Abstract
BackgroundLike mammalian MAP kinases, the mating-specific Fus3 MAPK of yeast accumulates in the nuclei of stimulated cells. Because Fus3 does not appear to be subjected to active nucleo-cytoplasmic transport, it is not clear how its activation by mating pheromone effects the observed change in its localization. One possibility is that the activation of Fus3 changes its affinity for nuclear and cytoplasmic tethers.ResultsDig1, Dig2, and Ste12 are nuclear proteins that interact with Fus3. We found that the pheromone-induced nuclear accumulation of a Fus3-GFP reporter is reduced in cells lacking Dig1 or Dig2, whereas Fus3T180AY182A-GFP localization was unaffected by the absence of these proteins. This suggests that Dig1 and Dig2 contribute to the retention of phosphorylated Fus3 in the nucleus. Moreover, overexpression of Ste12 caused the hyper-accumulation of Fus3-GFP (but not Fus3T180AY182A-GFP) in the nuclei of pheromone-treated cells, suggesting that Ste12 also plays a role in the nuclear retention of phosphorylated Fus3, either by directly interacting with it or by transcribing genes whose protein products are Fus3 tethers. We have previously reported that overexpression of the Msg5 phosphatase inhibits the nuclear localization of Fus3. Here we show that this effect depends on the phosphatase activity of Msg5, and provide evidence that both nuclear and cytoplasmic Msg5 can affect the localization of Fus3.ConclusionOur data are consistent with a model in which the pheromone-induced phosphorylation of Fus3 increases its affinity for nuclear tethers, which contributes to its nuclear accumulation and is antagonized by Msg5.
Highlights
Like mammalian MAP kinases, the mating-specific Fus3 Mitogen activated protein kinases (MAPKs) of yeast accumulates in the nuclei of stimulated cells
These data suggest that Dig1 and Dig2 contribute to the nuclear accumulation of Fus3 during the mating response, but either do not affect the localization of Fus3 in vegetative cells, or are functionally redundant in this regard
There is no detectable fluorescence in the nuclei of cells expressing Ptp3-GFP. This demonstrates that our imaging system can discern equal nuclear and cytoplasmic localization from purely cytoplasmic localization. These results suggest that Msg5 is distributed evenly in both compartments, and that its localization does not change upon pheromone treatment
Summary
Like mammalian MAP kinases, the mating-specific Fus MAPK of yeast accumulates in the nuclei of stimulated cells. Because Fus does not appear to be subjected to active nucleocytoplasmic transport, it is not clear how its activation by mating pheromone effects the observed change in its localization. One possibility is that the activation of Fus changes its affinity for nuclear and cytoplasmic tethers. Mitogen activated protein kinases (MAPKs) respond to a wide array of external signals, and are presumed to be present in all eukaryotes. MAPKs phosphorylate targets in the nucleus, in the cytoplasm, and at the cell cortex [5,6]. Because MAPKs affect targets throughout the cell, their localization is a critical aspect of signal regulation [7,8,9]. The duration of ERK nuclear localization has been reported to be very important in determining the outcome of ERK (page number not for citation purposes)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have