Synthesis and cellular characterization of the detransformation agent, (-)-depudecin.
BACKGROUND
(-)-Depudecin is a fungal metabolite that reverts the rounded phenotype of NIH3T3 fibroblasts transformed with v-ras and v-src oncogenes to the flat phenotype of the nontransformed parental cells. The mechanism of action of this detransformation agent is unknown. Although depudecin appears to be an excellent molecule for probing signaling pathways that regulate changes in the cytoskeletal architecture, reagents based on depudecin are not available as it has not yet been successfully synthesized. We therefore set out to synthesize (-)-depudecin.


RESULTS
An asymmetric synthesis of (-)-depudecin has been developed. A cell staining assay has been used to reveal the ability of synthetic depudecin, but not several structural variants, to induce a flattened morphology in v-Ha-ras-transformed NIH3T3 cells. This assay also shows that depudecin induces an intricate network of actin stress fibers in these cells and in MG63 osteosarcoma cells and reveals the essential role of the epoxide and hydroxyl moieties in depudecin. Cycloheximide and actinomycin D inhibited the ability of depudecin to induce a morphological change, suggesting that both mRNA synthesis and de novo protein synthesis are required for depudecin-mediated suppression of the transformed phenotypes in ras-transformed cells.


CONCLUSIONS
The synthetic procedure provides access to (-)-depudecin and could be readily modified to produce depudecin-related reagents for the identification of depudecin's cellular target(s). This target appears to be involved in the regulation of the assembly of the actin microfilament component of the cytoskeleton in mammalian cells.
Shimada
J
J
Howard Hughes Medical Institute, Department of Chemistry, Harvard University, Cambridge, MA 02138, USA.
Kwon
H J
HJ
Sawamura
M
M
Schreiber
S L
SL
D016475:3T3 Cells / Q000187:drug effects; D000199:Actins / Q000737:chemistry / Q000187:drug effects; D000466:Alkadienes / Q000138:chemical synthesis* / Q000494:pharmacology*; D000818:Animals; D001859:Bone Neoplasms / Q000737:chemistry; D002467:Cell Nucleus / Q000187:drug effects / Q000648:ultrastructure; D002471:Cell Transformation, Neoplastic / Q000187:drug effects*; D003513:Cycloheximide / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects; D003609:Dactinomycin / Q000494:pharmacology; D004852:Epoxy Compounds / Q000138:chemical synthesis* / Q000494:pharmacology*; D005233:Fatty Alcohols / Q000138:chemical synthesis* / Q000494:pharmacology*; D011905:Genes, ras / Q000187:drug effects; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D012516:Osteosarcoma / Q000737:chemistry; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D013055:Spectrophotometry, Infrared; D013237:Stereoisomerism; D014407:Tumor Cells, Cultured
D000199:Actins; D000466:Alkadienes; D004852:Epoxy Compounds; D005233:Fatty Alcohols; D011500:Protein Synthesis Inhibitors; C072251:depudecin; D003609:Dactinomycin; D003513:Cycloheximide