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Modified cell cycle status in a mouse model of altered neuronal vulnerability (slow Wallerian degeneration; Wlds).

Wishart TM, Pemberton HN, James SR, McCabe CJ, Gillingwater TH - Genome Biol. (2008)

Bottom Line: These include the following: elevated nicotinamide adenine dinucleotide (NAD) levels associated with nicotinamide mononucleotide adenylyltransferase 1 (Nmnat1; a part of the chimeric Wlds gene); altered mRNA expression levels of genes such as pituitary tumor transforming gene 1 (Pttg1); changes in the location/activity of the ubiquitin-proteasome machinery via binding to valosin-containing protein (VCP/p97); and modified synaptic expression of proteins such as ubiquitin-activating enzyme E1 (Ube1).We show that previous reports of diverse changes occurring downstream from Wlds expression converge upon modifications in cell cycle status.These data suggest a strong correlation between modified cell cycle pathways and altered vulnerability of axonal and synaptic compartments in postmitotic, terminally differentiated neurons.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Integrative Physiology, University of Edinburgh Medical School, Edinburgh, UK.

ABSTRACT

Background: Altered neuronal vulnerability underlies many diseases of the human nervous system, resulting in degeneration and loss of neurons. The neuroprotective slow Wallerian degeneration (Wlds) mutation delays degeneration in axonal and synaptic compartments of neurons following a wide range of traumatic and disease-inducing stimuli, providing a powerful experimental tool with which to investigate modulation of neuronal vulnerability. Although the mechanisms through which Wlds confers neuroprotection remain unclear, a diverse range of downstream modifications, incorporating several genes/pathways, have been implicated. These include the following: elevated nicotinamide adenine dinucleotide (NAD) levels associated with nicotinamide mononucleotide adenylyltransferase 1 (Nmnat1; a part of the chimeric Wlds gene); altered mRNA expression levels of genes such as pituitary tumor transforming gene 1 (Pttg1); changes in the location/activity of the ubiquitin-proteasome machinery via binding to valosin-containing protein (VCP/p97); and modified synaptic expression of proteins such as ubiquitin-activating enzyme E1 (Ube1).

Results: Wlds expression in mouse cerebellum and HEK293 cells induced robust increases in a broad spectrum of cell cycle-related genes. Both NAD-dependent and Pttg1-dependent pathways were responsible for mediating different subsets of these alterations, also incorporating changes in VCP/p97 localization and Ube1 expression. Cell proliferation rates were not modified by Wlds, suggesting that later mitotic phases of the cell cycle remained unaltered. We also demonstrate that Wlds concurrently altered endogenous cell stress pathways.

Conclusion: We report a novel cellular phenotype in cells with altered neuronal vulnerability. We show that previous reports of diverse changes occurring downstream from Wlds expression converge upon modifications in cell cycle status. These data suggest a strong correlation between modified cell cycle pathways and altered vulnerability of axonal and synaptic compartments in postmitotic, terminally differentiated neurons.

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Increased nuclear expression of cell cycle marker VCP corresponding with Wlds expression in mouse cerebellum. Confocal micrographs of cerebellar granule cells from (a-c) Wlds and (d-f) wild-type mice. Valosin-containing protein (VCP) is shown in green, the nuclear marker TOPRO3 is shown in blue, and Wlds protein in red (panels a and d show VCP and TOPRO3; panels b and e show Wlds and TOPRO3; and panels c and f show all three markers). Note how VCP protein can be seen in nuclear puncta with high frequency where Wlds is being expressed (arrows in panels a and c show four out of nine examples in this field of view). The majority of Wlds puncta coincided with VCP puncta. Nuclear puncta of VCP were rarely observed in wild-type cerebellar granule cells. As expected, VCP was detectable as diffuse staining in the cytoplasm of neurons in both strains of mice. Scale bar = 20 μm.
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Figure 11: Increased nuclear expression of cell cycle marker VCP corresponding with Wlds expression in mouse cerebellum. Confocal micrographs of cerebellar granule cells from (a-c) Wlds and (d-f) wild-type mice. Valosin-containing protein (VCP) is shown in green, the nuclear marker TOPRO3 is shown in blue, and Wlds protein in red (panels a and d show VCP and TOPRO3; panels b and e show Wlds and TOPRO3; and panels c and f show all three markers). Note how VCP protein can be seen in nuclear puncta with high frequency where Wlds is being expressed (arrows in panels a and c show four out of nine examples in this field of view). The majority of Wlds puncta coincided with VCP puncta. Nuclear puncta of VCP were rarely observed in wild-type cerebellar granule cells. As expected, VCP was detectable as diffuse staining in the cytoplasm of neurons in both strains of mice. Scale bar = 20 μm.

