencies in upstream negative regulators (i.e., phosphatase, PTEN, hamartin (tuberous sclerosis complex 1 (TSC1)) and tuberin (TSC2)), increases the axon regeneration capacity [118,119]. In PTEN-deleted RGC, overexpression of Armcx1 induces a substantial upregulation in the number of regenerating axons in comparison with PTEN deficiency alone [106]. ...
Regeneration of adult neural circuits after an injury is limited in the central nervous system (CNS). Heme oxygenase (HO) is an enzyme that produces HO metabolites, such as carbon monoxide (CO), biliverdin and iron by heme degradation. CO may act as a biological signal transduction effector in CNS regeneration by stimulating neuronal intrinsic and extrinsic mechanisms as well as mitochondrial biogenesis. CO may give directions by which the injured neurovascular system switches into regeneration mode by stimulating endogenous neural stem cells and endothelial cells to produce neurons and vessels capable of replacing injured neurons and vessels in the CNS. The present review discusses the regenerative potential of CO in acute and chronic neuroinflammatory diseases of the CNS, such as stroke, traumatic brain injury, multiple sclerosis and Alzheimer’s disease and the role of signaling pathways and neurotrophic factors. CO-mediated facilitation of cellular communications may boost regeneration, consequently forming functional adult neural circuits in CNS injury.
... The transparent body of C. elegans further allows analysis of such phenotypes, including axon regrowth rate, guidance, and fusion at a single neuron resolution in vivo with fluorescence microscopy. Past nerve regeneration studies in C. elegans have led to the discovery of many conserved nerve regeneration pathways [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26] , notably the dual leucine zipper kinase (DLK-1/p38) signaling cascade, whose role in neural development, regeneration, and degeneration has been confirmed in numerous vertebrate and invertebrate species 7, 27-31 . ...
... Through these steps, we aimed to identify TFs with significantly higher binding affinity to promoter regions of the identified DEGs using all genes expressed in neuron as a benchmark. Statistical analysis was performed by Gene Set Enrichment Analysis (GSEA) Pre-ranked module 22 . As the first input to GSEA Pre-ranked module, we generated a list of all genes expressed in neurons ranked by their Z statistics (generated during differential expression analysis, assigned a Z statistics of 0 if not a DEG). ...
Our understanding of nerve regeneration can be enhanced by delineating its underlying molecular activities at single neuron resolution in small model organisms such as Caenorhabditis elegans. Existing cell isolation techniques cannot isolate regenerating neurons from the nematode. We present femtosecond laser microdissection (fs-LM), a new single cell isolation method that dissects intact cells directly from living tissue by leveraging the micron-scale precision of fs-laser ablation. We show that fs-LM facilitated sensitive and specific gene expression profiling by single cell RNA-sequencing, while mitigating the stress related transcriptional artifacts induced by tissue dissociation. Single cell RNA-sequencing of fs-LM isolated regenerating C. elegans neurons revealed transcriptional program leading to successful regeneration in wild-type animals or regeneration failure in animals lacking DLK-1/p38 kinase. The ability of fs-LM to isolate specific neurons based on phenotype of interest allowed us to study the molecular basis of regeneration heterogeneity displayed by neurons of the same type. We identified gene modules whose expression patterns were correlated with axon regrowth rate at a single neuron level. Our results establish fs-LM as a highly specific single cell isolation method ideal for precision and phenotype-driven studies.
... In order to supply the amount of ATP energy to the growth cone of regenerating axons, mitochondria in the soma are actively transported into regenerating axons, and axonal proteins Aramcx1, DLK, and syntaphilin regulate mitochondrial transport by interacting with basic motility proteins (Zhou et al., 2016;Cartoni et al., 2016;Han et al., 2016). What would be the possible function of mitochondrial phospho-STAT3 in relation to axonal regeneration? ...
... It should however be noted that both mitochondrial and nuclear STAT3 activities contribute to enhanced axonal regeneration in retinal ganglion cells (Luo et al., 2016a). While our data strongly suggest the growthpromoting activity of mitochondrial phospho-STAT3 for axonal regeneration (Cartoni et al., 2016;Han et al., 2016), a possible function of STAT3 as a nuclear factor cannot be excluded. Since phospho-STAT3 at S727 is required for maximal transcriptional activity of Y705phosphorylated STAT3 (Wen et al., 1995), some of phospho-STAT3 (S727) proteins may be retrogradely transported into the cell body for transcriptional activation of target genes (Ben-Yaakov et al., 2012). ...
Cyclin-dependent kinase 5 (Cdk5) is involved in neural organization and synaptic functions in developing and adult brains, yet its role in axonal regeneration is not known well. Here, we characterize Cdk5 function for axonal regeneration after peripheral nerve injury. Levels of Cdk5 and p25 were elevated in sciatic nerve axons after injury. Cdk5 activity was concomitantly induced from injured nerve and increased the phosphorylation of signal transducer and activator of transcription 3 (STAT3) on the serine 727 residue. Pharmacological and genetic blockades of Cdk5 activity phosphorylating STAT3 resulted in the inhibition of axonal regeneration as evidenced by reduction of retrograde labeling of dorsal root ganglion (DRG) sensory neurons and spinal motor neurons and also of neurite outgrowth of preconditioned DRG neurons in culture. Cdk5 and STAT3 were found in mitochondrial membranes of the injured sciatic nerve. Cdk5-GFP, which was translocated into the mitochondria by the mitochondrial target sequence (MTS), induced STAT3 phosphorylation in transfected DRG neurons and was sufficient to induce neurite outgrowth. In the mitochondria, Cdk5 activity was positively correlated with increased mitochondrial membrane potential as measured by fluorescence intensity of JC-1 aggregates. Our data suggest that Cdk5 may play a role in modulating mitochondrial activity through STAT3 phosphorylation, thereby promoting axonal regeneration.
... Accordingly, enhanced mitochondrial transport contributes to energy recovery from injuries and promotes axon regeneration and neuronal survival. 56,57 However, it is still unclear whether improved mitochondrial quality in axons rescues axonal injuries merely by enhancing energy production or alternatively by repairing damaged mitochondria through mitochondrial fusion ( Figure 1A). Intriguingly, both mitochondrial fusion and fission proteins could interact with mitochondrial transport system. ...
