How cellular processes round and dispose of damaged proteins

Strieter says, “It took eight years to find out and I’m very proud of the work. We have had to develop many new methods and tools to understand what this enzyme does. ”

As he explains, a very large protease called a proteasome is responsible for breaking down the vast majority of proteins in a cell. It can contain up to 40 proteins. It has been known for more than 20 years that UCH37 is one of the regulatory enzymes associated with the proteasome, he adds, “but nobody understood what it did.”

It turns out that the crux of the whole process is how there can be complex changes in a small protein called ubiquitin. “In addition to modifying other proteins, ubiquitin modifies itself, which leads to a multitude of chains. Some of these chains can be highly branched. We found that UCH37 is removing the branch points from the chains, allowing mining to continue. ”

Write this week in Molecular cell, he and lead author and Ph.D. candidate Kirandeep Deol, who directed and conducted the experiments with co-authors Sean Crowe, Jiale Du, Heather Bisbee, and Robert Guenette, discussed how they answered the question. The work was supported by the National Institute of General Medical Sciences at the NIH.

This advance could eventually lead to a new cancer treatment, says Strieter, because cancer cells need the proteasome to grow and multiply. “Many cancer cells are essentially dependent on the function of the proteasome,” he points out. “Its cells produce proteins at such a rapid rate that mistakes are made, and if they are not corrected, the cells cannot function. Since UCH37 helps to eliminate proteins, it could be a useful therapeutic target for supplementing proteasome inhibitors that have already been successful in the clinic. ”

To begin his long-term process, Strieter said: “We had to find a way to create a multitude of ubiquitin chains that represent potential diversity in a cell. With this new ubiquitin chain library we were able to question the activity of UCH37 in a controlled environment. This series of experiments gave us the first indication that this enzyme does something unique. ”

Another new method they have developed uses mass spectrometry to characterize the architecture of ubiquitin chains in complex mixtures. “This allowed us to see that the activity we discovered with our substrate library was also present in a more heterogeneous mix,” says Strieter. Finally, the chemists used the CRISPR gene editing tool to remove UCH37 from cells to measure the impact of UCH37 on proteasome-mediated degradation in vitro and in cells.

This technique led to another surprise. “Instead of acting as intended and resisting the degradation process, UCH37 was found to remove the junction points from the ubiquitin chains to help break down the proteins,” says Strieter. “You might think that removing the signal for degradation would affect degradation,” he adds, “but it didn’t work that way.”

In future experiments, Strieter and colleagues hope to further investigate the degradation process and learn in more detail how UCH37 regulates cell function.

Proteasome phase separation for destruction

More information:
Kirandeep K. Deol et al., Proteasome-Bound UCH37 / UCHL5 disassembles ubiquitin chains to promote degradation, Molecular cell (2020). DOI: 10.1016 / j.molcel.2020.10.017

Provided by the University of Massachusetts Amherst