Mentions: Alongside identified changes in Pttg1/Ube1 expression and NAD pathways, previous studies have implicated VCP-mediated pathways (also known as p97 and CDC48) in Wlds-mediated neuroprotection, via its interaction with the Ube4b component of the Wlds chimeric protein [28]. Moreover, VCP is known to be important in early stages of cell cycle progression; VCP is normally localised in the endoplasmic reticulum during nonproliferative states (for example, terminally differentiated neurons), but relocates to the nucleus during late G1 phase in a cell cycle dependent manner [37]. Thus, VCP localisation would not normally be observed in the nucleus of terminally differentiated neurons unless cell cycle had been reactivated and they are progressing toward S phase. To examine whether VCP redistribution associated with modified cell cycle status is modified by Wlds, we examined VCP localization in the cerebellum of Wlds and wild-type mice. These experiments revealed an expected cytoplasmic, non-nuclear localization in wild-type neurons, but distinct, strong nuclear puncta in most cerebellar granule cells in Wlds mice (Figure 11). As predicted from the finding that VCP binds Wlds [28], VCP localization in the nucleus was consistently found in the same nuclear puncta as Wlds protein (Figure 11). These data provide further evidence that Wlds-expressing cells are being pushed toward the early phases of cell cycle re-entry and suggest that VCP binding may play a role in this process.


Modified cell cycle status in a mouse model of altered neuronal vulnerability (slow Wallerian degeneration; Wlds).

Wishart TM, Pemberton HN, James SR, McCabe CJ, Gillingwater TH - Genome Biol. (2008)

Increased nuclear expression of cell cycle marker VCP corresponding with Wlds expression in mouse cerebellum. Confocal micrographs of cerebellar granule cells from (a-c) Wlds and (d-f) wild-type mice. Valosin-containing protein (VCP) is shown in green, the nuclear marker TOPRO3 is shown in blue, and Wlds protein in red (panels a and d show VCP and TOPRO3; panels b and e show Wlds and TOPRO3; and panels c and f show all three markers). Note how VCP protein can be seen in nuclear puncta with high frequency where Wlds is being expressed (arrows in panels a and c show four out of nine examples in this field of view). The majority of Wlds puncta coincided with VCP puncta. Nuclear puncta of VCP were rarely observed in wild-type cerebellar granule cells. As expected, VCP was detectable as diffuse staining in the cytoplasm of neurons in both strains of mice. Scale bar = 20 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 11: Increased nuclear expression of cell cycle marker VCP corresponding with Wlds expression in mouse cerebellum. Confocal micrographs of cerebellar granule cells from (a-c) Wlds and (d-f) wild-type mice. Valosin-containing protein (VCP) is shown in green, the nuclear marker TOPRO3 is shown in blue, and Wlds protein in red (panels a and d show VCP and TOPRO3; panels b and e show Wlds and TOPRO3; and panels c and f show all three markers). Note how VCP protein can be seen in nuclear puncta with high frequency where Wlds is being expressed (arrows in panels a and c show four out of nine examples in this field of view). The majority of Wlds puncta coincided with VCP puncta. Nuclear puncta of VCP were rarely observed in wild-type cerebellar granule cells. As expected, VCP was detectable as diffuse staining in the cytoplasm of neurons in both strains of mice. Scale bar = 20 μm.
Mentions: Alongside identified changes in Pttg1/Ube1 expression and NAD pathways, previous studies have implicated VCP-mediated pathways (also known as p97 and CDC48) in Wlds-mediated neuroprotection, via its interaction with the Ube4b component of the Wlds chimeric protein [28]. Moreover, VCP is known to be important in early stages of cell cycle progression; VCP is normally localised in the endoplasmic reticulum during nonproliferative states (for example, terminally differentiated neurons), but relocates to the nucleus during late G1 phase in a cell cycle dependent manner [37]. Thus, VCP localisation would not normally be observed in the nucleus of terminally differentiated neurons unless cell cycle had been reactivated and they are progressing toward S phase. To examine whether VCP redistribution associated with modified cell cycle status is modified by Wlds, we examined VCP localization in the cerebellum of Wlds and wild-type mice. These experiments revealed an expected cytoplasmic, non-nuclear localization in wild-type neurons, but distinct, strong nuclear puncta in most cerebellar granule cells in Wlds mice (Figure 11). As predicted from the finding that VCP binds Wlds [28], VCP localization in the nucleus was consistently found in the same nuclear puncta as Wlds protein (Figure 11). These data provide further evidence that Wlds-expressing cells are being pushed toward the early phases of cell cycle re-entry and suggest that VCP binding may play a role in this process.

