Summary s-adenosylmethionine (SAM) is the sole methyl donor modifying histones, nucleic acids and phospholipids. Its fluctuation impacts hepatic phosphatidylcholine (PC) synthesis or may be linked to variations in DNA or histone methylation. Physiologically, low SAM is associated with lipid accumulation, tissue injury and immune responses in fatty liver disease. However, molecular connections between SAM limitation, methyltransferases and disease-associated phenotypes are unclear. We find that low SAM can activate or attenuate Caenorhabditis elegans immune responses. Immune pathways are stimulated downstream of PC production on a non-pathogenic diet. In contrast, distinct SAM-dependent mechanisms limit survival on pathogenic Pseudomonas aeruginosa. C. elegans undertakes a broad transcriptional response to pathogens and we find that low SAM restricts H3K4me3 at Pseudomonas-responsive promoters, limiting their expression. Furthermore, this response depends on the H3K4 methyltransferase set-16/MLL. Thus, our studies provide molecular links between SAM and innate immune functions and suggest that SAM depletion may limit stress-induced gene expression.

Summary Lipogenesis requires coordinated expression of genes for fatty acid, phospholipid, and triglyceride synthesis. Transcription factors, such as SREBP-1 (Sterol regulatory element binding protein), may be activated in response to feedback mechanisms linking gene activation to levels of metabolites in the pathways. SREBPs can be regulated in response to membrane cholesterol and we also found that low levels of phosphatidylcholine (a methylated phospholipid) led to SBP-1/SREBP-1 maturation in C. elegans or mammalian models. To identify additional regulatory components, we performed a targeted RNAi screen in C. elegans, finding that both lpin-1/Lipin 1 (converts phosphatidic acid to diacylglycerol) and arf-1.2/ARF1 (a GTPase regulating Golgi function) were important for low-PC activation of SBP-1/SREBP-1. Mechanistically linking the major hits of our screen, we find that limiting PC synthesis or LPIN1 knockdown in mammalian cells reduces levels of active GTP-bound ARF1. Thus, changes in distinct lipid ratios may converge on ARF1 to increase SBP-1/SREBP-1 activity.

Mycobacterium tuberculosis induces metabolic reprogramming in macrophages like the Warburg effect. This enhances antimicrobial performance at the expense of increased inflammation, which may promote a pathogen-permissive host environment. Since the NAD+-dependent protein deacetylase Sirtuin 3 (SIRT3) is an important regulator of mitochondrial metabolism and cellular redox homeostasis, we hypothesized that SIRT3 modulation mediates M. tuberculosis-induced metabolic reprogramming. Infection of immortalized and primary murine macrophages resulted in reduced levels of SIRT3 mRNA and protein and perturbation of SIRT3-regulated enzymes in the tricarboxylic acid cycle, electron transport chain, and glycolytic pathway. These changes were associated with increased reactive oxygen species and reduced antioxidant scavenging, thereby triggering mitochondrial stress and macrophage cell death. Relevance to tuberculosis disease in vivo was indicated by greater bacterial burden and immune pathology in M. tuberculosis-infected Sirt3−/− mice. CD11b+ lung leukocytes isolated from infected Sirt3−/− mice showed decreased levels of enzymes involved in central mitochondrial metabolic pathways, along with increased reactive oxygen species. Bacterial burden was also greater in lungs of LysMcreSirt3L2/L2 mice, demonstrating the importance of macrophage-specific SIRT3 after infection. These results support the model of SIRT3 as a major upstream regulatory factor, leading to metabolic reprogramming in macrophages by M. tuberculosis. IMPORTANCE Tuberculosis, the disease caused by the bacterium M. tuberculosis, remains one of the top 10 causes of death worldwide. Macrophages, the first cells to encounter M. tuberculosis and critical for defense against infection, are hijacked by M. tuberculosis as a protected growth niche. M. tuberculosis-infected macrophages undergo metabolic reprogramming where key mitochondrial pathways are modulated, but the mechanisms driving this metabolic shift is unknown. Our study demonstrates that M. tuberculosis downregulates Sirtuin 3 (SIRT3), an important regulator of mitochondrial metabolism, leading to SIRT3-dependent transcriptional downregulation of mitochondrial metabolic proteins, which is followed by oxidative stress and macrophage necrosis. This study identifies SIRT3 modulation as a key event in M. tuberculosis-induced metabolic reprograming in macrophages that defend against tuberculosis.

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The sterol regulatory element‐binding protein (SREBP‐1a, ‐1c, and 2) family of transcription factors control genes involved in cholesterol, fatty acid, and triacylglycerol biosynthesis. Misregulation of these genes can contribute to development of the metabolic syndrome. Although cholesterol regulation of SREBP's has been well characterized, regulation by nutritional cues is less defined. Previously, we identified an additional level of SREBP‐1 regulation whereby proteolytic maturation is controlled by levels of intracellular phosphatidylcholine (PC). Importantly, we have found that ablation of PC biosynthesis results in increased protein expression of nuclear, transcriptionally active SREBP‐1 in multiple metabolically active cells, including human hepatoma (HepG2) and mouse macrophage cells. Inhibition of PC biosynthesis may alter the lipid composition of intracellular membrane compartments leading to SREBP‐1 activation, and further experiments will elucidate the mechanisms involved in low‐PC based activation of SREBP‐1. All together, nutritional or genetic factors which block PC biosynthesis could inappropriately activate SREBP‐1, leading to increased risk for development of diseases associated with the metabolic syndrome. This work is funded by the NIH (R01 DK084352).