1. Automated High-Throughput Fluorescence Lifetime Imaging Microscopy to Detect Protein-Protein Interactions.

    Journal of Laboratory Automation 21(2):238 (2016) PMID 26384400

    Fluorescence resonance energy transfer (FRET) is widely used to study conformational changes of macromolecules and protein-protein, protein-nucleic acid, and protein-small molecule interactions. FRET biosensors can serve as valuable secondary assays in drug discovery and for target validation in...
  2. Galectin-1 dimers can scaffold Raf-effectors to increase H-ras nanoclustering.

    Scientific reports 6:24165 (2016) PMID 27087647 PMCID PMC4834570

    Galectin-1 (Gal-1) dimers crosslink carbohydrates on cell surface receptors. Carbohydrate-derived inhibitors have been developed for cancer treatment. Intracellularly, Gal-1 was suggested to interact with the farnesylated C-terminus of Ras thus specifically stabilizing GTP-H-ras nanoscale signal...
  3. ASPP2 Is a Novel Pan-Ras Nanocluster Scaffold.

    PLoS ONE 11(7):e0159677 (2016) PMID 27437940 PMCID PMC4954646

    Ras-induced senescence mediated through ASPP2 represents a barrier to tumour formation. It is initiated by ASPP2's interaction with Ras at the plasma membrane, which stimulates the Raf/MEK/ERK signaling cascade. Ras to Raf signalling requires Ras to be organized in nanoscale signalling complexes...
  4. Phenotypic Screening Identifies Protein Synthesis Inhibitors as H-Ras-Nanocluster-Increasing Tumor Growth Inducers.

    Biochemistry (Washington) 54(49):7212 (2015) PMID 26568031

    Ras isoforms H-, N-, and K-ras are each mutated in specific cancer types at varying frequencies and have different activities in cell fate control. On the plasma membrane, Ras proteins are laterally segregated into isoform-specific nanoscale signaling hubs, termed nanoclusters. As Ras nanocluste...
  5. GTP-specific fab fragment-based GTPase activity assay.

    Analytical chemistry 87(6):3527 (2015) PMID 25707436

    GTPases are central cellular signaling proteins, which cycle between a GDP-bound inactive and a GTP-bound active conformation in a controlled manner. Ras GTPases are frequently mutated in cancer and so far only few experimental inhibitors exist. The most common methods for monitoring GTP hydroly...
  6. Specific cancer-associated mutations in the switch III region of Ras increase tumorigenicity by nanocluster augmentation.

    eLife 4:e08905 (2015) PMID 26274561 PMCID PMC4563131

    Hotspot mutations of Ras drive cell transformation and tumorigenesis. Less frequent mutations in Ras are poorly characterized for their oncogenic potential. Yet insight into their mechanism of action may point to novel opportunities to target Ras. Here, we show that several cancer-associated mut...
  7. Rab-NANOPS: FRET biosensors for Rab membrane nanoclustering and prenylation detection in mammalian cells.

    Methods in Molecular Biology 1298:29 (2015) PMID 25800830

    Rab proteins constitute the largest subfamily of Ras-like small GTPases. They are central to vesicular transport and organelle definition in eukaryotic cells. Unlike their Ras counterparts, they are not a hallmark of cancer. However, a number of diseases, including cancer, show a misregulation o...
  8. S-7 : A new cytokine receptor activation paradigm: Activation of JAK2 by the growth hormone receptor

    Cytokine 70(1):22 (2014)

    The conformational changes required to transmit the GH binding signal from the extracellular domain of the GH receptor to its intracellular domain resulting in activation of JAK2 has been enigmatic. We have recently defined the first complete mechanistic model for JAK2 activation based...
  9. Synthesis and characterization of novel phosphonocarboxylate inhibitors of RGGT.

    European Journal of Medicinal Chemistry 84:77 (2014) PMID 25016230

    Phosphonocarboxylate (PC) analogs of the anti-osteoporotic drugs, bisphosphonates, represent the first class of selective inhibitors of Rab geranylgeranyl transferase (RabGGTase, RGGT), an enzyme implicated in several diseases including ovarian, breast and skin cancer. Here we present the synthe...
  10. A homogeneous quenching resonance energy transfer assay for H-Ras activation cycle monitoring and inhibitor screening

    New Biotechnology 31:S37 (2014)

  11. A homogeneous quenching resonance energy transfer assay for the kinetic analysis of the GTPase nucleotide exchange reaction.

