Markus R WENK
Cellular membranes share common classes of lipid
constituents, but differ in the relative proportions
of the individual molecular species. They segregate
cellular compartments and provide a structural
scaffold for the organization of enzymatic
machineries and for the anchoring of the cytoskeleton.
In addition, membrane lipids play an important
role in cellular signaling. This signaling
function is made possible by the very active
metabolism of some lipids leading to the transient
accumulation of compounds that play critical
roles in short and long range cellular regulation.
A well-known example of regulatory phospholipids
are the phosphoinositides (phosphorylated derivatives
of phosphatidylinositol) which contribute to
the regulation of nearly every cellular function
including membrane traffic, cell proliferation,
and cell motility. Similar signaling functions
are emerging for a number of other lipid classes,
including ceramides, lysolipids, and diacylglycerols.
The critical role of lipids in cell, tissue and organismal physiology is demonstrated by a large number of genetic studies and by the many human diseases involving the disruption of lipid metabolic enzymes and pathways. Examples of such diseases include cancer, diabetes and neurodegenerative diseases. Surprisingly, so far, the explosion of information in the fields of genomics and proteomics has not been matched by a corresponding advancement of knowledge in the field of lipids. Conventional methods of lipid analysis are hampered by low sensitivity and the difficulty to detect and measure lipids from unprocessed extracts. Furthermore, many available techniques of lipid analysis have not been suited to systematic studies. We plan to capitalize on the increased sensitivity and resolution power of mass spectrometry to advance the field and obtain biologically and medically important information.
The inositol head group of phosphatidylinositol can be reversibly phosphorylated at various positions resulting in seven naturally occurring phosphoinositides. Shown here is the molecular structure of 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphatidylinositol-4,5-bisphosphate, an abundant PI(4,5)P2 species in mammalian cells. Commonly used methods for the detection of phosphoinositides include fluorescent techniques, receptor displacement assays and radiometric measurements of chromatographically separated products after metabolic labeling with radioisotope precursors. As a result, some of these approaches report the position of the phosphate(s) on the inositol headgroup (isomer information). Electrospray ionization mass spectrometry, on the other hand, reveals a number of PIP2 species with different fatty acid compositions, depending on the number of carbon atoms and double bonds in their fatty acid moieties. This fatty acid information adds an additional layer of information to phosphoinositide signaling. The dotted line indicates the plane of the bilayer and roughly marks the aqueous-to-hydrophobic boundary of the membrane.
'Lipidomics of neuronal membranes - Identification of lipids involved in neurosecretion and neurodegenerative diseases'
The goal of this project is to identify membrane lipids which undergo stimulation induced metabolism during neurosecretion. This work will be done in collaboration with research groups at the National University of Singapore, the University of Western Australia, Perth and Yale University in New Haven, CT, USA.
profiling of Mycobacteria in different physiological
The aim of this project is to test whether tubercle bacilli change their membrane lipid composition upon entry into metabolically inert states which are resistant to conventional anti-tuberculosis treatment. Research collaboration with the Novartis Institute for Tropical Diseases, Singapore.
of enveloped viruses - novel molecular insights
into host-pathogen interactions'
Mass spectrometry will be used to determine the lipid inventory of enveloped viruses which will be the first systematic high resolution compositional analysis of viral envelope lipids. In collaboration with groups at the Institute for Molecular and Cell Biology, Singapore and Yale University, New Haven, CT, USA.
MR, Lucast L, Di Paolo G, Romanelli AJ, Suchy
SF, Nussbaum RL, Cline GW, Shulman GI, McMurray
W, and P De Camilli. 2003. Nat. Biotechnol. 7:813-817.
Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry.
MR, and P De Camilli. 2003. Methods
Assembly of endocytosis-associated proteins on liposomes.
Paolo G, Pellegrini L, Letinic K, Cestra
G, Zoncu R, Voronov S, Chang S, Guo J, Wenk
MR, and P De Camilli. 2002. Nature. 402:85-89.
PIP2 generation at sites of focal adhesion: Critical role for the interaction of PIP Kinase I-gamma with the FERM domain of talin.
E, Marcucci M, Daniell L, Pypaert M, Weisz
O, Ochoa GC, Farsad K, Wenk MR, and P De
Camilli. 2002. Science.
Amphiphysin 2 (Bin1) and T-tubule biogenesis in muscle.
MR, Pellegrini L, Klenchin VA, Di Paolo G,
Chang S, Daniell L, Arioka M, Martin TF,
and P De Camilli. 2001. Neuron 32:79-88.
PIP kinase I-gamma is the major PI(4,5)P2 synthesizing enzyme at the synapse.
O, Di Paolo G, Wenk MR, Lthi
A, Kim WT, Takei K, Daniell L, Nemoto Y,
Shears SB, Flavell R, McCormick DA, and P
De Camilli. 1999. Cell 99:179-188.
Essential role of phosphoinositide metabolism in synaptic vesicle recycling.
MR, and J Seelig. 1998. Biochim.
Biophys. Acta 1372:2227-2236.
Proton induced vesicle fusion and the isothermal Lalpha<-HII phase transition of lipid bilayers. A 31P-NMR and titration calorimetry study.
MR, and J Seelig. 1998. Biochemistry 37:3909-3916.
Magainin 2 amide interaction with lipid membranes: calorimetric detection of peptide binding and pore formation.
MR, and J Seelig. 1997. J. Phys.
Chem. B 101:5224-5231.
Vesicle-micelle transformation of phosphatidylcholine / octyl-beta-D-glucopyranoside mixtures as detected with titration calorimetry.