On a daily basis, we don't spend much time thinking about trash collection. We take out the garbage when needed, it gets picked up periodically, and the cycle begins again. But if the sanitation workers go on strike, their business suddenly becomes our business. Bags of trash pile up, creating a stinking mess and interfering with our ability to function and maintain a good quality of life.
Cells — which are basically protein factories — create garbage, too, in the form of damaged or defective proteins. Cellular trash disposal is critical for maintaining the quality of proteins in the cell. The cell has systems in various compartments like the endoplasmic reticulum (ER) and cytoplasm that remove the trash. But no such system has ever been identified in the "command center" of the cell — the nucleus — until now.
Researchers in Dr. Dan Gottschling's lab in the Basic Sciences Division spent the last three years searching for the nuclear rubbish route and found the first protein quality-control system in the yeast nucleus. Published in the March 25 issue of Cell, the scientists discovered that the system involves an ubiquitin-protein ligase called San1 — a type of protein that tags bad proteins for destruction.
"Virtually nothing was known about how the cell maintains protein quality in the nucleus," said Dr. Rich Gardner, lead author of the study and a postdoctoral fellow at the center. "That's somewhat surprising since proteins pretty much run the show in the nucleus. The DNA contains the genetic code, but it's wrapped around proteins, copied by proteins and read by proteins. Our goal was to see how the nucleus keeps these proteins pristine."
When trash happens
There are numerous examples of what happens when protein quality control fails in the cell. A number of diseases — Alzheimer's, Huntington's, Parkinson's and even mad-cow disease — are characterized by the accumulation of bad proteins in the brain. When proteins become structurally defective, their insides open up so that the sticky interiors are now exposed, causing the proteins to stick together and clump. "These quality-control diseases are like having a trash strike," Gardner said. "Bags of abnormal proteins are piling up on the front lawn, and that's bad for the cell."
Gardner, who conducted the study along with Gottschling and Zara Nelson, wanted to know how the bad proteins in some of these quality-control diseases elude trash collection in the nucleus. But first, they had to get a better understanding of the nuclear protein quality-control systems. Prior to this finding, some people didn't think a protein quality-control function even existed in the nucleus. The reason was because unlike in the ER and cytoplasm, no protein is made (synthesized) within the nucleus. If the cell is compared to a factory, the ER and cytoplasm are the protein-assembly line, where defectively produced proteins are destroyed as soon as they're detected. The nucleus, on the other hand, is like a retail store, receiving the "finished" protein products from the cytoplasm. Instead of an assembly line quality-control system, the nucleus would likely have a "point-of-service" quality-control system that detects and destroys proteins that became damaged during or after transport into the nucleus.
To chip away at the question, the team first conducted a so-called "virtual screen." They searched through scientific literature archives to find examples of abnormal proteins in the nucleus that shared a common suppressor, or mutation, that compensated for the abnormality and allowed the protein to function normally. They found one: San1. It shared all of the traits of the previously found proteins that provide quality control in the ER. From there, Gardner ran assays in the lab to learn more about San1's characteristics and confirm that this was a nuclear-specific pathway.
Gardner's research was validated by a key discovery. When San1 was disabled, the yeast cell showed a stress response. That meant that when the clean-up protein wasn't doing its job, the bad proteins built up and signaled to the cell that something was wrong in the nucleus. The finding cemented their theory.
Huntington's disease — a degenerative genetic disorder of the brain extensively studied at Fred Hutchinson — is a striking illustration of what happens when bad proteins are allowed to accumulate in the nucleus. Gardner and his team are now working to develop models of Huntington's in yeast to see if they can mimic what occurs in humans and determine if San1, or a similar pathway, is involved. In Huntington's disease, the Huntington protein has an abnormally long track of a single repeating amino acid called glutamine. The researchers speculate that the cell recognizes the abnormally long protein as bad, and a pathway like San1 targets it and sends it to the proteasome, the cellular equivalent of a paper shredder. The expanded protein track is like a thick stack of paper. The proteasome starts to shred it, jams and then grinds to a halt. The proteasome rejects this half-shredded protein mass and the refuse pile accumulates. "The shortened form of this protein is especially prone to aggregate, so if we can keep it in the full-length form, the disease may not occur," Gardner said. "If we can set up a model for Huntington's disease, then we can screen for a drug that inhibits this pathway from letting the protein get chewed down to the smaller form."
Challenges in road ahead
Though excited by the findings, Gardner understands that the road ahead holds many challenges. "We've initially characterized the pathway, but it's only a start," he said. "We want to come to as much of an understanding about protein quality control in the nucleus as we have about DNA quality control. There are so many crucial things that proteins do in the nucleus — including managing the blueprint of life — that nuclear protein quality control is likely to be extensive."
"Although protein quality control is considered a housekeeping function in the cell, we can see that it is exceedingly important in the nucleus and elsewhere in the cell," Gardner said. "Just like in the real world, if the cell doesn't have its house in order, it's going to have a very dysfunctional life."