Abstract
Background: The ribosomal protein S6 kinase 1 (S6K1) is one of the main components of the mTOR/S6K signal transduction pathway, which controls cellular metabolism, autophagy, growth, and proliferation. Overexpression of S6K1 was detected in tumors of different origin including breast cancer, and correlated with the worse disease outcome. In addition, significant accumulation of S6K1 was found in the nuclei of breast carcinoma cells suggesting the implication of kinase nuclear substrates in tumor progression. However, this aspect of S6K1 functioning is still poorly understood. The main aim of the present work was to study the subcellular localization of S6K1 in breast cancer cells with the focus on cell migration. Methods: Multicellular spheroids of MCF-7 cells were generated using agarose-coated Petri dishes. Cell migration was induced by spheroids seeding onto adhesive growth surface and subsequent cultivation for 24 to 72 hours. The subcellular localization of S6K1 was studied in human normal breast and cancer tissue samples, 2D and 3D MCF-7 cell cultures using immunofluorescence analysis and confocal microscopy. Results: Analysis of histological sections of human breast tissue samples revealed predominantly nuclear localization of S6K1 in breast malignant cells and its mainly cytoplasmic localization in conditionally normal cells. In vitro studies of MCF-7 cells demonstrated that the subcellular localization of S6K1 depends on the cell density in the monolayer culture. S6K1 relocalization from the cytoplasm into the nucleus was detected in MCF-7 cells migrating from multicellular spheroids onto growth surface. Immunofluorescence analysis of S6K1 and immunocoprecipitation assay revealed the colocalization and interaction between S6K1 and transcription factor TBR2 (T-box brain protein 2) in MCF-7 cells. Conclusions: Subcellular localization of S6K1 depends on the density and locomotor activity of the MCF-7 cells.
Highlights
Ribosomal protein S6 kinase 1 (S6K1) belongs to the AGC family of serine/threonine protein kinases (Ruvinsky & Meyuhas, 2006)
We focused on the study of subcellular localization of endogenous S6K1 in breast tumor and normal tissue, and in breast adenocarcinoma MCF-7 cells in monolayer culture, 3D multicellular spheroids, and in the course of induced cancer cell migration
Immunochemical detection of S6K1 subcellular localization in human breast cancer cells Firstly, the subcellular distribution of S6K1 was determined in the histological sections of human breast cancer and normal tissues
Summary
Ribosomal protein S6 kinase 1 (S6K1) belongs to the AGC family of serine/threonine protein kinases (Ruvinsky & Meyuhas, 2006). Recent studies based on nuclear-cytoplasmic fractionation revealed its presence in the cytoplasm of the breast cancer cells and primary human fibroblasts (Kim et al, 2009; Rosner & Hengstschläger, 2011). Significant accumulation of S6K1 was found in the nuclei of breast carcinoma cells suggesting the implication of kinase nuclear substrates in tumor progression. This aspect of S6K1 functioning is still poorly understood. The subcellular localization of S6K1 was studied in human normal breast and cancer tissue samples, 2D and 3D MCF-7 cell cultures using immunofluorescence analysis and confocal microscopy.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