... 57 Armcx1 overexpression promotes mitochondrial transport and neurite outgrowth in cortical neurons ( Figure 3B) and, more importantly, promotes neuronal survival and axon regeneration after optic nerve injury in vivo. 57 Taken together, mitochondrial transport resolves bioenergetic crisis in injured axons by replenishing axonal mitochondria, suggesting that modulating mitochondrial transport might provide a novel strategy to axonal regeneration in TAI. ...
Axonal mitochondrial quality is essential for neuronal health and functions. Compromised mitochondrial quality, reflected by loss of membrane potential, collapse of ATP production, abnormal morphology, burst of reactive oxygen species generation, and impaired Ca²⁺ buffering capacity, can alter mitochondrial transport. Mitochondrial transport in turn maintains axonal mitochondrial homeostasis in several ways. Newly generated mitochondria are anterogradely transported along with axon from soma to replenish axonal mitochondrial pool, while damaged mitochondria undergo retrograde transport for repair or degradation. Besides, mitochondria are also arrested in axon to quarantine damages locally. Accumulating evidence suggests abnormal mitochondrial transport leads to mitochondrial dysfunction and axon degeneration in a variety of neurological and psychiatric disorders. Further investigations into the details of this process would help to extend our understanding of various neurological diseases and shed light on the corresponding therapies.
... Upregulation of Armcx1 was reported to promote neuronal survival and repair of injured axons after optic nerve injury via augmenting mitochondrial trafficking in mature retinal ganglion cells, dependent on its mitochondrial targeting sequence. In contrast, the knockdown of Armcx1 exacerbated axonal lesions and the death of neurons (Cartoni et al., 2017). The proofs above indicate that Armcx1 regulates mitochondrial transport during neuronal repair. ...
Ensuring mitochondrial quality is essential for maintaining neuronal homeostasis, and mitochondrial transport plays a vital role in mitochondrial quality control. In this review, we first provide an overview of neuronal mitochondrial transport, followed by a detailed description of the various motors and adaptors associated with the anterograde and retrograde transport of mitochondria. Subsequently, we review the modest evidence involving mitochondrial transport mechanisms that has surfaced in acute neurological disorders, including traumatic brain injury, spinal cord injury, spontaneous intracerebral hemorrhage, and ischemic stroke. An in-depth study of this area will help deepen our understanding of the mechanisms underlying the development of various acute neurological disorders and ultimately improve therapeutic options.
... Firstly, because axon r e g e n e r a t i o n i s a h i g h l y e n e r g ydemanding process, the transfer of monocarboxylates from denervated SCs into proximal regenerating axon stumps could support axonal mitochondrial respiration and ATP production requisite for axon elongation (Figure 2a). Indeed, these energetic features combined with the striking mitochondrial redistribution into growing proximal axon stumps, have been identified as key determinants for successful axon regeneration (Cartoni et al., 2016;Han et al., 2016;Zhou et al., 2016). Secondly, the glial shuttling of energetic substrates into proximal axon stumps is expected to support neuronal survival and fitness through the ATP-dependent retrograde transport of neurotrophin-containing signaling endosomes. ...
... Across multiple populations, ARMCX3 (ALEX3) and the RNA anti-sense gene ARMCX3-AS1 were associated with apnea-hypopnea triggered intermittent hypoxia. ARMCX3 regulates mitochondrial aggregation and trafficking in multiple tissues and facilitates neuronal survival and axon regeneration [63][64][65]. Wnt signaling regulates reactive oxygen species (ROS) generation and ARMCX3-associated mitochondrial aggregation [64,66]. Potential mechanisms for further study include sensitized carotid body chemoreflexes, interaction with inflammatory mechanisms, and neuronal dysfunction within respiratory centers. ...
Background
Sleep-disordered breathing is a common disorder associated with significant morbidity. The genetic architecture of sleep-disordered breathing remains poorly understood. Through the NHLBI Trans-Omics for Precision Medicine (TOPMed) program, we performed the first whole-genome sequence analysis of sleep-disordered breathing.
Methods
The study sample was comprised of 7988 individuals of diverse ancestry. Common-variant and pathway analyses included an additional 13,257 individuals. We examined five complementary traits describing different aspects of sleep-disordered breathing: the apnea-hypopnea index, average oxyhemoglobin desaturation per event, average and minimum oxyhemoglobin saturation across the sleep episode, and the percentage of sleep with oxyhemoglobin saturation < 90%. We adjusted for age, sex, BMI, study, and family structure using MMSKAT and EMMAX mixed linear model approaches. Additional bioinformatics analyses were performed with MetaXcan, GIGSEA, and ReMap.
Results
We identified a multi-ethnic set-based rare-variant association ( p = 3.48 × 10 ⁻⁸ ) on chromosome X with ARMCX3 . Additional rare-variant associations include ARMCX3-AS1 , MRPS33 , and C16orf90 . Novel common-variant loci were identified in the NRG1 and SLC45A2 regions, and previously associated loci in the IL18RAP and ATP2B4 regions were associated with novel phenotypes. Transcription factor binding site enrichment identified associations with genes implicated with respiratory and craniofacial traits. Additional analyses identified significantly associated pathways.
Conclusions
We have identified the first gene-based rare-variant associations with objectively measured sleep-disordered breathing traits. Our results increase the understanding of the genetic architecture of sleep-disordered breathing and highlight associations in genes that modulate lung development, inflammation, respiratory rhythmogenesis, and HIF1A -mediated hypoxic response.
... A recent publication (Kalinski et al. 2019) reported that HDAC6 targets Miro1 to deacetylate it on lysine 105, blocking mitochondrial transport and attenuating axon growth in mouse neuronal cells. Inhibition or depletion of HDAC6 protected axons from mitochondrial damage, as mitochondrial respiration and transport are important for axonal growth (Cartoni et al. 2016;Lewis 2016). On the other hand, increasing the level of acetyl-Miro1 resulted in enhanced axon growth due to sustained mitochondrial transport (Kalinski et al. 2019). ...
