Identification of novel Saccharomyces cerevisiae proteins with nuclear export activity: Cell cycle-regulated transcription factor Ace2p shows cell cycle-independent nucleocytoplasmic shuttling

Nuclear export of proteins containing leucine-rich nuclear export signals ( NESs) is mediated by the NES receptor CRM1/Crm1p. We have carried out a yea st two-hybrid screen with Crm1p as a bait. The Crm1p-interacting clones wer e subscreened for nuclear export activity in a visual assay utilizing the C rm1p-inhibitor leptomycin B (LMB). This approach identified three Saccharom yces cerevisiae proteins not previously known to have nuclear export activi ty. These proteins are the 5' RNA triphosphatase Ctl1p, the cell cycle-regu lated transcription factor Ace2p, and a protein encoded by the previously u ncharacterized open reading frame YDR499W. Mutagenesis analysis show that Y DR499Wp contains an NES that conforms to the consensus sequence for leucine rich NESs. Mutagenesis of Ctl1p and Ace2p were unable to identify specific NES residues, However, a 29-amino-acid region of Ace2p, rich in hydrophobi c residues, contains nuclear export activity. Ace2p accumulates in the nucl eus at the end of mitosis and activates early-G(1)-specific genes. We now p rovide evidence that Ace2p is nuclear not only in late M-early G(1) but als o during other stages of the cell cycle. This feature of Ace2p localization explains its ability to activate genes such as CUP1, which are not express ed in a cell cycle-dependent manner.