Restoration of cellular ubiquitin reverses impairments in neuronal development caused by disruption of the polyubiquitin gene Ubb

Ryu, Han-Wook; Park, Chulwoo; Ryu, Kwon‐Yul

doi:10.1016/j.bbrc.2014.09.103

Cited by 17 publications

(16 citation statements)

References 23 publications

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“…1b ). As previously reported in Ubb −/− cells 13 , upregulation of the polyubiquitin gene Ubc was also observed in Ubb KD cells (Fig. 1c ).…”

Section: Resultssupporting

confidence: 89%

“…When cells were isolated from embryonic brains on 14.5 dpc, two-thirds of them were NSCs 12 . Upon culture in the neuronal growth medium, these cells differentiated into neurons 13 . In fact, immunofluorescence analysis using the NSC marker nestin showed the gradual decrease of the number of NSCs as culture progressed (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…In our previous studies, we used Ubb −/− cells, in which cellular Ub levels are reduced throughout the neuronal development; therefore, the requirement of Ub for a specific process could not be investigated 12 , 13 . Although we were able to establish a link between cellular Ub levels and neuronal development, most importantly, we could not answer whether the impaired neuronal maturation was caused by the reduced Ub levels during maturation process or by the defects residing in the neurons generated under Ub deficiency.…”

Section: Introductionmentioning

confidence: 99%

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Temporal downregulation of the polyubiquitin gene Ubb affects neuronal differentiation, but not maturation, in cells cultured in vitro

Jung

Park

Ryu

2018

Sci Rep

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Reduced levels of cellular ubiquitin (Ub) pools due to disruption of the polyubiquitin gene Ubb lead to dysregulation of neural stem cell (NSC) differentiation and impaired neuronal maturation in cells isolated from Ubb−/− mouse embryonic brains. However, it is currently unknown whether Ub is required for the specific stage of neuronal development or whether it plays a pleiotropic role throughout the process. To answer this question, we aimed to downregulate Ubb expression temporally during neuronal development, which could not be achieved in Ubb −/− cells. Therefore, we exploited lentivirusmediated knockdown (KD) of Ubb at different stages of neuronal development, and investigated their phenotypes. Here, we report the outcome of Ubb KD on two independent culture days in vitro (DIV): DIV1 and DIV7. We observed that NSCs did not differentiate properly via Ubb KD on DIV1, but the maturation of already differentiated neurons was intact via Ubb KD on DIV7. Intriguingly, Ubb KD activated Notch signaling when it had been suppressed, but exerted no effect when it had already been activated. Therefore, our study suggests that Ub plays a pivotal role in NSC differentiation to suppress Notch signaling, but not in the subsequent maturation stages of neurons that had already been differentiated.Ubiquitin (Ub) is one of the most abundant eukaryotic proteins involved in post-translational modifications 1-3 . Ubiquitylation of target substrates occurs via the actions of three enzymes: E1 Ub-activating enzymes, E2 Ub-conjugation enzymes, and E3 Ub ligases 4,5 . The fate of the substrates is determined by the type of ubiquitylation, i.e., monoubiquitylation or polyubiquitylation with a specific Ub chain linkages 6,7 . The best-known and most important outcome is the polyubiquitylation of substrates with Lys 48 (K48) linkages and their targeting to the 26S proteasome for degradation 8,9 . A timely degradation of substrates is also important for the differentiation of neural stem cells (NSCs), onset of neurogenesis, neuronal development, and neuronal function 10,11 . Indeed, reduced levels of cellular Ub via the disruption of the polyubiquitin gene Ubb compromised the degradation of substrates and resulted in the dysregulation of NSC differentiation with inhibition of neurogenesis and impaired neuronal maturation [12][13][14] . NSC differentiation is mainly regulated by Notch signaling 15,16 . Typically, Notch signaling is suppressed to promote neurogenesis during embryonic stages, while it is activated to promote gliogenesis and neuronal maturation during postnatal stages [17][18][19][20] . Notch signaling is initiated by the interaction between the Notch receptor and its ligand Delta (DLL1) in the neighboring cells, followed by the cleavage and release of Notch intracellular domain (NICD) via γ-secretase. NICD then translocates into the nucleus to form a transactivator complex and activates the transcription of target genes 21 . Notch target genes such as the Hairy/enhancer of split (Hes) and Hairy/enhancer-of-split related with...

