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Unique aspects of transcriptional regulation in neurons--nuances in NFkappaB and Sp1-related factors.

Mao XR, Moerman-Herzog AM, Chen Y, Barger SW - J Neuroinflammation (2009)

Bottom Line: These studies led us to conclude that this population of cells is nearly incapable of activating the NFkappaB that is nonetheless expressed at reasonable levels.A subset of the kappaB cis elements are instead bound by members of the Sp1 family in neurons.Also surprising was our discovery that Sp1 itself, typically described as ubiquitous, is severely restricted in expression within forebrain neurons; Sp4 seems to be substituted during neuronal differentiation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anesthesiology, Washington University School of Medicine, St Louis, MO 63110, USA. maox@morpheus.wustl.edu

ABSTRACT
The unique physiology and function of neurons create differences in their cellular physiology, including their regulation of gene expression. We began several years ago exploring the relationships between the NFkappaB transcription factor, neuronal survival, and glutamate receptor activation in telencephalic neurons. These studies led us to conclude that this population of cells is nearly incapable of activating the NFkappaB that is nonetheless expressed at reasonable levels. A subset of the kappaB cis elements are instead bound by members of the Sp1 family in neurons. Also surprising was our discovery that Sp1 itself, typically described as ubiquitous, is severely restricted in expression within forebrain neurons; Sp4 seems to be substituted during neuronal differentiation. These findings and their implications for neuronal differentiation--as well as potential dedifferentiation during degenerative processes--are discussed here.

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Glutamate is unable to activate neuronal NFκB in mixed cell cultures. Neocortical neurons from κB/β-gal transgenic mice were either plated on the top of wild-type astrocytes ("+glia") or retained as nearly pure neuronal cultures ("-glia"). A. Glutamate was applied at 50 μM for 10 min, then chased for the times indicated (h). B. Glutamate was applied at 20 μM continuously for the times indicated (h). β-gal activity was determined by a luminescent assay. Values represent activity relative to untreated cultures ± SEM in quadruplicate cultures. ANOVA followed by Scheffe post-hoc test showed significance between control and each treatment, and between the "+glia" and "-glia" in all comparisons except the 6 h timepoint in A. (p < 0.02).
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Figure 2: Glutamate is unable to activate neuronal NFκB in mixed cell cultures. Neocortical neurons from κB/β-gal transgenic mice were either plated on the top of wild-type astrocytes ("+glia") or retained as nearly pure neuronal cultures ("-glia"). A. Glutamate was applied at 50 μM for 10 min, then chased for the times indicated (h). B. Glutamate was applied at 20 μM continuously for the times indicated (h). β-gal activity was determined by a luminescent assay. Values represent activity relative to untreated cultures ± SEM in quadruplicate cultures. ANOVA followed by Scheffe post-hoc test showed significance between control and each treatment, and between the "+glia" and "-glia" in all comparisons except the 6 h timepoint in A. (p < 0.02).

Mentions: We have further analyzed in vitro neurons from the transgenic line developed by Bhakar et al. [33]. The main objective was to explore the possibility that pure neuronal cultures are altered in their handling of NFκB due to the absence of a physical interaction with glia. To test this idea, we cultured neurons from the κB/β-gal reporter line with wild-type glia so that typical neuron-glia interactions could take place, while the reporting of NFκB activity would be specific to the neurons in the coculture. Wild-type astrocytes were first plated and grown to confluency, and then E17 transgenic (κB/β-gal) cortical cells were added one week prior to treatment. The pre-plated wild-type astrocytes suppressed growth of transgenic glia but promoted survival of the κB/β-gal neurons, as histochemistry showed no astrocytes positive for β-gal. These cocultures were compared to cultures of κB/β-gal neurons alone. Two treatment paradigms were tested: 1) a transient exposure to glutamate (10 min) followed by a 6- to 24-hour chase period, or 2) continuous exposure to glutamate for 6 to 24 hours (Figure 2). Once again, neuronally localized NFκB activity could not be induced by glutamate. Indeed, basal β-gal levels were suppressed by glutamate exposure, an effect that was partially suppressed in the presence of glia, presumably due to the ability of astrocytes to metabolize glutamate. This negative effect of glutamate on κB-dependent transcription is consistent with copious EMSA data showing that glutamate reduces the levels of κB-binding Sp1-related factors in neurons (below).