Mitochondria are organelles present in most eukaryotic cells, where they play major and multifaceted roles. The classical notion of the main mitochondrial function as the powerhouse of the cell per se has been complemented by recent discoveries pointing to mitochondria as organelles affecting a number of other auxiliary processes. They go beyond the classical energy provision via acting as a relay point of many catabolic and anabolic processes, to signaling pathways critically affecting cell growth by their implication in de novo pyrimidine synthesis. These additional roles further underscore the importance of mitochondrial homeostasis in various tissues, where its deregulation promotes a number of pathologies. While it has long been known that mitochondria can move within a cell to sites where they are needed, recent research has uncovered that mitochondria can also move between cells. While this intriguing field of research is only emerging, it is clear that mobilization of mitochondria requires a complex apparatus that critically involves mitochondrial proteins of the Miro family, whose role goes beyond the mitochondrial transfer, as will be covered in this review.
... 59 In adult retinal ganglion cells, expressing mitochondrial protein Armcx1 promotes axon regeneration by recruiting mitochondria into motile pools. 60 In mouse DRG neurons, HDAC6mediated deacetylation of Miro-1 inhibits axon growth by blocking mitochondrial transport. 61 Therefore, activating an intrinsic regrowth program requires the recovery of energy supply in injured axons. ...
Mitochondria supply adenosine triphosphate (ATP) essential for neuronal survival and regeneration. Brain injury and ischemia trigger acute mitochondrial damage and a local energy crisis, leading to degeneration. Boosting local ATP supply in injured axons is thus critical to meet increased energy demand during nerve repair and regeneration in adult brains, where mitochondria remain largely stationary. Here, we elucidate an intrinsic energetic repair signaling axis that boosts axonal energy supply by reprogramming mitochondrial trafficking and anchoring in response to acute injury-ischemic stress in mature neurons and adult brains. P21-activated kinase 5 (PAK5) is a brain mitochondrial kinase with declined expression in mature neurons. PAK5 synthesis and signaling is spatiotemporally activated within axons in response to ischemic stress and axonal injury. PAK5 signaling remobilizes and replaces damaged mitochondria via the phosphorylation switch that turns off the axonal mitochondrial anchor syntaphilin. Injury-ischemic insults trigger AKT growth signaling that activates PAK5 and boosts local energy supply, thus protecting axon survival and facilitating regeneration in in vitro and in vivo models. Our study reveals an axonal mitochondrial signaling axis that responds to injury and ischemia by remobilizing damaged mitochondria for replacement, thereby maintaining local energy supply to support central nervous system (CNS) survival and regeneration.
... Mitochondrial function in neuron is quite essential for electrical activity, axon extension, regeneration, branching, synaptic formation, and synaptic neurotransmission 5,29,30 , and altered mitochondrial function leads to neurodegenerative diseases [31][32][33] . Ndufs4 is the gene responsible for Leigh syndrome 17 . ...
Altered function of mitochondrial respiratory chain in brain cells is related to many neurodegenerative diseases. NADH Dehydrogenase (Ubiquinone) Fe-S protein 4 (Ndufs4) is one of the subunits of mitochondrial complex I and its mutation in human is associated with Leigh syndrome. However, the molecular biological role of Ndufs4 in neuronal function is poorly understood. In this study, upon Ndufs4 expression confirmation in NeuN-positive neurons, and GFAP-positive astrocytes in WT mouse hippocampus, we found significant decrease of mitochondrial respiration in Ndufs4-KO mouse hippocampus. Although there was no change in the number of NeuN positive neurons in Ndufs4-KO hippocampus, the expression of synaptophysin, a presynaptic protein, was significantly decreased. To investigate the detailed mechanism, we silenced Ndufs4 in Neuro-2a cells and we observed shorter neurite lengths with decreased expression of synaptophysin. Furthermore, western blot analysis for phosphorylated extracellular regulated kinase (pERK) revealed that Ndufs4 silencing decreases the activity of ERK signalling. These results suggest that Ndufs4-modulated mitochondrial activity may be involved in neuroplasticity via regulating synaptophysin expression.
... This process is supported by a regenerative program through expression of the Regeneration-associated gene (Attwell et al., 2018;Mahar and Cavalli, 2018). Nevertheless, axotomy at the central branch has a minimal regenerative response and the Regenerationassociated gene response also seems to make no difference on axon regeneration (Chandran et al., 2016;Cartoni et al., 2017). While in the CNS, the ability of axon regeneration is dependent on the maturity of the neurons. ...
Mitochondria are organelles responsible for bioenergetic metabolism, calcium homeostasis, and signal transmission essential for neurons due to their high energy consumption. Accumulating evidence has demonstrated that mitochondria play a key role in axon degeneration and regeneration under physiological and pathological conditions. Mitochondrial dysfunction occurs at an early stage of axon degeneration and involves oxidative stress, energy deficiency, imbalance of mitochondrial dynamics, defects in mitochondrial transport, and mitophagy dysregulation. The restoration of these defective mitochondria by enhancing mitochondrial transport, clearance of reactive oxidative species (ROS), and improving bioenergetic can greatly contribute to axon regeneration. In this paper, we focus on the biological behavior of axonal mitochondria in aging, injury (e.g., traumatic brain and spinal cord injury), and neurodegenerative diseases (Alzheimer's disease, AD; Parkinson's disease, PD; Amyotrophic lateral sclerosis, ALS) and consider the role of mitochondria in axon regeneration. We also compare the behavior of mitochondria in different diseases and outline novel therapeutic strategies for addressing abnormal mitochondrial biological behavior to promote axonal regeneration in neurological diseases and injuries.
... The transparent body of C. elegans further allows analysis of such phenotypes, including axon regrowth rate, guidance, and fusion at a single neuron resolution in vivo with fluorescence microscopy. Past nerve regeneration studies in C. elegans have led to the discovery of many conserved nerve regeneration pathways [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26] , notably the dual leucine zipper kinase (DLK-1/p38) signaling cascade, whose role in neural development, regeneration, and degeneration has been confirmed in numerous vertebrate and invertebrate species 7, 27-31 . ...