Bottom Line: These include the following: elevated nicotinamide adenine dinucleotide (NAD) levels associated with nicotinamide mononucleotide adenylyltransferase 1 (Nmnat1; a part of the chimeric Wlds gene); altered mRNA expression levels of genes such as pituitary tumor transforming gene 1 (Pttg1); changes in the location/activity of the ubiquitin-proteasome machinery via binding to valosin-containing protein (VCP/p97); and modified synaptic expression of proteins such as ubiquitin-activating enzyme E1 (Ube1).We show that previous reports of diverse changes occurring downstream from Wlds expression converge upon modifications in cell cycle status.These data suggest a strong correlation between modified cell cycle pathways and altered vulnerability of axonal and synaptic compartments in postmitotic, terminally differentiated neurons.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Integrative Physiology, University of Edinburgh Medical School, Edinburgh, UK.

ABSTRACT

Background: Altered neuronal vulnerability underlies many diseases of the human nervous system, resulting in degeneration and loss of neurons. The neuroprotective slow Wallerian degeneration (Wlds) mutation delays degeneration in axonal and synaptic compartments of neurons following a wide range of traumatic and disease-inducing stimuli, providing a powerful experimental tool with which to investigate modulation of neuronal vulnerability. Although the mechanisms through which Wlds confers neuroprotection remain unclear, a diverse range of downstream modifications, incorporating several genes/pathways, have been implicated. These include the following: elevated nicotinamide adenine dinucleotide (NAD) levels associated with nicotinamide mononucleotide adenylyltransferase 1 (Nmnat1; a part of the chimeric Wlds gene); altered mRNA expression levels of genes such as pituitary tumor transforming gene 1 (Pttg1); changes in the location/activity of the ubiquitin-proteasome machinery via binding to valosin-containing protein (VCP/p97); and modified synaptic expression of proteins such as ubiquitin-activating enzyme E1 (Ube1).

Results: Wlds expression in mouse cerebellum and HEK293 cells induced robust increases in a broad spectrum of cell cycle-related genes. Both NAD-dependent and Pttg1-dependent pathways were responsible for mediating different subsets of these alterations, also incorporating changes in VCP/p97 localization and Ube1 expression. Cell proliferation rates were not modified by Wlds, suggesting that later mitotic phases of the cell cycle remained unaltered. We also demonstrate that Wlds concurrently altered endogenous cell stress pathways.

Conclusion: We report a novel cellular phenotype in cells with altered neuronal vulnerability. We show that previous reports of diverse changes occurring downstream from Wlds expression converge upon modifications in cell cycle status. These data suggest a strong correlation between modified cell cycle pathways and altered vulnerability of axonal and synaptic compartments in postmitotic, terminally differentiated neurons.

Show MeSH
Related in: MedlinePlus