May 07, 2014
In the Sherlock Holmes mystery story “The Adventure of the Crooked Man,” a man and his wife are heard having an argument behind closed doors. There is a crash, and the doors are opened to reveal that the man is dead in a pool of blood and his wife has fainted. No one else is nearby, and it seems beyond any doubt that the wife has murdered her husband…of course, until Sherlock Holmes delves into the details and uncovers the real story.
Something similar is going on in the nucleus behind those nuclear pores. The proteasome is a huge molecular machine that recognizes and degrades ubiquitinated proteins. Scientists have seen key parts of the proteasome in the nucleus, and nuclear proteins are degraded by this complex. Seems like an open and shut case that the proteasome degrades nuclear proteins in the nucleus. But like our Sherlock Holmes story, things aren’t always as they appear.
Chen and Madura applied some detective work to the details of our nuclear protein degradation mystery in a new study published in GENETICS and found that it probably doesn’t work this way at all. Nuclear proteins need to be exported out of the nucleus to be degraded by the proteasome.
The researchers first confirmed earlier work showing that the Sts1 protein is responsible for escorting proteasomes to the nucleus. In the temperature-sensitive sts1-2 mutant at the restrictive temperature of 37 degrees, two proteasome subunits from different subcomplexes of the proteasome (Rpn11p from the regulatory particle and Pup1p from the catalytic particle) didn’t make it into the nucleus.
They then looked at two nuclear proteins, Rad4p and Pol1p (also known as Cdc17p) that are substrates of proteasomal degradation. When the proteasome subunits Rpn11p and Pup1p didn’t make it into the nucleus because of the sts1-2 mutation, Rad4p and Pol1p were not degraded.
So the proteasome needs to get into the nucleus in order for nuclear proteins to get degraded. Sounds like the proteasome is guilty of degrading proteins in the nucleus. But like a good mystery novel, the story takes an interesting twist here.
Chen and Madura found that when they raised the temperature of the sts1-2 mutant cells, Rad4p and Pol1p were stabilized immediately. This didn’t really make sense though. Even if the temperature-sensitive mutation blocked import of proteasomes into the nucleus as soon as the temperature increased, the proteasomes already inside the nucleus should have been able to continue degrading their substrates.
Wondering whether the substrates might be exported from the nucleus to be degraded elsewhere, they tested what happened to Rad4p and Pol1p when nuclear export of proteins was blocked. Using a few different ways to prevent nuclear export (combinations of mutations, chemicals, and temperature), they showed that if Rad4p and Pol1p could not get out of the nucleus, they were not degraded by the proteasome.
So it’s clear that nuclear export is part of the degradation process for at least these two nuclear proteins. Chen and Madura also detected a general increase in multi-ubiquitinated proteins (tagged for proteasomal degradation) in the nucleus under conditions where export was blocked, suggesting that this mechanism may apply to other proteins as well. And it’s been shown in human cells that several specific proteins, including the tumor suppressor p53, need to get out of the nucleus to be degraded.
There are still a lot of details to be filled in about where degradation is really happening. An intriguing clue comes from the fact that Sts1p is distantly related to the Schizosaccharomyces pombe protein Cut8, which is a nuclear envelope protein that tethers the proteasome to the nuclear membrane. Might nuclear proteasomes work on the outside of the nuclear envelope?
More detective work is needed to answer this question. But it’s clearly a very important one. Nuclear export and protein degradation are highly conserved processes, and both are currently under study as potential targets of cancer treatment.
Let this be a warning to all of us not to take everything at face value. Just because someone is holding the bloody knife, that doesn’t mean he is the murderer. And just because subunits and subcomplexes of the proteasome machinery look to be in the nucleus, that doesn’t mean nuclear proteins are degraded there. It is elementary, my dear Watson.
by Maria Costanzo, Ph.D., Senior Biocurator, SGD
Categories: Research Spotlight