Our understanding of nerve regeneration can be enhanced by delineating its underlying molecular activities at single neuron resolution in small model organisms such as Caenorhabditis elegans. Existing cell isolation techniques cannot isolate regenerating neurons from the nematode. We present femtosecond laser microdissection (fs-LM), a new single cell isolation method that dissects intact cells directly from living tissue by leveraging the micron-scale precision of fs-laser ablation. We show that fs-LM facilitated sensitive and specific gene expression profiling by single cell RNA-sequencing, while mitigating the stress related transcriptional artifacts induced by tissue dissociation. Single cell RNA-sequencing of fs-LM isolated regenerating C. elegans neurons revealed transcriptional program leading to successful regeneration in wild-type animals or regeneration failure in animals lacking DLK-1/p38 kinase. The ability of fs-LM to isolate specific neurons based on phenotype of interest allowed us to study the molecular basis of regeneration heterogeneity displayed by neurons of the same type. We identified gene modules whose expression patterns were correlated with axon regrowth rate at a single neuron level. Our results establish fs-LM as a highly specific single cell isolation method ideal for precision and phenotype-driven studies.
... Robust mitochondria density is required to produce adequate ATP for axon regeneration. While Cartoni et al. (2016) demonstrated that the mammalian-specific mitochondrial protein Armcx1 mobilizes mitochondria and promotes neuronal survival and axonal regeneration in an optic nerve injury model. These significant studies demonstrate a critical role of mitochondrial transport and energy supply for axon regeneration in various models with different injury regimes. ...
... Our research showed that the mRNA level of PTEN, SOCS3, Klf9, and Mdm4 decreased signi cantly in AAV-STAT3 group. Overexpression of Dclk2, Armcx1, c-myc, and Nrn1 are closely related to the regeneration of RGCs axon (31)(32)(33)(34). The mRNA level of Dclk2, Armcx1, c-myc, and Nrn1 signi cantly increased in the AAV-STAT3 group. ...
BackgroundMüller differentiated RGCs have potential therapeutic value for glaucoma. However, axonal regeneration of differentiated RGCs has been a difficult problem. Studies have confirmed that STAT3 and Y27632 play essential roles in regulating neuronal axon regeneration. Whether STAT3 and Y27632 can induce the Müller differentiated RGCs axon regeneration is still unknown.Method
Retina Müller cells were isolated and purified from Day 21 SD rats’ retina and were differentiated into retinal stem cells. The stem cells were randomly divided into five groups (control group, AAV-STAT3 group, shSTAT3 group, Y27632 group and AAV-STAT3 + Y27632 group). The axon length in each group were measured by ImageJ. Immunofluorescence were used to label the RGCs. The mRNA level of pluripotent associated and differentiation-associated proteins was analysed by qRT-PCR. Stem cells in different groups were injected into mice model of glaucoma. Immunohistochemical, Immunohistochemistry and OCT were performed to access RGC layer thickness in glaucoma model. VEP was used to detect the optic nerve conduction function.ResultsIn this study, we found that overexpression of STAT3 could promote the growth of RGCs axons generated by Müller cell differentiation. Combined with Y27632, axonal regeneration was significantly longer than that of the STAT3 group. However, after STAT3 was knocked out, axonal regeneration significantly decreased or even stopped. The mRNA levels of Esrrb, Prdm14, Sox2, and Rex1 in Müller differentiated RGCs after overexpression STAT3 combined with Y27632 were significantly increased, while the mRNA levels of Nestin, Eomes, Mixl1 and Gata4 were significantly decreased. The mRNA levels of Socs3, Pten, Klf9, and Mdm4 were significantly decreased, while the mRNA levels of Dclk2, Armcx1, C-MYC, and Nrn1 were significantly increased. The mRNA levels of differentiation and pluripotency marker genes showed opposite results after STAT3 deletion. After injecting Müller differentiated RGCs intervened by STAT3 combined with Y27632 into the eyes of the glaucoma model mice, the axon length, OCT displayed RGC layer thickness and the electrophysiology indicated by VEP were superior to those of the glaucoma model group.Conclusions
These findings suggested that STAT3 combined with Y27632 can significantly improve the axonal growth level of RGCs, and reveal the potential mechanism to induce pluripotency of RGCs.
... PTEN knockout recruits ipRGCs for axon regeneration (Bray et al. 2019), as does AAV2-CNTF (Bray et al. 2019), but other RGCs have not been examined in detail with these models. Promoting mitochondrial transport in axons by overexpression of Armcx1 induced axon regeneration that was not enriched for α-RGCs (Cartoni et al. 2016). Finally, in a curious example, AAV2-Sox11 promoted robust RGC axon regeneration after crush but paradoxically killed α-RGCs even without axon injury (Norsworthy et al. 2017). ...
... Energy supply tends to promote neuronal survival following CNS injury [35,36]. Given that more internal energy failed to produce more surviving RGCs, we questioned whether supplying extrinsic ATP would confer any benefit for RGC survival. ...
Neurons, especially axons, are metabolically demanding and energetically vulnerable during injury. However, the exact energy budget alterations that occur early after axon injury and the effects of these changes on neuronal survival remain unknown. Using a classic mouse model of optic nerve-crush injury, we found that traumatized optic nerves and retinas harbor the potential to mobilize two primary energetic machineries, glycolysis and oxidative phosphorylation, to satisfy the robustly increased adenosine triphosphate (ATP) demand. Further exploration of metabolic activation showed that mitochondrial oxidative phosphorylation was amplified over other pathways, which may lead to decreased retinal ganglion cell (RGC) survival despite its supplement to ATP production. Gene set enrichment analysis of a microarray (GSE32309) identified significant activation of oxidative phosphorylation in injured retinas from wild-type mice compared to those from mice with deletion of phosphatase and tensin homolog (PTEN), while PTEN-/- mice had more robust RGC survival. Therefore, we speculated that the oxidation-favoring metabolic pattern after optic nerve-crush injury could be adverse for RGC survival. After redirecting metabolic flux toward glycolysis (magnifying the Warburg effect) using the drug meclizine, we successfully increased RGC survival. Thus, we provide novel insights into a potential bioenergetics-based strategy for neuroprotection.