Unique aspects of transcriptional regulation in neurons--nuances in NFkappaB and Sp1-related factors.

Mao XR, Moerman-Herzog AM, Chen Y, Barger SW - J Neuroinflammation (2009)

Glutamate is unable to activate neuronal NFκB in mixed cell cultures. Neocortical neurons from κB/β-gal transgenic mice were either plated on the top of wild-type astrocytes ("+glia") or retained as nearly pure neuronal cultures ("-glia"). A. Glutamate was applied at 50 μM for 10 min, then chased for the times indicated (h). B. Glutamate was applied at 20 μM continuously for the times indicated (h). β-gal activity was determined by a luminescent assay. Values represent activity relative to untreated cultures ± SEM in quadruplicate cultures. ANOVA followed by Scheffe post-hoc test showed significance between control and each treatment, and between the "+glia" and "-glia" in all comparisons except the 6 h timepoint in A. (p < 0.02).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2693111&req=5

Figure 2: Glutamate is unable to activate neuronal NFκB in mixed cell cultures. Neocortical neurons from κB/β-gal transgenic mice were either plated on the top of wild-type astrocytes ("+glia") or retained as nearly pure neuronal cultures ("-glia"). A. Glutamate was applied at 50 μM for 10 min, then chased for the times indicated (h). B. Glutamate was applied at 20 μM continuously for the times indicated (h). β-gal activity was determined by a luminescent assay. Values represent activity relative to untreated cultures ± SEM in quadruplicate cultures. ANOVA followed by Scheffe post-hoc test showed significance between control and each treatment, and between the "+glia" and "-glia" in all comparisons except the 6 h timepoint in A. (p < 0.02).
Mentions: We have further analyzed in vitro neurons from the transgenic line developed by Bhakar et al. [33]. The main objective was to explore the possibility that pure neuronal cultures are altered in their handling of NFκB due to the absence of a physical interaction with glia. To test this idea, we cultured neurons from the κB/β-gal reporter line with wild-type glia so that typical neuron-glia interactions could take place, while the reporting of NFκB activity would be specific to the neurons in the coculture. Wild-type astrocytes were first plated and grown to confluency, and then E17 transgenic (κB/β-gal) cortical cells were added one week prior to treatment. The pre-plated wild-type astrocytes suppressed growth of transgenic glia but promoted survival of the κB/β-gal neurons, as histochemistry showed no astrocytes positive for β-gal. These cocultures were compared to cultures of κB/β-gal neurons alone. Two treatment paradigms were tested: 1) a transient exposure to glutamate (10 min) followed by a 6- to 24-hour chase period, or 2) continuous exposure to glutamate for 6 to 24 hours (Figure 2). Once again, neuronally localized NFκB activity could not be induced by glutamate. Indeed, basal β-gal levels were suppressed by glutamate exposure, an effect that was partially suppressed in the presence of glia, presumably due to the ability of astrocytes to metabolize glutamate. This negative effect of glutamate on κB-dependent transcription is consistent with copious EMSA data showing that glutamate reduces the levels of κB-binding Sp1-related factors in neurons (below).

Bottom Line: These studies led us to conclude that this population of cells is nearly incapable of activating the NFkappaB that is nonetheless expressed at reasonable levels.A subset of the kappaB cis elements are instead bound by members of the Sp1 family in neurons.Also surprising was our discovery that Sp1 itself, typically described as ubiquitous, is severely restricted in expression within forebrain neurons; Sp4 seems to be substituted during neuronal differentiation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anesthesiology, Washington University School of Medicine, St Louis, MO 63110, USA. maox@morpheus.wustl.edu

ABSTRACT
The unique physiology and function of neurons create differences in their cellular physiology, including their regulation of gene expression. We began several years ago exploring the relationships between the NFkappaB transcription factor, neuronal survival, and glutamate receptor activation in telencephalic neurons. These studies led us to conclude that this population of cells is nearly incapable of activating the NFkappaB that is nonetheless expressed at reasonable levels. A subset of the kappaB cis elements are instead bound by members of the Sp1 family in neurons. Also surprising was our discovery that Sp1 itself, typically described as ubiquitous, is severely restricted in expression within forebrain neurons; Sp4 seems to be substituted during neuronal differentiation. These findings and their implications for neuronal differentiation--as well as potential dedifferentiation during degenerative processes--are discussed here.

Show MeSH
Related in: MedlinePlus