| The Streptomyces genus of actinobacteria has yielded a cornucopia of bio-active molecules, many of which have found application in the clinic. Recently, a group of Streptomyces polyketides garnered interest as cell migration inhibitors, centered on the macrolide migrastatin. We investigated the biological activity of 35 molecules related to migrastatin, including its parent compound isomigrastatin and its open-chain form dorrigocin. Despite their origin from the same biosynthetic machinery and structural homology, their mechanisms of action differ greatly. Though isomigrastatin and the closely related molecule lactimidomycin (LTM) had also been described as cell migration inhibitors, they proved to inhibit protein synthesis instead. LTM acted in a similar, though not identical fashion to the cycloheximide (CHX) in blocking translation elongation. Even though CHX is the most commonly used laboratory reagent for inhibiting protein synthesis, its mechanism of action remains incompletely understood. LTM and CHX share the glutarimide structural motif though CHX has only about half the molecular weight of the other molecule. Both inhibited eEF2 mediated translocation, thereby halting translation early in the elongation phase. However, CHX allowed one complete elongation cycle to proceed while LTM halted translation immediately.;CHX and LTM shared the same binding site located in the exit (E) site of the large ribosomal subunit. The larger LTM occluded deacylated tRNA from the E site, which prevented the first round of translocation to proceed. In contrast CHX bound together with the E-site tRNA after the first translocation step had occurred. Thus the dissection of LTM's mechanism finally explained the workings of CHX as well. Either mechanism differs from all previously known translation inhibitors, both eukaryotic and bacterial, in inhibiting from the E site rather than acting on the ribosome catalytic center, decoding or translation factor binding. The initially described inhibition of cell migration most likely resulted from the inhibition of translation and did not constitute an independent effect.;Since growth and development are translation dependent processes, which also drive tumorigenesis LTM was tested against an array of breast cancer cell lines. LTM proved highly cytotoxic and prevented cell proliferation at low nanomolar doses. It displayed selectivity for transformed cell lines and slowed tumor growth in mouse xenograft models. While not suited as a primary therapeutic agent, LTM may prove useful in conjunction with an established first-line drug.;With the characterization of its mechanism and structural requirements for activity LTM presents a specific and potent probe into eukaryotic translation with potential in both research and clinical applications. |