... Moreover, new players regulating mitochondrial trafficking in neurons have been recently described. Among them, the Miro-interacting mitochondrial protein Armcx1 enhanced mitochondrial transport in adult RGC and promoted axonal regeneration after injury (Cartoni et al. 2016). Nonetheless, the molecular role of motor and adaptor proteins mediating mitochondrial transport in RGC remains to be further elucidated. ...
Inherited retinal dystrophies (IRDs) are a broad group of neurodegenerative disorders associated with reduced or deteriorating visual system. In the retina, cells are under constant oxidative stress, leading to elevated reactive oxygen species (ROS) generation that induces mitochondrial dysfunction and alteration of the mitochondrial network. This mitochondrial dysfunction combined with mutations in mitochondrial DNA and nuclear genes makes photoreceptors and retinal ganglion cells more susceptible to cell death. In this minireview, we focus on mitochondrial dynamics and their contribution to neuronal degeneration underlying IRDs, with particular attention to Leber hereditary optic neuropathy (LHON) and autosomal dominant optic atrophy (DOA), and propose targeting cell resilience and mitochondrial dynamics modulators as potential therapeutic approaches for retinal disorders.
... Moreover, new players regulating mitochondrial trafficking in neurons have been recently described. Among them, the Miro-interacting mitochondrial protein Armcx1 enhanced mitochondrial transport in adult RGC and promoted axonal regeneration after injury (Cartoni et al. 2016). Nonetheless, the molecular role of motor and adaptor proteins mediating mitochondrial transport in RGC remains to be further elucidated. ...
Leber congenital amaurosis (LCA) caused by AIPL1 mutations is one of the most severe forms of inherited retinal degeneration (IRD). The rapid and extensive photoreceptor degeneration challenges the development of potential treatments. Nevertheless, preclinical studies show that both gene augmentation and photoreceptor transplantation can regenerate and restore retinal function in animal models of AIPL1-associated LCA. However, questions regarding long-term benefit and safety still remain as these therapies advance towards clinical application. Ground-breaking advances in stem cell technology and genome editing are examples of alternative therapeutic approaches and address some of the limitations associated with previous methods. The continuous development of these cutting-edge biotechnologies paves the way towards a bright future not only for AIPL1-associated LCA patients but also other forms of IRD.
... It is conceivable that local ATP concentrations fluctuate dramatically following axon injury due to dilution of the cytoplasm by extracellular flux and mitochondrial dysfunction, resulting in local deficits that may create an environment permissive to phase separation. This is supported by evidence that local injection of ATP supports regeneration of spinal sensory axons , increases in mitochondrial density correlate with improved regeneration (Cartoni et al., 2017;Han et al., 2016), and several mitochondrial genes regulate regeneration (Cartoni et al., 2016;Knowlton et al., 2017;Zhou et al., 2016). We speculate that injury triggers changes in the axonal microenvironment, such as increased Zn 2+ or reduced ATP, that alter the biophysical properties of TIAR-2 granules into a regeneration-inhibitory state. ...
Phase separation into liquid-like compartments is an emerging property of proteins containing prion-like domains (PrLDs), yet the in vivo roles of phase separation remain poorly understood. TIA proteins contain a C-terminal PrLD, and mutations in the PrLD are associated with several diseases. Here, we show that the C. elegans TIAR-2/TIA protein functions cell autonomously to inhibit axon regeneration. TIAR-2 undergoes liquid-liquid phase separation in vitro and forms granules with liquid-like properties in vivo. Axon injury induces a transient increase in TIAR-2 granule number. The PrLD is necessary and sufficient for granule formation and inhibiting regeneration. Tyrosine residues within the PrLD are important for granule formation and inhibition of regeneration. TIAR-2 is also serine phosphorylated in vivo. Non-phosphorylatable TIAR-2 variants do not form granules and are unable to inhibit axon regeneration. Our data demonstrate an in vivo function for phase-separated TIAR-2 and identify features critical for its function in axon regeneration.
... Notably, only severing the central branch of DRG neurons did not increase mitochondrial transport, nor did it lead to axonal regrowth (Mar et al., 2014). Secondly, mice with a co-deletion of phosphate and tensin homolog (PTEN) and suppressor of cytokine signaling 3 (SOCS3), known to robustly enhance RGC axonal regrowth, show an increased expression of armadillo repeat containing X-linked 1 (Armcx1), a mitochondrial protein that interacts with the transport machinery via mitochondrial Rho GTPase 1 (Cartoni et al., 2017). Subsequent overexpression and knockdown experiments indicated that Armcx1 is crucial for the positive axonal regeneration effect after ONC and does so by releasing stationary mitochondria and thus promoting mitochondrial mobility. ...
Dendrites form an essential component of the neuronal circuit have been largely overlooked in regenerative research. Nevertheless, subtle changes in the dendritic arbors of neurons are one of the first stages of various neurodegenerative diseases, leading to dysfunctional neuronal networks and ultimately cellular death. Maintaining dendrites is therefore considered an essential neuroprotective strategy. This mini-review aims to discuss an intriguing hypothesis, which postulates that dendritic shrinkage is an important stimulant to boost axonal regeneration, and thus that preserving dendrites might not be the ideal therapeutic method to regain a full functional network upon central nervous system damage. Indeed, our study in zebrafish, a versatile animal model with robust regenerative capacity recently unraveled that dendritic retraction is evoked prior to axonal regrowth after optic nerve injury. Strikingly, inhibiting dendritic pruning upon damage perturbed axonal regeneration. This constraining effect of dendrites on axonal regrowth has sporadically been proposed in literature, as summarized in this short narrative. In addition, the review discusses a plausible underlying mechanism for the observed antagonistic axon-dendrite interplay, which is based on energy restriction inside neurons. Axonal injury indeed leads to a high local energy demand in which efficient axonal energy supply is fundamental to ensure regrowth. At the same time, axonal lesion is known to induce mitochondrial depolarization, causing energy depletion in the axonal compartment of damaged neurons. Mitochondria, however, become mostly stationary after development, which has been proposed as a potential underlying reason for the low regenerative capacity of adult mammals. Per contra, upon reduced neuronal activity, mitochondrial mobility enhances. In this view, dendritic shrinkage after axonal injury in zebrafish could result in less synaptic input and hence, a release of mitochondria within the soma-dendrite compartment that then translocate to the axonal growth cone to stimulate axonal regeneration. If this hypothesis proofs to be correct, i.e. dendritic remodeling serving as fuel for axonal regeneration, we envision a major shift in the research focus within the neuroregenerative field and in the potential uncovering of various novel therapeutic targets.
