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Nuclear Arc Interacts with the Histone Acetyltransferase Tip60 to Modify H4K12 Acetylation(1,2,3).

Wee CL, Teo S, Oey NE, Wright GD, VanDongen HM, VanDongen AM - eNeuro (2014)

Bottom Line: Although Arc protein is found at synapses, it also localizes to the neuronal nucleus, where its function is less understood.Neuronal activity-induced expression of Arc (1) increases endogenous nuclear Tip60 puncta, (2) recruits Tip60 to PML bodies, and (3) increases histone acetylation of Tip60 substrate H4K12, a learning-induced chromatin modification.These mechanisms point to an epigenetic role for Arc in regulating memory consolidation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Program in Neuroscience and Behavioral Disorders, Duke-NUS Graduate Medical School , Singapore 169857.

ABSTRACT
Arc is an immediate-early gene whose genetic ablation selectively abrogates long-term memory, indicating a critical role in memory consolidation. Although Arc protein is found at synapses, it also localizes to the neuronal nucleus, where its function is less understood. Nuclear Arc forms a complex with the β-spectrin isoform βSpIVΣ5 and associates with PML bodies, sites of epigenetic regulation of gene expression. We report here a novel interaction between Arc and Tip60, a histone-acetyltransferase and subunit of a chromatin-remodelling complex, using biochemistry and super-resolution microscopy in primary rat hippocampal neurons. Arc and βSpIVΣ5 are recruited to nuclear Tip60 speckles, and the three proteins form a tight complex that localizes to nuclear perichromatin regions, sites of transcriptional activity. Neuronal activity-induced expression of Arc (1) increases endogenous nuclear Tip60 puncta, (2) recruits Tip60 to PML bodies, and (3) increases histone acetylation of Tip60 substrate H4K12, a learning-induced chromatin modification. These mechanisms point to an epigenetic role for Arc in regulating memory consolidation.

No MeSH data available.


Arc and βSpIVΣ5 interact with Tip60. A, Tip60 interacts with Arc. HEK293T cells were transfected with either Arc in pCDNA3.1 and YFP vector, or Arc-pCDNA3.1 and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc was detected with a rabbit anti-Arc antibody. B, Tip60 interacts with βSpIVΣ5. HEK293T cells were transfected with either βSpIVΣ5-HA and YFP vector or βSpIVΣ5-HA and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and βSpIVΣ5-HA was detected with a mouse anti-HA antibody. C, Tip60 interacts with both Arc and βSpIVΣ5. HEK293T cells were transfected with Arc, βSpIVΣ5-HA, and either YFP or Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc or βSpIVΣ5-HA was detected with a rabbit anti-Arc or mouse anti-HA antibody respectively. The asterix (*) denotes an unidentified band.
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f5: Arc and βSpIVΣ5 interact with Tip60. A, Tip60 interacts with Arc. HEK293T cells were transfected with either Arc in pCDNA3.1 and YFP vector, or Arc-pCDNA3.1 and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc was detected with a rabbit anti-Arc antibody. B, Tip60 interacts with βSpIVΣ5. HEK293T cells were transfected with either βSpIVΣ5-HA and YFP vector or βSpIVΣ5-HA and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and βSpIVΣ5-HA was detected with a mouse anti-HA antibody. C, Tip60 interacts with both Arc and βSpIVΣ5. HEK293T cells were transfected with Arc, βSpIVΣ5-HA, and either YFP or Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc or βSpIVΣ5-HA was detected with a rabbit anti-Arc or mouse anti-HA antibody respectively. The asterix (*) denotes an unidentified band.

Mentions: The colocalization seen in the imaging results discussed above suggests that Tip60 can bind to both Arc and βSpIVΣ5. To test this idea more directly, we performed coimmunoprecipitation experiments (Fig. 5). Arc was coexpressed in HEK293 cells with either Tip60-YFP, or YFP as a negative control. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody; the precipitate was resolved by SDS-PAGE and transferred to a nitrocellulose filter, where Arc was detected with a rabbit anti-Arc antibody. These experiments showed that Arc coimmunoprecipitated with Tip60 (Fig. 5A). Tip60 was also found to bind to βSpIVΣ5. HEK293 cells were transfected with either βSpIVΣ5-HA and YFP (negative control) or βSpIVΣ5-HA and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and βSpIVΣ5-HA was detected with a mouse anti-HA antibody (Fig. 5B). Finally, Tip60 was seen to physically interact with the complex formed by Arc and βSpIVΣ5: HEK293 cells were transfected with Arc, βSpIVΣ5-HA, and either YFP or Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc or βSpIVΣ5-HA was detected with a rabbit anti-Arc or mouse anti-HA antibody, respectively (Fig. 5C), suggesting that complex formation between Arc and βSpIVΣ5 does not interfere with Tip60 binding.


Nuclear Arc Interacts with the Histone Acetyltransferase Tip60 to Modify H4K12 Acetylation(1,2,3).

