Lipidomics meeting: Fat mapping in the sun
Lipidomics Gateway (27 May 2010) [doi:10.1038/lipidmaps.2010.16]
Idyllically set, the LIPID MAPS annual conference focused on locations and functions of lipids in disease.
Lipidomics has matured; powerful, reliable methods are delivering unprecedented detail about the functions of lipids. The title of the second annual public LIPID MAPS meeting, Lipidomics Impact on Cell Biology, Atherosclerosis and Inflammatory Disease, reflects knowledge gained but also an intention: as key roles of lipids in disease emerge, lipid scientists must engage the wider community and share their expertise.
Perceptions old and new
Lipid biologists frequently bemoan the difficulty of persuading other researchers that lipids are 'more than membranes'. Nevertheless, the first topic eased any lipid newcomers in on familiar ground, as William Dowhan described how bilayer lipids interact with membrane proteins. A subset of the latter, including lactose permease (LacY) from Escherichia coli, requires sufficient membrane phosphatidylethanolamine (PE) for correct topology and function. Using E. coli mutants lacking PE, and introducing genes for the synthesis of foreign lipids, Dowhan and colleagues found that the influence of charged amino acids on protein topology is modified by the cumulative effect of local lipid headgroups, such that protein topology responds to the lipid environment 1 . Furthermore, both the lipid headgroup and the fatty acid composition of phospholipids influence protein function.
Roger Tsien is familiar with the staying power of perceptions. Irrevocably known for his work on green fluorescent protein (GFP) – for which he shared the Nobel Prize in Chemistry – he cannot use green in his cell imaging figures without the assumption that it denotes GFP. He commiserated with the audience over the tendency to presume that useful molecules are peptides, a fate that occasionally befalls one of his latest tools, dendrimers. These large, spherical molecules can contain contrast agents or fluorescent groups used for imaging. Tsien and colleagues developed activatable cell-penetrating peptides (ACPPs) and conjugated them to dendrimers to enhance their uptake by cells 2 3 . ACPPs consist of a polycation peptide linked to and masked by a polyanion sequence. Specific cleavage of the linker unmasks the polycation, which adheres to and is taken up by cells alongside conjugated molecules. For tumor imaging, used to enhance surgical resection, the linker is cleaved by matrix metalloproteinases expressed by tumor cells 3 . Tsien reported that using a linker with a thrombin target sequence instead directs the ACPPs to atherosclerotic plaques, especially vulnerable ones, enabling visualization. These ACPPs could guide bypass surgery, guard against risky plaque nudging during other invasive procedures, or target drugs to the plaques.
After fatty plaques, Robert Murphy focused the imaging section of the talks onto individual lipid species. Matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) allows images to be constructed from sequential mass spectra of ions blasted from a target tissue surface, and is a powerful way to show the abundance of specific lipid species across a sample 4 . Three of the key insights that have been gained so far with this technique are: arachidonic acid-containing species of phosphatidylcholine (PC) are concentrated around airways; in a rat model of stroke, various ceramide molecular species accumulate in specific hippocampal regions; and although there is no change in PC content overall at sites of traumatic injury, more sodiated PC adduct ions emanate from the region of injury. Validation and optimization continues, and the technique only seems to produce false negatives for a few molecular species. In a short talk, Hay-Yan Wang described a new tissue-processing method that enhances MALDI-MS imaging of pathological samples that contain an interfering overabundance of sodium.
Updating the Lipidomics update
A few speakers described work that we have highlighted since our inception a year ago. Alex Brown updated Lipid analysis: Click, stick... and release with news that alkynyl lipids are converted to eicosanoids, further validating their use for metabolic studies. During a shared sphingolipid talk, Al Merrill and Marion Sewer interweaved developments to two threads: nuclear sphingolipid signaling (see Lipid signaling: S1P's nuclear program) and the novel deoxy sphinganine bases (deoxySa) introduced in Fungal toxicity: Based in sphinganine. DeoxySa's are produced by the use of alanine or glycine in place of serine by serine palmitoyltransferase (SPT). They are found in normal tissues, but can be elevated by mutations in SPT to cause human hereditary sensory neuropathy type 1 5 . Whether deoxySa's are merely an 'accident of nature' is unresolved said Merrill, but the centrality of serine, glycine and alanine in intermediary metabolism suggests that these sphingoid bases could serve as regulatory mediators. Indeed, Sewer reported that the most abundant sphingoid base sphingosine is, unusually, an antagonist for a nuclear receptor. Essential for sex differentiation, steroidogenic factor-1 (SF-1) regulates the expression of genes involved in steroid metabolism and endocrine development. Besides sphingosine, SF-1 also binds to phosphatidic acid, which conversely increases transcription. These findings develop the nascent concept of transcriptional regulation by products of both sphingolipid and glycerolipid metabolism.
