Neuronal non-CG methylation is an essential target for MeCP2 function.

DNA methylation is implicated in neuronal biology via the protein MeCP2, the mutation of which causes Rett syndrome. MeCP2 recruits the NCOR1/2 co-repressor complexes to methylated cytosine in the CG dinucleotide, but also to sites of non-CG methylation, which are abundant in neurons. To test the biological significance of the dual-binding specificity of MeCP2, we replaced its DNA binding domain with an orthologous domain from MBD2, which can only bind mCG motifs. Knockin mice expressing the domain-swap protein displayed severe Rett-syndrome-like phenotypes, indicating that normal brain function requires the interaction of MeCP2 with sites of non-CG methylation, specifically mCAC. The results support the notion that the delayed onset of Rett syndrome is due to the simultaneous post-natal accumulation of mCAC and its reader MeCP2. Intriguingly, genes dysregulated in both Mecp2 null and domain-swap mice are implicated in other neurological disorders, potentially highlighting targets of relevance to the Rett syndrome phenotype.

Dopey-dependent regulation of extracellular vesicles maintains neuronal morphology.

Mature neurons maintain their distinctive morphology for extended periods in adult life. Compared to developmental neurite outgrowth, axon guidance, and target selection, relatively little is known of mechanisms that maintain the morphology of mature neurons. Loss of function in C. elegans dip-2, a member of the conserved lipid metabolic regulator Dip2 family, results in progressive overgrowth of neurites in adults. We find that dip-2 mutants display specific genetic interactions with sax-2, the C. elegans ortholog of Drosophila Furry and mammalian FRY. Combined loss of dip-2 and sax-2 results in failure to maintain neuronal morphology and elevated release of neuronal extracellular vesicles (EVs). By screening for suppressors of dip-2(0) sax-2(0) double mutant defects, we identified gain-of-function (gf) mutations in the conserved Dopey family protein PAD-1 and its associated phospholipid flippase TAT-5/ATP9A that restore normal neuronal morphology and normal levels of EV release to dip-2(0) sax-2(0) double mutants. Neuron-specific knockdown suggests that PAD-1(gf) can act cell autonomously in neurons. PAD-1(gf) displays increased association with the plasma membrane in oocytes and inhibits EV release in multiple cell types. Our findings uncover a novel functional network of DIP-2, SAX-2, PAD-1, and TAT-5 that maintains neuronal morphology and modulates EV release.

Dopey-dependent regulation of extracellular vesicles maintains neuronal morphology.

Mature neurons maintain their distinctive morphology for extended periods in adult life. Compared to developmental neurite outgrowth, axon guidance, and target selection, relatively little is known of mechanisms that maintain mature neuron morphology. Loss of function in C. elegans DIP-2, a member of the conserved lipid metabolic regulator Dip2 family, results in progressive overgrowth of neurites in adults. We find that dip-2 mutants display specific genetic interactions with sax-2, the C. elegans ortholog of Drosophila Furry and mammalian FRY. Combined loss of DIP-2 and SAX-2 results in severe disruption of neuronal morphology maintenance accompanied by increased release of neuronal extracellular vesicles (EVs). By screening for suppressors of dip-2 sax-2 double mutant defects we identified gain-of-function (gf) mutations in the conserved Dopey family protein PAD-1 and its associated phospholipid flippase TAT-5/ATP9A. In dip-2 sax-2 double mutants carrying either pad-1(gf) or tat-5(gf) mutation, EV release is reduced and neuronal morphology across multiple neuron types is restored to largely normal. PAD-1(gf) acts cell autonomously in neurons. The domain containing pad-1(gf) is essential for PAD-1 function, and PAD-1(gf) protein displays increased association with the plasma membrane and inhibits EV release. Our findings uncover a novel functional network of DIP-2, SAX-2, PAD-1, and TAT-5 that maintains morphology of neurons and other types of cells, shedding light on the mechanistic basis of neurological disorders involving human orthologs of these genes.

Neuronal postdevelopmentally acting SAX-7S/L1CAM can function as cleaved fragments to maintain neuronal architecture in Caenorhabditis elegans.

Whereas remarkable advances have uncovered mechanisms that drive nervous system assembly, the processes responsible for the lifelong maintenance of nervous system architecture remain poorly understood. Subsequent to its establishment during embryogenesis, neuronal architecture is maintained throughout life in the face of the animal's growth, maturation processes, the addition of new neurons, body movements, and aging. The Caenorhabditis elegans protein SAX-7, homologous to the vertebrate L1 protein family of neural adhesion molecules, is required for maintaining the organization of neuronal ganglia and fascicles after their successful initial embryonic development. To dissect the function of sax-7 in neuronal maintenance, we generated a null allele and sax-7S-isoform-specific alleles. We find that the null sax-7(qv30) is, in some contexts, more severe than previously described mutant alleles and that the loss of sax-7S largely phenocopies the null, consistent with sax-7S being the key isoform in neuronal maintenance. Using a sfGFP::SAX-7S knock-in, we observe sax-7S to be predominantly expressed across the nervous system, from embryogenesis to adulthood. Yet, its role in maintaining neuronal organization is ensured by postdevelopmentally acting SAX-7S, as larval transgenic sax-7S(+) expression alone is sufficient to profoundly rescue the null mutants' neuronal maintenance defects. Moreover, the majority of the protein SAX-7 appears to be cleaved, and we show that these cleaved SAX-7S fragments together, not individually, can fully support neuronal maintenance. These findings contribute to our understanding of the role of the conserved protein SAX-7/L1CAM in long-term neuronal maintenance and may help decipher processes that go awry in some neurodegenerative conditions.

