Abstract 76: KPNA7 nuclear import protein - a critical regulator of cancer cell growth

Abstract

Abstract Background. In eukaryotic cells, the nucleus is separated from the cytosol by the nuclear envelope, thus requiring a specific machinery for the transport of macromolecules between the two cellular compartments. The nuclear import cycle is strictly regulated and its malfunction results in incorrect localization of proteins, ultimately leading to variety of diseases including cancer. Karyopherin alpha 7 (KPNA7) is the newest member of the karyopherin alpha family of nuclear importers. KPNA7 is mainly expressed in early embryogenesis and oocytes as well as in some cancer cells. We previously showed that silencing of KPNA7 in pancreatic cancer cell lines with high endogenous expression dramatically reduced cell proliferation and anchorage-independent growth, through a G1 cell cycle arrest and transcriptional induction of p21 expression. Here, we further explored the functional importance of KPNA7 in a large panel of pancreatic and breast cancer cell lines and identified its cargo proteins that may be responsible for the observed phenotypes. Methods. KPNA7 was silenced in pancreatic and breast cancer cells using siRNAs. Cell numbers was determined 72 to 96 h after transfection and compared to those in corresponding controls. To isolate KPNA7 cargo proteins, stable tagged KPNA7-overexpressing pancreatic cancer cell lines were established. An affinity-based protein pull-down was performed and the KPNA7-interacting partners were identified with mass spectrometry. Results. The silencing of KPNA7 in six pancreatic and breast cell lines led to decreased proliferation in all cell lines with low to medium expression level, whereas both cancer and normal cell lines with absolutely no KPNA7 expression were not affected, thus confirming the specificity of the phenotype to KPNA7 silencing. The results were also consistent in breast cancer cell lines, implicating a role not restricted to pancreatic cancer. Interestingly, in some cell lines a distinct change from round to lobular nuclear morphology was observed, suggesting that KPNA7 may also participate in the maintenance of nuclear architecture. The protein pull-down followed by mass spectrometry yielded multiple proteins co-purifying with KPNA7. These included several proteins involved in RNA processing and nucleopore proteins. Many of these were also involved in the regulation of cell cycle and p21 pathway, consistent with the phenotype observed in the functional studies. Conclusions. These results implicate KPNA7 as an important regulator of cancer cell growth and suggest that it might also have other functions in the maintenance of cancer cell homeostasis. We were the first to link KPNA7 to a human malignancy and here we expand that notion beyond pancreatic cancer. We have also identified many putative KPNA7 cargos with functions matching the observed cellular phenotypes. This study provides additional evidence on the role of altered nuclear transfer in cancer pathogenesis. Citation Format: Elisa M. Vuorinen, Nina Rajala, Hanna E. Rauhala, Anne Kallioniemi. KPNA7 nuclear import protein - a critical regulator of cancer cell growth. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 76.