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Using dynamic gene module map analysis to identify targets that modulate free fatty acid induced cytotoxicity.

ABSTRACT: The objective of this study was to identify pathways that regulate the cytotoxicity induced by free fatty acids (FFAs) in human hepatoblastoma cells (HepG2/C3A). Gene expression profiles of HepG2/C3A cells were obtained at three time points, after 24, 48, and 72 h of exposure to different types of FFA. Saturated fatty acid (palmitate) was found to be cytotoxic. The pathways activated by the different FFAs at the different time points were identified using global gene module map analysis. Unsaturated FFAs exerted transcriptional regulation mainly within the first 24 h, whereas saturated FFA, palmitate, regulated energy production pathways, such as the electron transport chain (ETC) and tricarboxylic acid cycle, within the first 24 h. In the next 24 h, palmitate up-regulated 36 cell death relevant pathways and down-regulated several protective pathways, such as the pentose phosphate pathway and glutathione-related pathways. In the final 24 h, the FFAs did not induce significant transcriptional regulation. We hypothesized that palmitate induced cytotoxicity by first perturbing metabolic pathways in the initial 24 h, resulting in changes to factors, such as metabolites or signaling molecules, which subsequently triggered cell death relevant pathways in the next 24 h. The uptake and release of 27 metabolites were measured to further elucidate the metabolic changes in the first 24 h. It was determined that ketone bodies such as beta-hydroxybutyrate and acetoacetate were important in separating the toxic from the nontoxic phenotypes. A regression model was used to identify the genes relevant to these metabolites. Some of the genes identified to be important were experimentally validated. It was found that ETC genes such as NADH dehydrogenase and succinate dehydrogenase were involved in palmitate induced cytotoxicity.

SUBMITTER: Li Z

PROVIDER: S-EPMC4059368 | biostudies-literature | 2008 Jan-Feb

REPOSITORIES: biostudies-literature

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Using dynamic gene module map analysis to identify targets that modulate free fatty acid induced cytotoxicity.

Li Zheng Z Srivastava Shireesh S Findlan Robert R Chan Christina C

Biotechnology progress 20071204 1

The objective of this study was to identify pathways that regulate the cytotoxicity induced by free fatty acids (FFAs) in human hepatoblastoma cells (HepG2/C3A). Gene expression profiles of HepG2/C3A cells were obtained at three time points, after 24, 48, and 72 h of exposure to different types of FFA. Saturated fatty acid (palmitate) was found to be cytotoxic. The pathways activated by the different FFAs at the different time points were identified using global gene module map analysis. Unsatur ...[more]

PMID: 18052188

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Project description:Streptococcus pneumoniae, a major cause of pneumonia, sepsis, and meningitis worldwide, has the nasopharynges of small children as its main ecological niche. Depletion of pneumococci from this niche would reduce the disease burden and could be achieved using small molecules with narrow-spectrum antibacterial activity. We identified the alkylated dicyclohexyl carboxylic acid 2CCA-1 as a potent inducer of autolysin-mediated lysis of S. pneumoniae, while having low activity against Staphylococcus aureus 2CCA-1-resistant strains were found to have inactivating mutations in fakB3, known to be required for uptake of host polyunsaturated fatty acids, as well as through inactivation of the transcriptional regulator gene fabT, vital for endogenous, de novo fatty acid synthesis regulation. Structure activity relationship exploration revealed that, besides the central dicyclohexyl group, the fatty acid-like structural features of 2CCA-1 were essential for its activity. The lysis-inducing activity of 2CCA-1 was considerably more potent than that of free fatty acids and required growing bacteria, suggesting that 2CCA-1 needs to be metabolized to exert its antimicrobial activity. Total lipid analysis of 2CCA-1 treated bacteria identified unique masses that were modeled to 2CCA-1 containing lysophosphatidic and phosphatidic acid in wild-type but not in fakB3 mutant bacteria. This suggests that 2CCA-1 is metabolized as a fatty acid via FakB3 and utilized as a phospholipid building block, leading to accumulation of toxic phospholipid species. Analysis of FabT-mediated fakB3 expression elucidates how the pneumococcus could ensure membrane homeostasis and concurrent economic use of host-derived fatty acids.IMPORTANCE Fatty acid biosynthesis is an attractive antibiotic target, as it affects the supply of membrane phospholipid building blocks. In Streptococcus pneumoniae, it is not sufficient to target only the endogenous fatty acid synthesis machinery, as uptake of host fatty acids may bypass this inhibition. Here, we describe a small-molecule compound, 2CCA-1, with potent bactericidal activity that upon interactions with the fatty acid binding protein FakB3, which is present in a limited number of Gram-positive species, becomes metabolized and incorporated as a toxic phospholipid species. Resistance to 2CCA-1 developed specifically in fakB3 and the regulatory gene fabT These mutants reveal a regulatory connection between the extracellular polyunsaturated fatty acid metabolism and endogenous fatty acid synthesis in S. pneumoniae, which could ensure balance between efficient scavenging of host polyunsaturated fatty acids and membrane homeostasis. The data might be useful in the identification of narrow-spectrum treatment strategies to selectively target members of the Lactobacillales such as S. pneumoniae.

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Using dynamic gene module map analysis to identify targets that modulate free fatty acid induced cytotoxicity.

Publications

Using dynamic gene module map analysis to identify targets that modulate free fatty acid induced cytotoxicity.

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