    Analytical and Bioanalytical Chemistry 406(17):4147 (2014) PMID 24760397

    A quenching resonance energy transfer (QRET) assay for small GTPase nucleotide exchange kinetic monitoring is demonstrated using nanomolar protein concentrations. Small GTPases are central signaling proteins in all eukaryotic cells acting as a "molecular switches" that are active in the GTP-stat...
  12. The efficacy of Raf kinase recruitment to the GTPase H-ras depends on H-ras membrane conformer-specific nanoclustering.

    Journal of Biological Chemistry 289(14):9519 (2014) PMID 24569991 PMCID PMC3975003

    Solution structures and biochemical data have provided a wealth of mechanistic insight into Ras GTPases. However, information on how much the membrane organization of these lipid-modified proteins impacts on their signaling is still scarce. Ras proteins are organized into membrane nanoclusters, ...
  13. ColonyArea: an ImageJ plugin to automatically quantify colony formation in clonogenic assays.

    PLoS ONE 9(3):e92444 (2014) PMID 24647355 PMCID PMC3960247

    The clonogenic or colony formation assay is a widely used method to study the number and size of cancer cell colonies that remain after irradiation or cytotoxic agent administration and serves as a measure for the anti-proliferative effect of these treatments. Alternatively, this assay is used t...
  14. Nanoclustering and heterogeneous membrane diffusion of Ras studied by FRAP and RICS analysis.

    Methods in Molecular Biology 1120:307 (2014) PMID 24470034

    Fluorescence Recovery After Photobleaching (FRAP) and Raster Image Correlation Spectroscopy (RICS) are two powerful techniques to study the diffusion dynamics of fluorescently labeled proteins. FRAP and RICS can be easily applied on any commercial confocal microscope. In this chapter, we describ...
  15. H-Ras Membrane Orientation Affects Galectin-1 Dependent Nanoclustering

    Biophysical Journal 104(2):119a (2013)

  16. Lipid-Anchored Ras is Sorted by Membrane Curvature Both In Vitro and in Living Cells

    Biophysical Journal 104(2):96a (2013)

  17. Ski-interacting protein (SKIP) interacts with androgen receptor in the nucleus and modulates androgen-dependent transcription.

    BMC Biochemistry 14:10 (2013) PMID 23566155 PMCID PMC3668167

    The androgen receptor (AR) is a member of the nuclear receptor (NR) superfamily of ligand-inducible DNA transcription factors, and is the major mediator of male sexual development, prostate growth and the pathogenesis of prostate cancer. Cell and gene specific regulation by the AR is determined ...
  18. Cellular FRET-Biosensors to Detect Membrane Targeting Inhibitors of N-Myristoylated Proteins.

    PLoS ONE 8(6):e66425 (2013) PMID 23824448 PMCID PMC3688908

    Hundreds of eukaryotic signaling proteins require myristoylation to functionally associate with intracellular membranes. N-myristoyl transferases (NMT) responsible for this modification are established drug targets in cancer and infectious diseases. Here we describe NANOMS (NANOclustering and My...
  19. The effects of transmembrane sequence and dimerization on cleavage of the p75 neurotrophin receptor by γ-secretase.

    Journal of Biological Chemistry 287(52):43810 (2012) PMID 23105112 PMCID PMC3527965

    Cleavage of transmembrane receptors by γ-secretase is the final step in the process of regulated intramembrane proteolysis (RIP) and has a significant impact on receptor function. Although relatively little is known about the molecular mechanism of γ-secretase enzymatic activity, it is becoming ...
  20. Quantitative analysis of prenylated RhoA interaction with its chaperone, RhoGDI.

    Journal of Biological Chemistry 287(32):26549 (2012) PMID 22628549 PMCID PMC3410996

    Small GTPases of the Rho family regulate cytoskeleton remodeling, cell polarity, and transcription, as well as the cell cycle, in eukaryotic cells. Membrane delivery and recycling of the Rho GTPases is mediated by Rho GDP dissociation inhibitor (RhoGDI), which forms a stable complex with prenyla...