Wee CL, Teo S, Oey NE, Wright GD, VanDongen HM, VanDongen AM - eNeuro (2014)

Arc and βSpIVΣ5 interact with Tip60. A, Tip60 interacts with Arc. HEK293T cells were transfected with either Arc in pCDNA3.1 and YFP vector, or Arc-pCDNA3.1 and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc was detected with a rabbit anti-Arc antibody. B, Tip60 interacts with βSpIVΣ5. HEK293T cells were transfected with either βSpIVΣ5-HA and YFP vector or βSpIVΣ5-HA and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and βSpIVΣ5-HA was detected with a mouse anti-HA antibody. C, Tip60 interacts with both Arc and βSpIVΣ5. HEK293T cells were transfected with Arc, βSpIVΣ5-HA, and either YFP or Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc or βSpIVΣ5-HA was detected with a rabbit anti-Arc or mouse anti-HA antibody respectively. The asterix (*) denotes an unidentified band.
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Related In: Results  -  Collection

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f5: Arc and βSpIVΣ5 interact with Tip60. A, Tip60 interacts with Arc. HEK293T cells were transfected with either Arc in pCDNA3.1 and YFP vector, or Arc-pCDNA3.1 and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc was detected with a rabbit anti-Arc antibody. B, Tip60 interacts with βSpIVΣ5. HEK293T cells were transfected with either βSpIVΣ5-HA and YFP vector or βSpIVΣ5-HA and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and βSpIVΣ5-HA was detected with a mouse anti-HA antibody. C, Tip60 interacts with both Arc and βSpIVΣ5. HEK293T cells were transfected with Arc, βSpIVΣ5-HA, and either YFP or Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc or βSpIVΣ5-HA was detected with a rabbit anti-Arc or mouse anti-HA antibody respectively. The asterix (*) denotes an unidentified band.
Mentions: The colocalization seen in the imaging results discussed above suggests that Tip60 can bind to both Arc and βSpIVΣ5. To test this idea more directly, we performed coimmunoprecipitation experiments (Fig. 5). Arc was coexpressed in HEK293 cells with either Tip60-YFP, or YFP as a negative control. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody; the precipitate was resolved by SDS-PAGE and transferred to a nitrocellulose filter, where Arc was detected with a rabbit anti-Arc antibody. These experiments showed that Arc coimmunoprecipitated with Tip60 (Fig. 5A). Tip60 was also found to bind to βSpIVΣ5. HEK293 cells were transfected with either βSpIVΣ5-HA and YFP (negative control) or βSpIVΣ5-HA and Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and βSpIVΣ5-HA was detected with a mouse anti-HA antibody (Fig. 5B). Finally, Tip60 was seen to physically interact with the complex formed by Arc and βSpIVΣ5: HEK293 cells were transfected with Arc, βSpIVΣ5-HA, and either YFP or Tip60-YFP. Tip60-YFP was immunoprecipitated with a mouse anti-GFP antibody, and Arc or βSpIVΣ5-HA was detected with a rabbit anti-Arc or mouse anti-HA antibody, respectively (Fig. 5C), suggesting that complex formation between Arc and βSpIVΣ5 does not interfere with Tip60 binding.

Bottom Line: Although Arc protein is found at synapses, it also localizes to the neuronal nucleus, where its function is less understood.Neuronal activity-induced expression of Arc (1) increases endogenous nuclear Tip60 puncta, (2) recruits Tip60 to PML bodies, and (3) increases histone acetylation of Tip60 substrate H4K12, a learning-induced chromatin modification.These mechanisms point to an epigenetic role for Arc in regulating memory consolidation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Program in Neuroscience and Behavioral Disorders, Duke-NUS Graduate Medical School , Singapore 169857.

ABSTRACT
Arc is an immediate-early gene whose genetic ablation selectively abrogates long-term memory, indicating a critical role in memory consolidation. Although Arc protein is found at synapses, it also localizes to the neuronal nucleus, where its function is less understood. Nuclear Arc forms a complex with the β-spectrin isoform βSpIVΣ5 and associates with PML bodies, sites of epigenetic regulation of gene expression. We report here a novel interaction between Arc and Tip60, a histone-acetyltransferase and subunit of a chromatin-remodelling complex, using biochemistry and super-resolution microscopy in primary rat hippocampal neurons. Arc and βSpIVΣ5 are recruited to nuclear Tip60 speckles, and the three proteins form a tight complex that localizes to nuclear perichromatin regions, sites of transcriptional activity. Neuronal activity-induced expression of Arc (1) increases endogenous nuclear Tip60 puncta, (2) recruits Tip60 to PML bodies, and (3) increases histone acetylation of Tip60 substrate H4K12, a learning-induced chromatin modification. These mechanisms point to an epigenetic role for Arc in regulating memory consolidation.

No MeSH data available.