Proteins with short lifespans control gene expression in cells and perform many crucial tasks, from aiding in brain connectivity to assisting the body in immune defense. These proteins are produced in the cell nucleus and are rapidly degraded once their tasks are completed. Despite their significance, scientists have been puzzled for decades about how these proteins undergo degradation and are cleared from cells when they are no longer needed.
In a recent study, researchers from Harvard Medical School in the United States identified a protein called “midnolin” that plays a key role in the degradation of many short-lived nuclear proteins. Their research indicates that midnolin directly seizes these proteins and pulls them into the cell’s waste disposal system, the proteasome, where they are broken down. The research findings were published in the August 25, 2023, issue of the journal Science, titled “The midnolin-proteasome pathway catches proteins for ubiquitination-independent degradation.”
Xin Gu, a co-first author of the study and a neurobiology researcher at Harvard Medical School, said, “These specific short-lived proteins have been known for over 40 years, but no one knew exactly how they underwent degradation.”
As the short-lived proteins degraded in this process regulate critical functions related to the brain, immune system, and development, scientists may eventually be able to target this process as a means of controlling protein levels, altering these functions, and correcting any functional disorders.
Christopher Nardone, a co-first author of the study and a doctoral student in genetics at Harvard Medical School, added, “The mechanism we discovered is very simple and elegant. It’s a fundamental scientific discovery with far-reaching implications for the future.”
Unraveling the Molecular Mystery
It is well known that cells can degrade proteins by marking them with a small molecule called ubiquitin. The marking signals to the proteasome that these proteins are no longer needed, leading to their degradation. Much of the pioneering research on this process was conducted by the late Fred Goldberg at Harvard Medical School. However, there are times when proteins are degraded by the proteasome without the assistance of ubiquitin marking, leaving scientists to speculate about another ubiquitin-independent protein degradation mechanism.
Nardone stated, “There have been sporadic pieces of evidence in the literature suggesting that the proteasome can somehow directly degrade proteins without ubiquitin, but nobody understood how it worked.”
A group of proteins that seemed to be degraded through an alternative mechanism are stimuli-induced transcription factors: rapidly produced proteins in response to cell stimuli that enter the cell nucleus to turn genes on and are then rapidly degraded.
Gu explained, “What initially struck me was how unstable these proteins are; they have very short half-lives, meaning they are rapidly degraded once they’ve performed their function.”
Michael Greenberg, a co-corresponding author of the study and a professor of neurobiology at Harvard Medical School’s Blavatnik Institute, mentioned that these transcription factors play vital roles in a range of biological processes in the body but, even after decades of research, “their turnover mechanisms are largely unknown.”
From a Few to Hundreds
To investigate this mechanism, the researchers began by focusing on two well-studied transcription factors, Fos and EGR1. The Greenberg lab extensively studied Fos for its role in learning and memory, while EGR1 is involved in cell division and survival.
Using complex protein and gene analysis methods developed by the Elledge lab, the researchers honed in on midnolin as a protein that contributes to the degradation of both Fos and EGR1. Subsequent experiments revealed that, in addition to Fos and EGR1, midnolin may also be involved in degrading hundreds of other transcription factors found in the cell nucleus.
Gu and Nardone recalled feeling shocked and skeptical about their own research findings. To confirm their discovery, they set out to understand how midnolin targeted and degraded such a wide array of proteins.
Nardone said, “Once we identified all these proteins, there were still many puzzling questions about how this degradation mechanism of midnolin actually worked.”
Through the use of a tool that predicts protein structures, coupled with results from a series of laboratory experiments, the researchers filled in the details of this mechanism. They found that midnolin possesses a “catch domain,” a region of the protein capable of capturing other proteins and delivering them directly to the proteasome for subsequent degradation. This catch domain consists of two independent regions linked by amino acids, allowing midnolin to capture various types of proteins by grasping a relatively unstructured region within each protein.
Notably, proteins like Fos are responsible for turning genes on, allowing neurons in the brain to connect and reconnect in response to stimuli. Other proteins, such as IRF4, activate genes that ensure the production of functional B cells and T cells to support the immune system.
Elledge stated, “The most exciting aspect of this new study is that we now understand a general mechanism for a new ubiquitin-independent degradation protein.”
Promising Transformative Potential
In the short term, the researchers hope to delve deeper into the mechanism they’ve uncovered. They plan to conduct structural studies to better understand how midnolin captures and degrades proteins in detail. They are also creating mice that lack midnolin to investigate the role of this protein in different cells and developmental stages.
The researchers believe that their discovery holds promising transformative potential. It may provide a pathway for scientists to control the levels of transcription factors, thereby regulating gene expression and influencing related processes within the body.
Greenberg remarked, “Protein degradation is a critical process, and dysregulation of protein degradation is at the basis of many diseases,” including certain neurological and psychiatric disorders, as well as some cancers.
For instance, when transcription factors like Fos are present in excess or deficiency, it may lead to learning and memory problems. In multiple myeloma, cancer cells become addicted to the immune protein IRF4, exacerbating the disease. The authors are particularly interested in identifying diseases that might be treated by developing therapies based on the midnolin-proteasome pathway.