... Mitochondria are cellular organelles whose main functions are to generate ATP, buffer cytosolic calcium, and generate reactive oxygen species. Mitochondria also play an important role in the mechanism of axon extension, regeneration, and axon branching (1,2). The elongating axon requires a continuous supply of energy in areas distal from the cell body (1); proper mitochondria distribution is required for axonal regeneration (3), and indeed mitochondria movement is increased following axonal injury (4). ...
Neuronal regeneration is a highly energy‐demanding process that greatly relies on axonal mitochondrial transport to meet the enhanced metabolic requirements. Mature neurons typically fail to regenerate after injury, partly because of mitochondrial motility and energy deficits in injured axons. Retinoic acid receptor (RAR)‐β signaling is involved in axonal and neurite regeneration. Here we investigate the effect of RAR‐β signaling on mitochondrial trafficking during neurite outgrowth and find that it enhances their proliferation, speed, and movement toward the growing end of the neuron via hypoxia‐inducible factor 1α signaling. We also show that RAR‐β signaling promotes the binding of the mitochondria to the anchoring protein, glucose‐related protein 75, at the growing tip of neurite, thus allowing them to provide energy and metabolic roles required for neurite outgrowth.—Trigo, D., Goncalves, M. B., Corcoran, J. P. T. The regulation of mitochondrial dynamics in neurite outgrowth by retinoic acid receptor β signaling. FASEB J. 33, 7225–7235 (2019). www.fasebj.org
... Similarly, after optic nerve crush in mice, the mitochondrial protein Armcx1 is upregulated during injury in a regenerative condition, and further overexpression enhances both survival and regeneration of RGCs. This effect is hypothesized to be due to an increased in mobilization of mitochondria after injury, consistent with the results seen in C. elegans and in the mammalian sciatic nerve (Cartoni et al., 2016;Han et al., 2016;Zhou et al., 2016). Thus promoting increased mitochondrial transport promotes regenerative responses in the mammalian optic nerve, although it is not yet understood how regulation of this mitochondrial redistribution and energetics modulation contributes to survival and regeneration. ...
Retinal ganglion cells and other central nervous system neurons fail to regenerate after injury. Understanding the obstacles to survival and regeneration, and overcoming them, is key to preserving and restoring function. While comparisons in the cellular changes seen in these non‐regenerative cells with those that do have intrinsic regenerative ability has yielded many candidate genes for regenerative therapies, complete visual recovery has not yet been achieved. Insights gained from neurodegenerative diseases, like glaucoma, underscore the importance of axonal transport of organelles, mRNA, and effector proteins in injury and disease. Targeting molecular motor networks, and their cargoes, may be necessary for realizing complete axonal regeneration and vision restoration. This article is protected by copyright. All rights reserved.
... 8-Br-cAMP treatment also reduced the signal from an oxidative stress sensor ( Figures S2A and S2B). Although the cAMP analog is likely to affect many processes in wing neurons of aged flies, these results are consistent with previous evidence that mitochondrial transport has a protective role in adult neurons of Drosophila [5,8,15], C. elegans [6,16], and mice [17]. Several studies have reported that cAMP concentration modulates mitochondrial transport in cultured mammalian cells [18][19][20], although the underlying mechanisms have not been resolved. ...
... The Atf3:BAC Tg mouse could facilitate the study of axonal transport of mitochondria, which is closely associated with axonal growth and axonal degeneration. 25,[41][42][43][44][45] Another advantage is that the mice express Cre recombinase in response to nerve injury. Although we did not use this Cre system in this study, further gene deletion specifically in nerve-injured neurons is possible. ...
Damage-induced neuronal endopeptidase (DINE)/endothelin-converting enzyme-like 1 (ECEL1) is a membrane-bound metalloprotease that we identified as a nerve regeneration-associated molecule. The expression of DINE is upregulated in response to nerve injury in both the peripheral and central nervous systems, while its transcription is regulated by the activating transcription factor 3 (ATF3), a potent hub-transcription factor for nerve regeneration. Despite its unique hallmark of injury-induced upregulation, the physiological relevance of DINE in injured neurons has been unclear. In this study, we have demonstrated that the expression of DINE is upregulated in injured retinal ganglion cells (RGCs) in a coordinated manner with that of ATF3 after optic nerve injury, whereas DINE and ATF3 are not observed in any normal retinal cells. Recently, we have generated a mature DINE-deficient (KOTg) mouse, in which exogenous DINE is overexpressed specifically in embryonic motor neurons to avoid aberrant arborization of motor nerves and lethality after birth that occurs in the conventional DINE KO mouse. The DINE KOTg mice did not show any difference in retinal structure and the projection to brain from that of wild–type (wild type) mice under normal conditions. However, injured RGCs of DINE KOTg mice failed to regenerate even after the zymosan treatment, which is a well-known regeneration-promoting reagent. Furthermore, a DINE KOTg mouse crossed with a Atf3:BAC Tg mouse, in which green fluorescent protein (GFP) is visualized specifically in injured RGCs and optic nerves, has verified that DINE deficiency leads to regeneration failure. These findings suggest that injury-induced DINE is a crucial endopeptidase for injured RGCs to promote axonal regeneration after optic nerve injury. Thus, a DINE-mediated proteolytic mechanism would provide us with a new therapeutic strategy for nerve regeneration.
