We examined survival, growth and protein synthesis in mosquito cells that had been maintained for up to 21 days in serum-free medium. On polyacrylamide gels, protein bands from "starved" cells remained discrete, and despite low levels of incorporation, radiolabeled bands were detectable, suggesting that low levels of protein synthesis were sustained. A prominent band that accumulated in serum-starved cells was digested with trypsin and analyzed by tandem mass spectrometry, which identified the protein as eukaryotic elongation factor (EF)-1α EF-1α is well-conserved among species, and differential accumulation of EF-1α in serum-starved cells was verified by western blotting using a primary antibody to the homologous protein from Trypanosoma brucei. Aside from its importance in the elongation step of protein synthesis, EF-1α has been shown to have a number of non-canonical functions, including interaction with viral RNA and a potential role in apoptosis. We anticipate that the prolonged viability of mosquito cells in serum-free medium may provide a system to explore whether EF-1α accumulation is an adaptive response compatible with resumption of growth in the event that nutrients are replenished, or whether the excess EF-1α represents an irreversible commitment to an apoptotic pathway.
Bibliographical noteFunding Information:
This work was supported by National Institutes of Health grants AI 20385 and AI 43971 and by the University of Minnesota Agricultural Experiment Station, St. Paul, MN. The authors recognize the Mass Spectrometry Consortium for the Life Sciences at the University of Minnesota and various supporting agencies, including the National Science Foundation for Major Research Instrumentation grant 9871237, used to purchase the instruments described in this study.
- Elongation factor-1α
- Mosquito cell line
- Protein synthesis