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“…1b ). As previously reported in Ubb −/− cells 13 , upregulation of the polyubiquitin gene Ubc was also observed in Ubb KD cells (Fig. 1c ).…”

Section: Resultssupporting

confidence: 89%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Temporal downregulation of the polyubiquitin gene Ubb affects neuronal differentiation, but not maturation, in cells cultured in vitro

Jung

Park

Ryu

2018

Sci Rep

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“…To produce a lentivirus harboring shNrf2, packaging plasmid psPAX2 (8 μg), envelope plasmid pMD2.G (3 μg), and transfer plasmid pLKO.1-shNrf2 (10 μg), were transfected into the HEK-293T cells and the viral pellet was obtained as previously described [56]. The MEFs were infected with lentivirus harboring scrambled or Nrf2 shRNA for 16 h and cultured for another 48-72 h before harvesting the infected cells.…”

Section: Generation Of Lentivirus Harboring Shrna Against Nrf2mentioning

confidence: 99%

Disruption of polyubiquitin gene Ubc leads to attenuated resistance against arsenite-induced toxicity in mouse embryonic fibroblasts

Kim¹,

Choi²,

Ryu³

et al. 2015

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

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The polyubiquitin gene Ubc is upregulated under oxidative stress induced by arsenite [As(III)]. However, the detailed mechanism of Ubc upregulation and the exact role of ubiquitin (Ub) to protect cells against As(III)-induced toxicity remain unknown. Here, we found that Ubc-/- mouse embryonic fibroblasts (MEFs) exhibited reduced viability under As(III) exposure, although the Nrf2-Keap1 pathway was activated as a cytoprotective response. Intriguingly, due to the reduced polyubiquitination and delayed onset of degradation of Nrf2 in Ubc-/- MEFs, the basal expression levels of Nrf2 target genes were elevated. As(III)-induced accumulation of Ub conjugates occurred in an Nrf2-independent manner, probably due to cellular stress conditions, including reduced proteasomal activity. Increased cellular Ub levels were essential to polyubiquitinate misfolded proteins generated under As(III) exposure and to degrade them by the proteasome. However, when cellular Ub levels decreased, these misfolded proteins were not efficiently polyubiquitinated, but rather accumulated as large protein aggregates inside the cells, causing cytotoxicity. Furthermore, increased activity of the autophagic pathway to clear these aggregates was not observed in Ubc-/- MEFs. Therefore, reduced viability of Ubc-/- MEFs under As(III) exposure may not be due to dysregulation of the Nrf2-Keap1 pathway, but mostly to reduced efficacy to polyubiquitinate and degrade misfolded protein aggregates.

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“…UB genes are known to play key roles during neuronal development, including neuritogenesis, neurogenesis, and synaptogenesis (Kawabe & Brose, 2011). Ryu et al (Ryu, Park, & Ryu, 2014) found that neuronal morphology, neurite outgrowth, and synaptic development were impaired in UBB −/− neurons. UBC and UBA52 may play a critical role in compensating for disruption of UBB in neurons and astrocytes (Sinnar et al, 2011), and disruption of the UBB gene can cause hypothalamic neurodegeneration and sleep abnormalities in mice (Ryu et al, 2010).…”

Section: Discussionmentioning

confidence: 99%

Systematic genetic analyses of GWAS data reveal an association between the immune system and insomnia

Xiang

Liu

et al. 2019

Molec Gen & Gen Med

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Background Previous studies have inferred a strong genetic component for insomnia. However, the etiology of insomnia is still unclear. The aim of the current study was to explore potential biological pathways, gene networks, and brain regions associated with insomnia. Methods Using pathways (gene sets) from Reactome, we carried out a two‐stage gene set enrichment analysis strategy. From a large genome‐wide association studies (GWASs) of insomnia symptoms (32,155 cases/26,973 controls), significant gene sets were tested for replication in other large GWASs of insomnia complaints (32,384 cases/80,622 controls). After the network analysis of unique genes within the replicated pathways, a gene set analysis for genes in each cluster/module of the enhancing neuroimaging genetics through meta‐analysis GWAS data was performed for the volumes of the intracranial and seven subcortical regions. Results A total of 31 of 1,816 Reactome pathways were identified and showed associations with insomnia risk. In addition, seven functionally and topologically interconnected clusters (clusters 0–6) and six gene modules (named Yellow, Blue, Brown, Green, Red, and Turquoise) were associated with insomnia. Moreover, significant associations were detected between common variants of the genes in Cluster 2 with hippocampal volume ( p = 0.035; family wise error [FWE] correction) and the red module with intracranial volume ( p = 0.047; FWE correction). Functional enrichment for genes in the Cluster 2 and the Red module revealed the involvement of immune responses, nervous system development, NIK/NF‐kappaB signaling, and I‐kappaB kinase/NF‐kappaB signaling. Core genes ( UBC , UBB , and UBA52 ) in the interconnected functional network were found to be involved in regulating brain development. Conclusions The current study demonstrates that the immune system and the hippocampus may play central roles in neurodevelopment and insomnia risk.

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Restoration of cellular ubiquitin reverses impairments in neuronal development caused by disruption of the polyubiquitin gene Ubb

Cited by 17 publications

References 23 publications

Temporal downregulation of the polyubiquitin gene Ubb affects neuronal differentiation, but not maturation, in cells cultured in vitro

Temporal downregulation of the polyubiquitin gene Ubb affects neuronal differentiation, but not maturation, in cells cultured in vitro

Disruption of polyubiquitin gene Ubc leads to attenuated resistance against arsenite-induced toxicity in mouse embryonic fibroblasts

Systematic genetic analyses of GWAS data reveal an association between the immune system and insomnia

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