... Axon regeneration of mammalian neurons also appears to involve axonal mitochondrial transport, as loss of function in the mammalian-specific mitochondrial anchor protein syntaphilin results in enhanced axon regeneration (Zhou et al., 2016). Furthermore, the mammalian-specific transport protein Armcx1 is required for the enhanced regenerative capacity of some retinal ganglion axons in a regeneration-enhanced background (Cartoni et al., 2016). In contrast, Rawson and colleagues reported that in C. elegans ric-7 mutant worms, which cannot transport mitochondria into PLM axons, severed axons retained their regenerative competence (Rawson et al., 2014 ...
The role of mitochondria within injured neurons is an area of active interest since these organelles are vital for the production of cellular energy in the form of ATP. Using mechanosensory neurons of the nematode Caenorhabditis elegans to test regeneration after neuronal injury in vivo, we surveyed genes related to mitochondrial function for effects on axon regrowth after laser axotomy. Genes involved in mitochondrial transport, calcium uptake, mitophagy, or fission and fusion were largely dispensable for axon regrowth, with the exception of eat-3/Opa1. Surprisingly, many genes encoding components of the electron transport chain were dispensable for regrowth, except for the iron-sulfur proteins gas-1, nduf-2.2, nduf-7, and isp-1, and the putative oxidoreductase rad-8. In these mutants, axonal development was essentially normal and axons responded normally to injury by forming regenerative growth cones, but were impaired in subsequent axon extension. Overexpression of nduf-2.2 or isp-1 was sufficient to enhance regrowth, suggesting that mitochondrial function is rate-limiting in axon regeneration. Moreover, loss of function in isp-1 reduced the enhanced regeneration caused by either a gain-of-function mutation in the calcium channel EGL-19 or overexpression of the MAP kinase DLK-1. While the cellular function of RAD-8 remains unclear, our genetic analyses place rad-8 in the same pathway as other electron transport genes in axon regeneration. Unexpectedly, rad-8 regrowth defects were suppressed by altered function in the ubiquinone biosynthesis gene clk-1. Furthermore, we found that inhibition of the mitochondrial unfolded protein response via deletion of atfs-1 suppressed the defective regrowth in nduf-2.2 mutants. Together, our data indicate that while axon regeneration is not significantly affected by general dysfunction of cellular respiration, it is sensitive to the proper functioning of a select subset of electron transport chain genes, or to the cellular adaptations used by neurons under conditions of injury.
Oncomodulin (Ocm) is a myeloid cell-derived growth factor that enables axon regeneration in mice and rats after optic nerve injury or peripheral nerve injury, yet the mechanisms underlying its activity are unknown. Using proximity biotinylation, coimmunoprecipitation, surface plasmon resonance, and ectopic expression, we have identified armadillo-repeat protein C10 (ArmC10) as a high-affinity receptor for Ocm. ArmC10 deletion suppressed inflammation-induced axon regeneration in the injured optic nerves of mice. ArmC10 deletion also suppressed the ability of lesioned sensory neurons to regenerate peripheral axons rapidly after a second injury and to regenerate their central axons after spinal cord injury in mice (the conditioning lesion effect). Conversely, Ocm acted through ArmC10 to accelerate optic nerve and peripheral nerve regeneration and to enable spinal cord axon regeneration in these mouse nerve injury models. We showed that ArmC10 is highly expressed in human-induced pluripotent stem cell-derived sensory neurons and that exposure to Ocm altered gene expression and enhanced neurite outgrowth. ArmC10 was also expressed in human monocytes, and Ocm increased the expression of immune modulatory genes in these cells. These findings suggest that Ocm acting through its receptor ArmC10 may be a useful therapeutic target for nerve repair and immune modulation.
Our understanding of nerve regeneration can be enhanced by delineating its underlying molecular activities at single-neuron resolution in model organisms such as Caenorhabditis elegans. Existing cell isolation techniques cannot isolate neurons with specific regeneration phenotypes from C. elegans. We present femtosecond laser microdissection (fs-LM), a single-cell isolation method that dissects specific cells directly from living tissue by leveraging the micrometer-scale precision of fs-laser ablation. We show that fs-LM facilitates sensitive and specific gene expression profiling by single-cell RNA sequencing (scRNA-seq), while mitigating the stress-related transcriptional artifacts induced by tissue dissociation. scRNA-seq of fs-LM isolated regenerating neurons revealed transcriptional programs that are correlated with either successful or failed regeneration in wild-type and dlk-1 (0) animals, respectively. This method also allowed studying heterogeneity displayed by the same type of neuron and found gene modules with expression patterns correlated with axon regrowth rate. Our results establish fs-LM as a spatially resolved single-cell isolation method for phenotype-to-genotype mapping.
Mitochondrial transport along microtubules is mediated by Miro1 and TRAK adaptors that recruit kinesin-1 and dynein-dynactin. To understand how these opposing motors are regulated during mitochondrial transport, we reconstitute the bidirectional transport of Miro1/TRAK along microtubules in vitro. We show that the coiled-coil domain of TRAK activates dynein-dynactin and enhances the motility of kinesin-1 activated by its cofactor MAP7. We find that TRAK adaptors that recruit both motors move towards kinesin-1’s direction, whereas kinesin-1 is excluded from binding TRAK transported by dynein-dynactin, avoiding motor tug-of-war. We also test the predictions of the models that explain how mitochondrial transport stalls in regions with elevated Ca²⁺. Transport of Miro1/TRAK by kinesin-1 is not affected by Ca²⁺. Instead, we demonstrate that the microtubule docking protein syntaphilin induces resistive forces that stall kinesin-1 and dynein-driven motility. Our results suggest that mitochondrial transport stalls by Ca²⁺-mediated recruitment of syntaphilin to the mitochondrial membrane, not by disruption of the transport machinery.
Purpose:
Axon transport of organelles and neurotrophic factors is necessary for maintaining cellular function and survival of retinal ganglion cells (RGCs). However, it is not clear how trafficking of mitochondria, essential for RGC growth and maturation, changes during RGC development. The purpose of this study was to understand the dynamics and regulation of mitochondrial transport during RGC maturation using acutely purified RGCs as a model system.
