The nuclear export of mRNA is mediated by proteins that associate with it, during transcriptional elongation. Some of them target the mRNP to the NPC, for export out of the nucleus. Next the mRNP is disassembled in the cytoplasm, RNA is used for translation, and the export factors are returned to the nucleus. Using tandem affinity purification (TAP) of genomic tagged rae1, we identified several nuclear pore proteins that associate with Rae1 in fission yeast. Nup189, Nup186, Nup184, Nup170, Nup146, Nup132, Nup124, Nup120, Nup107, Npp106, Nup97, Nup61, Nup45, Nup44, Nup40 and Nsp1 co-purified with TAP tagged Rae1. Further studies revealed Rae1 associates and functions with proteins found on both the nuclear and cytoplasmic side of the pore. Suggesting, Rae1 may have a role not only in assembling the mRNP in the nucleus, but also in its disassembly in the cytoplasm. Dbp5, an ATP-dependent RNA helicase, also co-purified with Rae1. It has been shown to be involved in releasing mRNA in the cytoplasm. We found that it associates with proteins, Dss1and Uap56, in addition, to Rae1. In these studies, we show that the above proteins not only function in export of the mRNP, from the nucleus to the cytoplasm, but also in its potential disassembly in the cytoplasm. Additionally, we found that Amo1, an NPC associated protein, also, co-purified with Rae1. It has been shown to be required for proper microtubule organization. Further studies revealed its NPC localization is Rae1-dependent. One implication of these studies is that Rae1 may have a role in the proper organization of microtubules. Indeed, we identified a mutation within Rae1in which the microtubules were not properly assembled. Previously, we reported that Dss1, a multifunctional protein involved in mRNA export with Rae1, plays a critical role in linking DNA repair and checkpoint proteins, to damaged DNA sites, by recruiting Rad24 and Cdc25 to the double strand break sites. We have suggested, the sequestration of Cdc25 to DNA damage sites, likely provides a mechanism for S. pombe cells to arrest at G2/M boundary, in response to DNA damage. The involvement of mRNA export factors Rae1 and Dss1, in multiple cellular processes, raises the possibility that mRNA export, DNA-damage/cell cycle processes and cytoskeletal organization, could be coupled.
|Selvanathan, Saravana P; Thakurta, Anjan G; Dhakshnamoorthy, Jothy et al. (2010) Schizosaccharomyces pombe Dss1p is a DNA damage checkpoint protein that recruits Rad24p, Cdc25p, and Rae1p to DNA double-strand breaks. J Biol Chem 285:14122-33|