Transfer RNA-mediated enhancement of RSK/MSK signaling and its connection to tumorigenesis

Abstract

Transfer RNA (tRNA) is essential RNA for protein synthesis, transporting amino acids to the ribosome. tRNA has been known for house-keeping RNA for a long time, however, recent studies suggest that tRNA can be a multi-functional molecule that controls various cellular events including translation initiation and cell apoptosis. In various cancer cell lines and tissues, although increased expression of tRNA was observed, the function of the overexpressed tRNA was unclear. We hypothesized that tRNA would play a pro-oncogenic role during tumorigenesis, therefore we overexpressed tRNA in 293T cells and investigated the effect of tRNA overexpression on cell proliferation and survival. Interestingly, 293T cells with high expression level of tRNA, especially tRNALeu, showed increased resistance to cell death and enhanced phosphorylation of 90-kDa S6 kinase under amino acid starvation condition. Phosphorylation of 90-kDa S6 kinase was reduced by anti-tRNALeu RNA, p38MAPK inhibitor and ERK inhibitor, but not by treatment of mTOR inhibitor, rapamycin. RSK1 (Ribosomal S6 kinase 1) and MSK2 (Mitogen-and stress-activated protein kinase 2) were identified as the tRNALeu-responsive effectors among the tested 7 candidate proteins categorized as 90-kDa S6 kinase. Considering that RSK1 and MSK2 are downstream regulators of p38MAPK and ERK and play an important role in hormonal cancer such as estrogen-responsive breast cancer, we analyzed the expression level of tRNALeu in breast cancer cell lines and its responsiveness to ß-estradiol. Estrogen receptor (ER) positive MCF7 cell line showed the highest expression levels of tRNALeu isotypes compared with ER negative cell lines and its tRNALeu expression was induced by ß-estradiol treatment. We also investigated the protein binding partner of tRNALeu under amino acid starvation condition using SILAC (Stable Isotope Labeling by/with amino acids in cell culture) mass spectrometry and identified EBP1 (ErbB3-binding protein), which is the upstream regulator of RSK/MSK signal pathway. These results indicate that tRNALeu can be induced by estrogen and enhance EGFR signaling via binding with EBP1, subsequently activating RSK/MSK to increase survival of breast cancer cells under amino acid starvation condition. This study also suggests a possibility that tRNA can work as a new cellular regulator beyond translation and its novel function in tumorigenesis.