Author: Brittany Shimanski
https://www.nature.com/articles/nature22405.pdf
Epigenetic modifications have been identified in cognitive processing, specifically in learning and memory. Lack of histone acetylation has been implicated in neurodevelopmental disorders, neurodegeneration, and aging.1 Expression of neuronal genes that are involved in chromatin modification processes, such as histone acetylation, have important involvement in storing and forming memories, mostly in the hippocampus. Transcription factor CREB, coactivator CREB binding protein (CBP)2, particularly histone acetyltransferase (HAT)3, and histone deacetylase inhibitors are all known to be required and helpful for consolidation of memories, though the mechanisms of histone acetylation are unknown. As known from Biochemistry 441, acetyl CoA has various roles in bodily metabolism in different pathways, so its importance in another mechanism of our body is not surprising. Sensing of metabolites, such as acetyl CoA levels, by acetyltransferases can alter the structure of chromatin and the expression of genes. Therefore, the enzymes responsible for the making of acetyl CoA, acetate-dependent acetyl-CoA synthetase 2 (ACSS2) and citrate-dependent ATP-citrate lyase (ACL), control the acetylation of histones as well as gene expression, but their role in post-mitotic neuronal cells and specifically in memory function is not established.
Mews et al. used the Cath-a-differentiated (CAD) cell line, a neuronal cell line from a mouse that demonstrates histochemical fluorescence, to look into the function of ACSS2. Earlier in Biochemistry 441, we read an article discussing a different type of cell differentiation and its role in metabolism.
ACSS2 remained in the cytoplasm when undifferentiated, but the moved to the nucleus and increased once differentiated from CAD cells into neurons, while ACL expression remained constant in the cytoplasm4. When neuron-specific protein markers were upregulated in differentiated neuronal cells, pre-differentiation knockdown of ACSS2 reduced expression of nuclear markers and ACL was not decreased, therefore ACSS2 has a significant role in neuronal differentiation. mRNA-seq identified 894 upregulated genes upon CAD neuronal differentiation and gene ontology analysis identified these genes were specific to each neuron4.
Mews et al. examined the acetylation of histones during differentiation by using chromatin immunoprecipitation with high-throughput DNA Sequencing (ChIP-sep) for histone H3 lysine 9 acetylation (H3K9ac), H4K5ac, and H4K12ac. The results indicated the 894 genes that were upregulated showed high levels of acetylation than all others. When reducing levels of ACSS2 and ACL using short hairpin RNAs before differentiation and RNA-seq and discovered that when ACSS2 was knocked down, levels of upregulation were lowered and showed a genome-wide defect, and when ACL was knocked down, there was no change in upregulation and cells showed lower global transcript levels4. The authors concluded that ACL has a non-specific effect on gene expression while ACSS2 has a specific requirement for upregulation of gene expression when CAD cells are differentiating into neurons4.
Mews et al. then used ChIP-seq to access the connection between ACSS2 and chromatin, and found a strong correlation using ACSS2 antibodies. The binding of ACSS2 correlated with an increase in histones in differentiated CAD cells. Being that genes proximal to ACSS2 peaks were linked to cell differentiation, chromatin-associated, neuronal gene promoter-proximal ACSS2 could very well be a source of acetyl-CoA to HAT enzymes4. ACSS2 binding was found to be connected to histone acetylation due to ACSS2 peaks directly overlapping with H3 and H4 acetylation peaks and the height of the peaks were correlating with each other, suggesting that H4 acetylation is the most responsive to acetyl CoA from ACSS2, specifically H4K12ac which is interestingly linked to ineffective forming of memories.5 The overlapping of peaks with different genes involved in memory are shown in the figure. Recruitment of ACSS2 by transcription factor binding demonstrated that the most enriched motif recruits CBP and E1A binding protein (p300), which are both acetyl-CoA dependent, meaning that ACSS2 in involved in HAT activity nearby. In fact, genes with the largest change in differential AcSS2 binding had the highest levels of histone acetylation and were neuron-specific. The CAD cell genomic data demonstrated that the enrichment of ACSS2 in differentiated neuronal cells is linked to higher levels of acetylation of histones and an involvement the upregulation of neuronal genes4.
