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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 localizes the Arc-β-Spectrin-Tip60 complex to the perichromatin region. A, Fluorescently-tagged versions of Arc (green), β-Spectrin (red), and Tip60 (blue) were coexpressed in HEK293 cells and imaged as described in Fig. 3. Because of their strict overlap, Arc-β-Spectrin-Tip60 complexes appear as white puncta. The DAPI signal is shown in magenta. The insets (B−E) show the relationship between individual complexes and chromatin with higher magnification. Segmentation of the DAPI structures is indicated with a magenta line. B shows a complex in the center of an interchromatin domain. C and D show complexes localized to the perichromatin region, while E illustrates a complex surrounded by dense DAPI staining. Analysis of five nuclei indicated that the majority of puncta were localized to the perichromatin region (84%, n = 151), while the puncta were less frequently observed in the center of the interchromatin region (9%, n = 16) or in densely packed chromatin (7%, n = 12). F, Tip60-YFP was expressed in HEK293 cells, which were fixed as stained for DNA by DAPI. Tip60 occupies large parts of the interchromatin domains. Insets (G−J) show individual Tip60 speckles in higher magnification. Not the larger size of Tip60 compared to the tripartite complexes shown in B−E. K, Arc-YFP was expressed in HEK293 cells, which were fixed and stained with DAPI. Many small Arc puncta are seen, which associate with the interface of the IC domains and the dense chromatin strongly labeled with DAPI, indicated with a red line that segments the DAPI structure. Insets (L−M) show the relationship of Arc puncta with chromatin at higher magnification. P, Arc-YFP was expressed in hippocampal neurons (18 DIV), which were fixed and labeled with DAPI. White arrows indicate Arc puncta that localize to the interchromatin -chromatin interface indicated with the red line. Insets (Q−T) show Arc localization in higher magnification.
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f4: Arc localizes the Arc-β-Spectrin-Tip60 complex to the perichromatin region. A, Fluorescently-tagged versions of Arc (green), β-Spectrin (red), and Tip60 (blue) were coexpressed in HEK293 cells and imaged as described in Fig. 3. Because of their strict overlap, Arc-β-Spectrin-Tip60 complexes appear as white puncta. The DAPI signal is shown in magenta. The insets (B−E) show the relationship between individual complexes and chromatin with higher magnification. Segmentation of the DAPI structures is indicated with a magenta line. B shows a complex in the center of an interchromatin domain. C and D show complexes localized to the perichromatin region, while E illustrates a complex surrounded by dense DAPI staining. Analysis of five nuclei indicated that the majority of puncta were localized to the perichromatin region (84%, n = 151), while the puncta were less frequently observed in the center of the interchromatin region (9%, n = 16) or in densely packed chromatin (7%, n = 12). F, Tip60-YFP was expressed in HEK293 cells, which were fixed as stained for DNA by DAPI. Tip60 occupies large parts of the interchromatin domains. Insets (G−J) show individual Tip60 speckles in higher magnification. Not the larger size of Tip60 compared to the tripartite complexes shown in B−E. K, Arc-YFP was expressed in HEK293 cells, which were fixed and stained with DAPI. Many small Arc puncta are seen, which associate with the interface of the IC domains and the dense chromatin strongly labeled with DAPI, indicated with a red line that segments the DAPI structure. Insets (L−M) show the relationship of Arc puncta with chromatin at higher magnification. P, Arc-YFP was expressed in hippocampal neurons (18 DIV), which were fixed and labeled with DAPI. White arrows indicate Arc puncta that localize to the interchromatin -chromatin interface indicated with the red line. Insets (Q−T) show Arc localization in higher magnification.

Mentions: As shown in the insets of Figure 2, A and B, Arc and βSpIVΣ5 puncta localize to interchromatin domains, but are often found adjacent to DAPI-dense structures. Figure 4 illustrates the relationship of the Arc-βSpIVΣ5-Tip60 complex with DNA labeled by DAPI staining (Fig. 4A), and compares this with the localization of Tip60 (Fig. 4F) and Arc (Fig. 4K) alone, following expression of GFP fusion constructs in HEK293 cells. Arc-βSpIVΣ5-Tip60c complexes appear as white puncta due to the tight colocalization of the three components. Many of these complexes localize to the interface between the interchromatin space, which is devoid of DNA, and the compact chromatin domains, which is stained heavily by DAPI. This interface is sometimes referred to as the perichromatin region, and it has important functions, including transcription and RNA processing (Fakan and van Driel, 2007; Niedojadlo et al., 2011). In several instances, the Arc-βSpIVΣ5-Tip60 complexes appear to be engulfed by a small amount of DAPI, indicative of loosely packed DNA characteristic of perichromatin regions. In Figure 4, the interchromatin domains are delineated by a line created by segmenting the DAPI structure to show various locations of the tripartite complex puncta: in the center of an interchromatin domain (Fig. 4B), adjacent to or embedded in the perichromatin region (Fig. 4C,D), and completely surround by chromatin (Fig. 4E). The Arc-containing complexes are seen only rarely at locations B and E, and predominantly localize to the perichromatin regions CD.


