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Cegelski et al. (2009) demonstrate the importance of amyloid production for biofilm formation and host colonization using several mutant strains of pathogenic E. coli and small molecule inhibitors. This work reveals a path forward for studying the role of bacterial amyloids in vivo and suggests the potential for small molecules to target multiple biofilm formation pathways., (Copyright 2009 Elsevier Ltd. All rights reserved.)

Small molecule natural products have aided in the discovery and characterization of many proteins critical to the progression and maintenance of the cell cycle. Identification of the direct target of a natural product gives scientists a tool to control a specific aspect of the cell cycle, thus facilitating the study of the cell-cycle machinery.

Background: The marine natural product (+)-discodermolide has potent immunosuppressive activity. It inhibits proliferation of a wide range of human and murine cells, induces cell cycle arrest in the G2 or M phase and was recently shown to stabilize microtubules. Total synthesis of discodermolide has made it possible to generate variants of the compound to study its intracellular function in detail., Results: We have determined that (+)-discodermolide arrests MG63 cells at M phase, and has a stabilizing effect on microtubules. In vitro studies show that discodermolide induces polymerization of purified tubulin in the absence of microtubule-associated proteins, and that it binds to tubulin dimers in microtubules at 1:1 stoichiometry. Discodermolide binds taxol-polymerized microtubules at near stoichiometric level, whereas taxol binds discodermolide-induced microtubules poorly. Competition data show that the binding of microtubules by discodermolide and taxol are mutually exclusive; discodermolide binds with higher affinity than taxol. The results of binding assays carried out in vivo or in cell lysates also suggest that the microtubule network is discodermolide's cellular target., Conclusions: (+)-Discodermolide causes cell cycle arrest at the metaphase-anaphase transition in mitosis, presumably due to its stabilizing effect on microtubules. In vitro, discodermolide polymerizes purified tubulin potently in the absence of MAPs. It binds microtubules at one molecule per tubulin dimer with a higher affinity than taxol, and the binding of microtubules by discodermolide and taxol are mutually exclusive. In total cell lysates discodermolide displays binding activity that is consistent with its effects on microtubules.

Background: Cell permeable ligands of low molecular weight can be used to dissect complex cellular processes. During the past several years this approach has been particularly important in the study of intracellular signal transduction. Discodermolide, a marine natural product, appears to inhibit a signaling pathway in immune cells. The structure of natural discodermolide is known, but its absolute stereochemistry is not. We set out to make both enantiomers and to investigate their biological activity., Results: Both enantiomers of discodermolide were prepared by total synthesis. Surprisingly, both enantiomers have biological activity, and their effects seem to be distinct in that they arrest cells at different stages of the cell cycle. A specific binding activity was identified for (+)-discodermolide but not for (-)-discodermolide, and the binding of the two enantiomers was not competitive., Conclusions: Both enantiomers of discodermolide have antiproliferative activity, but they act by distinct mechanisms and appear to have distinct cellular targets. The natural product is the (+)-enantiomer, which blocks the cell cycle in the G2 or M phase. The (-)-enantiomer blocks cells in S phase. Both may be useful in studies of the regulation of the cell cycle; we have also identified a specific binding activity for (+)-discodermolide, and have provided evidence that it interacts with a functionally relevant receptor.