(801.17) Contribution of stress granules and spliceosomal components to the formation of chemoresistance in ovarian cancer cells

Poster Board Number: A219

Polina Shnaider (Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical and Biological Agency), Irina Malyants (Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical and Biological Agency), Olga Ivanova (Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical and Biological Agency), Georgij Arapidi (Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical and Biological Agency, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences), Maria Lagarkova (Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical and Biological Agency), Vadim Govorun (Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical and Biological Agency), Victoria Shender (Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical and Biological Agency, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences)

The emergence of chemoresistance in cancer cells is one of the main reasons for the high mortality from oncological diseases. We have shown that signaling molecules from dying cancer cells contribute to a more aggressive therapy-resistant phenotype. However, the molecular mechanisms underlying this effect remain elusive. 

Here we aimed to find out which proteins secreted by dying tumor cells get into recipient cells and to determine the molecular cascades triggered in response to incubation with therapy-induced secretomes. We modified the SILAC (Stable Isotope Labeling by Amino acids in Cell culture) technology. A heavy-labeled primary culture of ovarian cancer cells was treated or untreated with cisplatin for 48 h. Then using ultrafiltration cartridges, a fraction of extracellular vesicles (EV) were isolated from culture media. The obtained EV were resuspended in a medium containing Medium-labeled amino acids and added to the recipient chemonaive unlabeled tumor cells for 24 h. During incubation, recipient cells absorbed EV with Heavy-labeled proteins from donor cells and synthesized new proteins with Medium label. Then, we performed proteomic analysis of recipient cells. In total, we identified 4224 proteins. Interestingly, dying tumor cells secrete and recipient cells absorb a large number of spliceosomal and ribosomal proteins along with translation initiation complex. These proteins are often presented in stress granules. In response to EV from dying tumor cells, recipient cells upregulated the translation of genes responsible for the regulation of the cell cycle and oxidative phosphorylation.

To our knowledge, this is the first report about the secretion of stress granules and spliceosomal components into the extracellular space and their internalization in other cells. To confirm these data we performed transfection of donor cells with constructs encoding the fluorescent proteins TIA1 (protein of stress granules) and SRSF4 (protein of the spliceosome). We showed that both proteins are re-located together from the nucleus into the cytoplasm under the cisplatin treatment. Then, we demostrated the absorption of fluorescent-labeled proteins TIA1 and SRSF4 by recipient cells after incubation with therapy-induced EV. Our findings demonstrate that in response to cisplatin treatment tumor cells secrete spliceosomal components into the extracellular space (presumably as a part of stress granules), then these components are absorbed by recipient chemonaive cells and trigger a restructuring of metabolism and cell cycle.

Support or Funding Information

The work was supported by the grant 075-15-2019-1669 from the Ministry of Science and Higher Education of the Russian Federation (for LC-MS-MS analysis) and the Russian Science Foundation project 19-75-10123 (for molecular cloning).