Inflammatory and immune responses are rich in lipid pathways. Stimulating macrophages with KDO2-Lipid A, and in the future, oxidized low density lipoprotein (oxLDL) components, and monitoring subsequent lipid profiles is integral to the LIPID MAPS approach. As well as our highlight this month Inflammatory pain: Calm resolve, related articles include Inflammation: Finding the T in fat, Fat finding in disease: Lipids coming 'ome, Macrophage lipid droplets: Inert? Eicosa' not! and Cancer and Prostaglandin E2: Don't make it, or break it?.
Central to this theme is cyclooxygenase 2 (COX2), the rate limiting enzyme in prostaglandin and related lipid synthesis, and a target of non-steroidal anti-inflammatory drugs (NSAIDs). Harvey Herschman was one of the discoverers of the gene for COX2, and he spoke about its complex roles in colitis and colon cancer. Paradoxically, NSAIDS exacerbate colon inflammation in mouse models of inflammatory bowel disease (IBD). Macrophage-specific knockouts of COX2 in these mice increases clinical colitis symptoms and causes greater apoptosis of epithelial cells, indicating a protective role for the enzyme. IBD can lead to colon cancer, and knocking out COX2 expression in mice prevents colon tumorigenesis caused by a mutagenic agent, but not that caused by mimicking inflammation. Compensatory pathways involving inflammatory cytokines take over in the absence of COX2.
We highlighted work on the Lipidomics gateway showing that activation of Toll-like receptor 4 (TLR4) stimulates the production of 25-hydroxycholesterol in macrophages, and this suppresses production of immunoglobulin A (Immune response: Oxysterol takes Toll on IgA). Although this is the main oxysterol produced in macrophages in response to TLR4 signaling, Chris Glass told us that in foam cells, 24,25-epoxycholesterol predominates. The two oxysterols induce different patterns of macrophage gene expression: genes involved in the ER stress response for 25-hydroxycholesterol, and in cholesterol homeostasis for 24,25-epoxycholesterol. Now the focus is on how these programs of expression influence two potential outcomes of macrophage activation: resolution of inflammation, or cell death and tissue damage.
The mechanisms and consequences of macrophage death in atherosclerotic lesions are Ira Tabas's domain. Whereas an apoptotic response to stress can be appropriate, stressed macrophages are needed to fight infection. Tabas and colleagues found that TLR signaling in macrophages modifies the unfolded protein response, with a key role for the phosphatase PP2A in the signaling pathway.
Consort and collaborate
The meeting overall was a balance of LIPID MAPS investigators, collaborators and newcomers. Catherine Costello presented her work as an independent investigator and Eoin Fahy represented the home team; Costello spoke about the suitability of different MS methodologies for particular lipid analyses, whereas Fahy outlined the bioinformatic tools that the consortium provides. Other speakers discussed how lipidomics data could be integrated into external metabolomics, inflammation and chemical databases, to ensure that the roles of lipids are considered by researchers primarily interested in a disease or pathway. Still more, including Philip Beachy speaking about the role of lipids in Hedgehog signaling, exemplified how lipidomics is penetrating areas of research that have typically been focused on genetic or proteomic data. Increasing numbers of LIPID MAPS publications are now the result of collaborations, and eager seaside discussions over lunch suggest that several new ones have just been conceived.
There were many excellent talks that we do not have space to elaborate on here, or about which we expect imminent publications that may be featured in future updates. For those present, the meeting was an unqualified success. “People left this meeting with a great appreciation of the role of lipids in disease and advances that lipidomics is contributing to that understanding,” said Ed Dennis, director of LIPID MAPS.
It may be happening one collaboration at a time, but as the lipidomics field turns outwards, its impact is certain to be felt.
Emma Leah- Copyright © 2010 Nature Publishing Group, a division of Macmillan Publishers Limited; used with permission
Dowhan, W. and Bogdanov, M. Lipid-dependent membrane protein topogenesis.
Annu. Rev. Biochem. 78, 515-40 (2009). doi:10.1146/annurev.biochem.77.060806.091251
Olson, E. S. et al. Activatable cell penetrating peptides linked to nanoparticles as dual probes for in vivo fluorescence and MR imaging of proteases.
Proc. Natl Acad. Sci. USA 107, 4311-4316 (2010). doi:10.1073/pnas.0910283107
Nguyen, Q. T. et al. Surgery with molecular fluorescence imaging using activatable cell-penetrating peptides decreases residual cancer and improves survival.
Proc. Natl Acad. Sci. USA 107, 4317-4322 (2010). doi:10.1073/pnas.0910261107
Murphy, R. C., Hankin, J. A. and Barkley, R. M. Imaging of lipid species by MALDI mass spectrometry.
J. Lipid Res. 50, S317-S322 (2009). doi:10.1194/jlr.R800051-JLR200
Penno, A. et al. Hereditary Sensory Neuropathy Type 1 is caused by the accumulation of two neurotoxic sphingolipids.
J. Biol. Chem. 285, 11178-11187 (2010). doi:10.1074/jbc.M109.092973