Thomas Kaufmann's research while affiliated with Hannover Medical School and other places

In search of a more comprehensive structure activity relationship (SAR) regarding the inhibitory effect of cytochalasin B (CB) on actin polymerization, a virtual docking of CB onto monomeric actin was conducted. This led to the identification of potentially important functional groups of CB (i.e. the NH group of the isoindolone core (N-2), and the hydroxyl groups at C-7 and C-20) involved in interactions with the residual amino acids of the binding pocket of actin. Chemical modifications of CB at positions C-7, N-2, and C-20 led to derivatives CB1-CB4, which were analyzed for their bioactivities. CB1-CB4 exhibited reduced or no cytotoxicity in murine L929 fibroblasts compared to CB. Moreover, short- and long-term treatments of human osteosarcoma cells (U-2OS) affected the actin network to variable extent, partially accompanied by the induction of multinucleation. Derivatives displaying acetylation at C-20 and N-2 were subject to slow intracellular conversion to highly cytotoxic CB. Together, this study highlights the importance of the hydroxy group at C-7 and the NH function at N-2 for CB potency on the inhibition of actin polymerization.

In search of a more comprehensive structure activity relationship (SAR) regarding the inhibitory effect of cytochalasin B (CB) on actin polymerization, a virtual docking of CB onto monomeric actin was conducted. This led to the identification of potentially important functional groups of CB (i.e. the NH group of the isoindolone core (N-2), and the hydroxyl groups at C-7 and C-20) involved in interactions with the residual amino acids of the binding pocket of actin. Chemical modifications of CB at positions C-7, N-2, and C-20 led to derivatives CB1-CB4, which were analyzed for their bioactivities. CB1-CB4 exhibited reduced or no cytotoxicity in murine L929 fibroblasts compared to CB. Moreover, short-and long-term treatments of human osteosarcoma cells (U-2OS) affected the actin network to variable extent, partially accompanied by the induction of multinucleation. Derivatives displaying acetylation at C-20 and N-2 were subject to slow intracellular conversion to highly cytotoxic CB. Together, this study highlights the importance of the hydroxy group at C-7 and the NH function at N-2 for CB potency on the inhibition of actin polymerization.

Background: Filopodia are dynamic, finger-like actin-filament bundles that overcome membrane tension by forces generated through actin polymerization at their tips to allow extension of these structures a few microns beyond the cell periphery. Actin assembly of these protrusions is regulated by accessory proteins including heterodimeric capping protein (CP) or Ena/VASP actin polymerases to either terminate or promote filament growth. Accordingly, the depletion of CP in B16-F1 melanoma cells was previously shown to cause an explosive formation of filopodia. In Ena/VASP-deficient cells, CP depletion appeared to result in ruffling instead of inducing filopodia, implying that Ena/VASP proteins are absolutely essential for filopodia formation. However, this hypothesis was not yet experimentally confirmed. Methods: Here, we used B16-F1 cells and CRISPR/Cas9 technology to eliminate CP either alone or in combination with Ena/VASP or other factors residing at filopodia tips, followed by quantifications of filopodia length and number. Results: Unexpectedly, we find massive formations of filopodia even in the absence of CP and Ena/VASP proteins. Notably, combined inactivation of Ena/VASP, unconventional myosin-X and the formin FMNL3 was required to markedly impair filopodia formation in CP-deficient cells. Conclusions: Taken together, our results reveal that, besides Ena/VASP proteins, numerous other factors contribute to filopodia formation.