Acetyl CoA levels in knocked down ACSS2 cells with ACSS2 inhibitor were decreased, supporting the idea that ACSS2 is a supplier to acetyl-CoA. Knocked down cells also showed reduced global histone acetylation levels in markers with transcriptional coactivators (CBP and P300) with roles in long term memory. ACSS2, CBP, and acetylated chromatin co-immunoprecipitated, showing that ACSS2 binds chromatin at active genes to drive the increase in the acetylation of histones when memories are forming4. ACSS2 inhibitor treatment reduced marker expression and histone acetylation, but didn’t lower ACSS2 levels or the ACL levels in mouse neurons. Using ChIP-seq, ACSS2 in the hippocampus and H3K9ac corresponded over three neuronal genes involved in memory formation. Genes enriched for H3k9ac were transcribed, but if enriched for ACSS2 and H3k9ac only then did they have the highest levels of expression4.
Spatial memory in the hippocampus occurs through epigenetic modifications, such as histone acetylation. ACSS2 could mediate histone acetylation in order to upregulate gene expression during the consolidation of memories6. ACSS2 knockdown mice were found to have impaired spatial object memory, a lower discrimination index, and reduced total object exploration during an object-location memory test4. The authors concluded that AcSS2 has a role in dorsal hippocampus-meditated long-term spatial memory. Being that long-term spatial memory requires an increase in histone acetylation and immediate gene transcription and memory consolidation can be inhibited by stopping mRNA synthesis, Mews et al. used mRNA-seq on the dorsal hippocampus to access whether ACSS2 was involved in gene upregulation for hippocampal memory consolidation. Upregulation of early genes during the sensitive memory consolidation period was reduced when ACSS2 was knocked down. In addition, induction of early genes during the sensitive period was stopped by the knock down of ACSS2.4
In conclusion, the authors demonstrated a link between metabolism and neuronal plasticity, showing the function of ACSS2 as a chromatin-bound transcriptional coactivator, stimulating histone acetylation and upregulating gene expression, required for spatial memory. Post-mitotic neurons have a reliance on ACSS2 to be a supplier of the acetyl-CoA for the acetylation of histones, as HAT enzymes are controlled by changing acetyl CoA levels. The authors also show the role of ACSS2 of upregulation in early genes which can lead to a role in long-term memory consolidation. Understanding of epigenetic mechanisms that regulate neural functions can be linked to a better understanding of neuropsychiatric diseases. In fact, drug abuse is even linked to altering chromatin states7, so this new information can potentially be used to help in understanding drug addiction and treatment. Due to the complexity of understanding and memory, ACSS2 can be identified as enzymatic target to develop therapeutic agents to help to treat various different neurological and cognitive disorders, especially involving memory and learning.
References:
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(3) Korzus, E.; Rosenfeld, M. G.; Mayford, M. CBP Histone Acetyltransferase Activity Is a Critical Component of Memory Consolidation. Neuron 2004, 42 (6), 961–972.
(4) Mews, P.; Donahue, G.; Drake, A. M.; Luczak, V.; Abel, T.; Berger, S. L. Acetyl-CoA Synthetase Regulates Histone Acetylation and Hippocampal Memory. Nature 2017, 546 (7658), 381–386.
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(6) Barrett, R. M.; Malvaez, M.; Kramar, E.; Matheos, D. P.; Arrizon, A.; Cabrera, S. M.; Lynch, G.; Greene, R. W.; Wood, M. A. Hippocampal Focal Knockout of CBP Affects Specific Histone Modifications, Long-Term Potentiation, and Long-Term Memory. Neuropsychopharmacology 2011, 36 (8), 1545–1556.
(7) Walker, D. M.; Cates, H. M.; Heller, E. A.; Nestler, E. J. Regulation of Chromatin States by Drugs of Abuse. Curr. Opin. Neurobiol. 2015, 30, 112–121.
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