Acetyl-CoA flux regulates the proteome and acetyl-proteome to maintain intracellular metabolic crosstalk - Acetylation Stoichiometry Dataset
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Q Exactive
ORGANISM(S): Mus Musculus (ncbitaxon:10090)
SUBMITTER: 
John Denu Luigi Puglielli
PROVIDER: MSV000083855 | MassIVE | Fri May 24 13:18:00 BST 2019
SECONDARY ACCESSION(S): PXD014013
REPOSITORIES: MassIVE
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Project description:AT-1/SLC33A1 is a key member of the endoplasmic reticulum (ER) acetylation machinery, transporting acetyl-CoA from the cytosol into the ER lumen where acetyl-CoA serves as the acetyl-group donor for Nε-lysine acetylation. Dysfunctional ER acetylation, as caused by heterozygous or homozygous mutations as well as gene duplication events of AT-1/SLC33A1, has been linked to both developmental and degenerative diseases. Here, we investigate two models of AT-1 dysregulation and altered acetyl-CoA flux: AT-1S113R/+ mice, a model of AT-1 haploinsufficiency, and AT-1 sTg mice, a model of AT-1 overexpression. The animals display distinct metabolic adaptation across intracellular compartments, including reprogramming of lipid metabolism and mitochondria bioenergetics. Mechanistically, the perturbations to AT-1-dependent acetyl-CoA flux result in global and specific changes in both the proteome and the acetyl-proteome (protein acetylation). Collectively, our results suggest that AT-1 acts as an important metabolic regulator that maintains acetyl-CoA homeostasis by promoting functional crosstalk between different intracellular organelles.
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Project description:The acetyl-CoA transporter, AT-1 (also referred to as SLC33A1), is a key member of the endoplasmic reticulum (ER) acetylation machinery; it transports acetyl-CoA from the cytosol into the ER lumen where it serves as donor of the acetyl group for Nε-lysine acetylation 1,2. Dysfunctional ER acetylation, as caused by heterozygous or homozygous mutations as well as gene duplication events of AT-1/SLC33A1, has been linked to both developmental and age-associated human diseases 3-7. Mice with reduced or increased AT-1 expression mimic associated human diseases 8-10. In this study, we investigated the pervasive effects that dysregulated AT-1 has on intracellular acetyl-CoA homeostasis. Specifically, we used AT-1S113R/+ mice 8, a model of AT-1 haploinsufficiency, and AT-1 sTg mice 10, a model of AT-1 overexpression. We found that reduced AT-1 activity in AT-1S113R/+ mice led to increased availability of acetyl-CoA in the cytosol and spontaneous steatosis. Conversely, increased AT-1 activity decreased the availability of acetyl-CoA in the cytosol and made the animals resistant to diet-induced steatosis. Both models displayed significant metabolic adaptation involving different cellular organelles and compartments. Mechanistically, the metabolic adaptation was driven by changes in both protein levels (proteome) and stoichiometry of acetylation (acetylome) within fundamental pathways. Collectively, our results suggest that AT-1 acts as an important metabolic regulator that maintains acetyl-CoA homeostasis by promoting functional “cross-talk” between different intracellular organelles and compartments.
2019-08-05 | PXD013736 | Pride
Project description:Nε-lysine acetylation has emerged as a central mechanism to maintain quality control and protein homeostasis within the Endoplasmic Reticulum (ER) and secretory pathway. The ER acetylation machinery includes AT-1/SLC33A1, a membrane transporter that translocates acetyl-CoA from the cytosol to the ER lumen, and ATase1 and ATase2, two ER membrane-bound acetyltransferases that acetylate ER cargo proteins. Dysfunctional AT-1, as caused by loss-of-function mutations or gene duplication events, results in neurodevelopmental or neurodegenerative disorders. Experiments in our lab have demonstrated that these human diseases can be effectively recapitulated in mouse models. In this thesis, we used two models of dysregulated acetyl-CoA flux: AT-1S113R/+, a model of AT-1 haploinsufficiency, and AT-1 sTg, a model of systemic overexpression of AT-1. First, we examined upstream processes of cytosol-to-ER acetyl-CoA flux by evaluating the cytosolic pool of acetyl-CoA. The aberrant AT-1 models demonstrated distinct metabolic reprogramming of lipid metabolism and mitochondria bioenergetics. Dysregulated acetyl-CoA flux resulted in global changes at the level of the proteome and the acetyl-proteome. Second, we examined the downstream consequences of cytosol-to-ER acetyl-CoA flux. Specifically, we investigated the engagement and functional organization of the secretory pathway and used N-glycoproteomics to determine the quality of secreted proteins. Aberrant AT-1 models demonstrated reorganization of the ER, Golgi, and ERGIC, as well as a delay in glycoproteins clearing the Golgi apparatus. Additionally, AT-1 sTg mice showed a marked delay in protein trafficking to the cell surface. The N-glycoproteome revealed significant alterations, highlighting changes in the quality of the secretome.