Methods:
Primary RGCs were immunopanned from rats of either sex during three stages of development. MitoTracker dye and live-cell imaging were used to quantify mitochondrial motility. Analysis of single-cell RNA sequencing was used to identify Kinesin family member 5A (Kif5a) as a relevant motor candidate for mitochondrial transport. Kif5a expression was manipulated with either short hairpin RNA (shRNA) or exogenous expression adeno-associated virus viral vectors.
Results:
Anterograde and retrograde mitochondrial trafficking and motility decreased through RGC development. Similarly, the expression of Kif5a, a motor protein that transports mitochondria, also decreased during development. Kif5a knockdown decreased anterograde mitochondrial transport, while Kif5a expression increased general mitochondrial motility and anterograde mitochondrial transport.
Conclusions:
Our results suggested that Kif5a directly regulates mitochondrial axonal transport in developing RGCs. Future work exploring the role of Kif5a in vivo in RGCs is indicated.
Mitochondria generate ATP essential for neuronal growth, function, and regeneration. Due to their polarized structures, neurons face exceptional challenges to deliver mitochondria to and maintain energy homeostasis throughout long axons and terminal branches where energy is in high demand. Chronic mitochondrial dysfunction accompanied by bioenergetic failure is a pathological hallmark of major neurodegenerative diseases. Brain injury triggers acute mitochondrial damage and a local energy crisis that accelerates neuron death. Thus, mitochondrial maintenance defects and axonal energy deficits emerge as central problems in neurodegenerative disorders and brain injury. Recent studies have started to uncover the intrinsic mechanisms that neurons adopt to maintain (or reprogram) axonal mitochondrial density and integrity, and their bioenergetic capacity, upon sensing energy stress. In this review, we discuss recent advances in how neurons maintain a healthy pool of axonal mitochondria, as well as potential therapeutic strategies that target bioenergetic restoration to power neuronal survival, function, and regeneration.
Armcx1 is highly expressed in the brain and is located in the mitochondrial outer membrane of neurons, where it mediates mitochondrial transport. Mitochondrial transport promotes the removal of damaged mitochondria and the replenishment of healthy mitochondria, which are essential for neuronal survival after traumatic brain injury (TBI). This study investigated the role of Armcx1 and its underlying regulator(s) in secondary brain injury (SBI) after TBI. An in vivo TBI model was established in C57BL/6 mice via controlled cortical impact (CCI). Adeno-associated viruses with Armcx1 overexpression and knockdown were constructed and administered to mice by stereotactic cortical injection. Exogenous miR-223-3P mimic or inhibitor was transfected into cultured cortical neurons, which were then scratched to simulate TBI in vitro. The Armcx1 protein level was found to be decreased in peri-lesion tissue, particularly in neurons. The overexpression of Armcx1 significantly reduced TBI-induced neurological dysfunction, apoptosis, axonal injury, and mitochondrial dysfunction, while knockdown of Armcx1 had the opposite effect. Armcx1 was a direct target of miR-223-3P. The miR-223-3P mimic significantly reduced the Armcx1 protein level, while the miR-223-3P inhibitor had the opposite effect. Finally, the miR-223-3P inhibitor significantly improved mitochondrial membrane potential and increased the total length of the neurites without affecting branching numbers, while the miR-223-3P mimic had the opposite effect. In summary, our results suggest that the decreased expression of Armcx1 protein in neurons after experimental TBI aggravates secondary brain injury, which may be regulated by miR-223-3P. Therefore, this study provides a potential therapeutic approach for treating TBI.
Mitochondria are cellular organelles with roles in adenosine triphosphate production, cytosolic calcium buffering and reactive oxygen species production. A complex molecular motor system mediates the transport and positioning of mitochondria along axons. Recent studies have identified the transport and positioning of axonal mitochondria as crucial to multiple aspects of nervous system development including the developmental and regenerative extension of axons and the formation of axon collateral/interstitial branches, which underlie the formation of complex circuitry. Advances had been made possible by the development of novel experimental methods for targeting mitochondria and importantly for imaging mitochondria in living axons and in vivo. This chapter reviews the current understanding of the role of mitochondria in axon development.
Dual oxidase (duox)-derived reactive oxygen species (ROS) have been correlated with neuronal polarity, cerebellar development, and neuroplasticity. However, there have not been many comprehensive studies of the effect of individual duox isoforms on central-axon regeneration in vivo. Here, we explored this question in zebrafish, an excellent model organism for central-axon regeneration studies. In our research, mutation of the duox gene with CRISPR/Cas9 significantly retarded the single-axon regeneration of the zebrafish Mauthner cell in vivo. Using deep transcriptome sequencing, we found that the expression levels of related functional enzymes in mitochondria were down-regulated in duox mutant fish. In vivo imaging showed that duox mutants had significantly disrupted mitochondrial transport and redox state in single Mauthner-cell axon. Our research data provide insights into how duox is involved in central-axon regeneration by changing mitochondrial transport.
The failure of axons to regenerate in the damaged mammalian CNS is the main impediment to functional recovery. There are many molecules and structures in the environment of the injured nervous system that can inhibit regeneration, but even when these are removed or replaced with a permissive environment, most CNS neurons exhibit little regeneration of their axons. This contrasts with the extensive and vigorous axon growth that may occur when embryonic neurons are transplanted into the adult CNS. In the peripheral nervous system, the axons usually respond to axotomy with a vigorous regenerative response accompanied by a regenerative program of gene expression, usually referred to as the regeneration‐associated gene (RAG) program. These different responses to axotomy in the mature and immature CNS and the PNS lead to the concept of the intrinsic regenerative response of axons. Analysis of the many mechanisms and issues that affect the intrinsic regenerative response is the topic of this special issue of Developmental Neurobiology. The review articles highlight the control of expression of growth and regeneration‐associated genes, emphasizing the role of epigenetic mechanisms. The reviews also discuss changes within axons that lead to the developmental loss of regenerative ability. This is caused by changes in axonal transport and trafficking, in the cytoskeleton and in signaling pathways.
ResearchGate has not been able to resolve any references for this publication.