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 localizes the Arc-β-Spectrin-Tip60 complex to the perichromatin region. A, Fluorescently-tagged versions of Arc (green), β-Spectrin (red), and Tip60 (blue) were coexpressed in HEK293 cells and imaged as described in Fig. 3. Because of their strict overlap, Arc-β-Spectrin-Tip60 complexes appear as white puncta. The DAPI signal is shown in magenta. The insets (B−E) show the relationship between individual complexes and chromatin with higher magnification. Segmentation of the DAPI structures is indicated with a magenta line. B shows a complex in the center of an interchromatin domain. C and D show complexes localized to the perichromatin region, while E illustrates a complex surrounded by dense DAPI staining. Analysis of five nuclei indicated that the majority of puncta were localized to the perichromatin region (84%, n = 151), while the puncta were less frequently observed in the center of the interchromatin region (9%, n = 16) or in densely packed chromatin (7%, n = 12). F, Tip60-YFP was expressed in HEK293 cells, which were fixed as stained for DNA by DAPI. Tip60 occupies large parts of the interchromatin domains. Insets (G−J) show individual Tip60 speckles in higher magnification. Not the larger size of Tip60 compared to the tripartite complexes shown in B−E. K, Arc-YFP was expressed in HEK293 cells, which were fixed and stained with DAPI. Many small Arc puncta are seen, which associate with the interface of the IC domains and the dense chromatin strongly labeled with DAPI, indicated with a red line that segments the DAPI structure. Insets (L−M) show the relationship of Arc puncta with chromatin at higher magnification. P, Arc-YFP was expressed in hippocampal neurons (18 DIV), which were fixed and labeled with DAPI. White arrows indicate Arc puncta that localize to the interchromatin -chromatin interface indicated with the red line. Insets (Q−T) show Arc localization in higher magnification.
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f4: Arc localizes the Arc-β-Spectrin-Tip60 complex to the perichromatin region. A, Fluorescently-tagged versions of Arc (green), β-Spectrin (red), and Tip60 (blue) were coexpressed in HEK293 cells and imaged as described in Fig. 3. Because of their strict overlap, Arc-β-Spectrin-Tip60 complexes appear as white puncta. The DAPI signal is shown in magenta. The insets (B−E) show the relationship between individual complexes and chromatin with higher magnification. Segmentation of the DAPI structures is indicated with a magenta line. B shows a complex in the center of an interchromatin domain. C and D show complexes localized to the perichromatin region, while E illustrates a complex surrounded by dense DAPI staining. Analysis of five nuclei indicated that the majority of puncta were localized to the perichromatin region (84%, n = 151), while the puncta were less frequently observed in the center of the interchromatin region (9%, n = 16) or in densely packed chromatin (7%, n = 12). F, Tip60-YFP was expressed in HEK293 cells, which were fixed as stained for DNA by DAPI. Tip60 occupies large parts of the interchromatin domains. Insets (G−J) show individual Tip60 speckles in higher magnification. Not the larger size of Tip60 compared to the tripartite complexes shown in B−E. K, Arc-YFP was expressed in HEK293 cells, which were fixed and stained with DAPI. Many small Arc puncta are seen, which associate with the interface of the IC domains and the dense chromatin strongly labeled with DAPI, indicated with a red line that segments the DAPI structure. Insets (L−M) show the relationship of Arc puncta with chromatin at higher magnification. P, Arc-YFP was expressed in hippocampal neurons (18 DIV), which were fixed and labeled with DAPI. White arrows indicate Arc puncta that localize to the interchromatin -chromatin interface indicated with the red line. Insets (Q−T) show Arc localization in higher magnification.
Mentions: As shown in the insets of Figure 2, A and B, Arc and βSpIVΣ5 puncta localize to interchromatin domains, but are often found adjacent to DAPI-dense structures. Figure 4 illustrates the relationship of the Arc-βSpIVΣ5-Tip60 complex with DNA labeled by DAPI staining (Fig. 4A), and compares this with the localization of Tip60 (Fig. 4F) and Arc (Fig. 4K) alone, following expression of GFP fusion constructs in HEK293 cells. Arc-βSpIVΣ5-Tip60c complexes appear as white puncta due to the tight colocalization of the three components. Many of these complexes localize to the interface between the interchromatin space, which is devoid of DNA, and the compact chromatin domains, which is stained heavily by DAPI. This interface is sometimes referred to as the perichromatin region, and it has important functions, including transcription and RNA processing (Fakan and van Driel, 2007; Niedojadlo et al., 2011). In several instances, the Arc-βSpIVΣ5-Tip60 complexes appear to be engulfed by a small amount of DAPI, indicative of loosely packed DNA characteristic of perichromatin regions. In Figure 4, the interchromatin domains are delineated by a line created by segmenting the DAPI structure to show various locations of the tripartite complex puncta: in the center of an interchromatin domain (Fig. 4B), adjacent to or embedded in the perichromatin region (Fig. 4C,D), and completely surround by chromatin (Fig. 4E). The Arc-containing complexes are seen only rarely at locations B and E, and predominantly localize to the perichromatin regions CD.

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.