The where, what, and when of membrane protein degradation in neurons.

Membrane protein turnover and degradation are required for the function and health of all cells. Neurons may live for the entire lifetime of an organism and are highly polarized cells with spatially segregated axonal and dendritic compartments. Both longevity and morphological complexity represent challenges for regulated membrane protein degradation. To investigate how neurons cope with these challenges, an increasing number of recent studies investigated local, cargo-specific protein sorting, and degradation at axon terminals and in dendritic processes. In this review, we explore the current answers to the ensuing questions of where, what, and when membrane proteins are degraded in neurons. © 2017 The Authors Developmental Neurobiology Published by Wiley Periodicals, Inc. Develop Neurobiol 78: 283-297, 2018.

Chromatin Changes Associated with Neuronal Maintenance and Their Pharmacological Application.

BACKGROUND: The transcriptional control of neuronal specification and early development has been intensively studied over the past few decades. However, relatively little is known about transcriptional programs associated with the maintenance of terminally differentiated neuronal cells with respect to their functions, structures, and cell type-specific identity features. METHODS: Notably, largely because of the recent advances in related techniques such as next generation sequencing and chromatin immunoprecipitation sequencing, the physiological implications of system-wide regulation of gene expression through changes in chromatin states have begun to be extensively studied in various contexts and systems, including the nervous system. RESULTS: Here, we attempt to review our current understanding of the link between chromatin changes and neuronal maintenance in the period of life after the completion of neuronal development. Perturbations involving chromatin changes in the system-wide transcriptional control are believed to be closely associated with diverse aspects of neuronal aging and neurodegenerative conditions. CONCLUSION: In this review, we focused on heterochromatin and epigenetic dysregulation in neurodegenerative conditions as well as neuronal aging, the most important risk factor leading to neuronal degeneration, in order to highlight the close association between chromatin changes and neuronal maintenance. Lastly, we reviewed the currently available and potential future applications of pharmacological control of the chromatin states associated with neuronal maintenance.

Emerging nexus between RAB GTPases, autophagy and neurodegeneration.

The RAB class of small GTPases includes the major regulators of intracellular communication, which are involved in vesicle generation through fusion and fission, and vesicular trafficking. RAB proteins also play an imperative role in neuronal maintenance and survival. Recent studies in the field of neurodegeneration have also highlighted the process of autophagy as being essential for neuronal maintenance. Here we review the emerging roles of RAB proteins in regulating macroautophagy and its impact in the context of neurodegenerative diseases.

Effects of ADAM10 deletion on Notch-1 signaling pathway and neuronal maintenance in adult mouse brain.

A disintegrin and metalloproteinase 10 (ADAM10) has been demonstrated to regulate embryonic brain development by initiating Notch signaling. However, it is still unclear whether ADAM10 is required to activate the Notch signaling pathway in adult brain. To investigate the physiological role of ADAM10, we generated conditional knockout (cKO) mice lacking the Adam10 gene primarily in the cortex and hippocampus. We found that conditional disruption of ADAM10 resulted in a prominent decrease in the number of proliferating neuronal progenitor cells in the subgranular zone (SGZ), and a significant increase in the number of adult-generated postmitotic neurons in the hippocampal dentate gyrus (DG) due to premature neuronal differentiation. Moreover, the mutant mice also displayed an age-dependent reduction in the number of granule neurons in the hippocampal DG. It was further showed that the activation of Notch-1 and its downstream target genes Hes1, Hes5, Hey1, and Hey2 was impaired in ADAM10-deficient hippocampal tissues. Finally, Adam10 cKO mice had impaired learning and memory in the Morris water-maze. Thus, we provided experimental evidence to demonstrate that ADAM10 plays an essential role in the activation of Notch-1 signaling and has a remarkable effect on neuronal maintenance in adult mouse brain.

NMNATs, evolutionarily conserved neuronal maintenance factors.

Proper brain function requires neuronal homeostasis over a range of environmental challenges. Neuronal activity, injury, and aging stress the nervous system, and lead to neuronal dysfunction and degeneration. Nevertheless, most organisms maintain healthy neurons throughout life, implying the existence of active maintenance mechanisms. Recent studies have revealed a key neuronal maintenance and protective function for nicotinamide mononucleotide adenylyl transferases (NMNATs). We review evidence that NMNATs protect neurons through multiple mechanisms in different contexts, and highlight functions that either require or are independent of NMNAT catalytic activity. We then summarize data supporting a role for NMNATs in neuronal maintenance and raise intriguing questions on how NMNATs preserve neuronal integrity and facilitate proper neural function throughout life.