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Project description:Metabolic production of acetyl coenzyme A (acetyl-CoA) is linked to histone acetylation and gene regulation, but the precise mechanisms of this process are largely unknown. Here we show that the metabolic enzyme acetyl-CoA synthetase 2 (ACSS2) directly regulates histone acetylation in neurons and spatial memory in mammals. In a neuronal cell culture model, ACSS2 increases in the nuclei of differentiating neurons and localizes to upregulated neuronal genes near sites of elevated histone acetylation. A decrease in ACSS2 lowers nuclear acetyl-CoA levels, histone acetylation, and responsive expression of the cohort of neuronal genes. In adult mice, attenuation of hippocampal ACSS2 expression impairs long-term spatial memory, a cognitive process that relies on histone acetylation. A decrease in ACSS2 in the hippocampus also leads to defective upregulation of memory-related neuronal genes that are pre-bound by ACSS2. These results reveal a connection between cellular metabolism, gene regulation, and neural plasticity and establish a link between acetyl-CoA generation 'on-site' at chromatin for histone acetylation and the transcription of key neuronal genes.
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Project description:Cellular metabolism plays a crucial role in controlling the proliferation, differentiation, and quiescence of neural stem cells (NSCs). The metabolic transition from aerobic glycolysis to oxidative phosphorylation has been regarded as a hallmark of neuronal differentiation. Understanding what triggers metabolism reprogramming and how glucose metabolism directs NSC differentiation may provide new insight into the regenerative potential of the brain. TP53 inducible glycolysis and apoptosis regulator (TIGAR) is an endogenous inhibitor of glycolysis and is highly expressed in mature neurons. However, its function in embryonic NSCs has not yet been explored. In this study, we aimed to investigate the precise roles of TIGAR in NSCs and the possible involvement of metabolic reprogramming in the TIGAR regulatory network. We observed that TIGAR is significantly increased during brain development as neural differentiation proceeds, especially at the peak of NSC differentiation (E14.5-E16.5). In cultured NSCs, knockdown of TIGAR reduced the expression of microtubule-associated protein 2 (MAP2), neuron-specific class III beta-tubulin (Tuj1), glial fibrillary acidic protein (GFAP), Ngn1, and NeuroD1, and enhanced the expression of REST, suggesting that TIGAR is an important regulator of NSC differentiation. Furthermore, TIGAR enhanced the expression of lactate dehydrogenase B (LDHB) and the mitochondrial biogenesis and oxidative phosphorylation (OXPHOS) markers, peroxisome proliferator-activated receptor gamma coactivator 1 (PGC-1?), nuclear respiratory factor (NRF1), and MitoNEET during NSC differentiation. TIGAR can decrease lactate production and accelerate oxygen consumption and ATP generation to maintain a high rate of OXPHOS in differentiated NSCs. Interestingly, knockdown of TIGAR decreased the level of acetyl-CoA and H3K9 acetylation at the promoters of Ngn1, Neurod1, and Gfap. Acetate, a precursor of acetyl-CoA, increased the level of H3K9 acetylation and rescued the effect of TIGAR deficiency on NSC differentiation. Together, our data demonstrated that TIGAR promotes metabolic reprogramming and regulates NSC differentiation through an epigenetic mechanism.
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