Researchers are discovering how too much oxygen damages cells and tissues

When it comes to oxygen, you can have too many good things. Breathing air containing higher levels of oxygen than the usual 21 percent found in Earth’s atmosphere can cause organ damage, seizures, and even death in humans and animals, particularly if it exceeds the body’s oxygen needs. Until now, however, scientists have mostly speculated about the mechanisms behind this phenomenon, known as oxygen toxicity or hyperoxia.

Now, researchers at the Gladstone Institutes have discovered how excess oxygen changes a handful of proteins in our cells that contain iron and sulfur – a chemical process similar to iron rusting. In turn, these “rusty” proteins trigger a cascade of events that destroy cells and tissues. The findings were published in the journal Molecular Cellthey have implications for conditions such as heart attacks and sleep apnea.

“This study allowed us to create a very specific timeline of what happens in hyperoxia,” says Gladstone research assistant Isha Jain, Ph.D., senior author of the new study. “The results were not at all what we expected, but it is very interesting and exciting to now know how this series of events is unfolding.”

A spoiled question

At high levels, oxygen is toxic to all life forms, from bacteria and plants to animals and humans. Of course, insufficient oxygen is also deadly. There is an intermediate amount, “Goldilocks” below which most life on Earth thrives — neither too much nor too little.

While clinicians have long studied the details of how a lack of oxygen affects cells and tissues (for example, in heart attacks and strokes), the effects of excess oxygen have been relatively understudied.

“For many years, the medical teaching was that to some degree, more oxygen was better, or at least benign, when treating patients with conditions like heart attacks,” says Alan Baik, MD, a postdoctoral researcher in the Jain lab and a cardiologist at UC San Francisco (UCSF). “But there are now a growing number of clinical studies showing that excess oxygen actually leads to worse outcomes. This motivated us to better understand why excess oxygen can be toxic.”

Studies have recently revealed, for example, that breathing too much supplemental oxygen can be harmful to heart attack patients and premature infants. Similarly, in obstructive sleep apnea, the sudden bursts of oxygen that follow pauses in breathing have been shown to be a key component of how the disorder increases patients’ risk of chronic health problems.

However, the mechanisms of these effects remained obscure. Many researchers hypothesized that reactive oxygen species—unstable and highly reactive oxygen derivatives that can damage our genome and many molecules in our cells—probably played a role in hyperoxia, but there was little evidence of how excess oxygen affected specific enzymes and streets.

How CRISPR found the answer

Jain’s team—including Baik, postdoctoral fellow Galih Haribowo, Ph.D., and graduate student Kirsten Xuewen Chen, who are first authors of the new paper—turned to the CRISPR genome-editing technology to test the roles of a gene diversity in hyperoxia.

Using CRISPR, the researchers removed, one at a time, more than 20,000 different genes from human cells grown in the lab, then compared the growth of each group of cells in 21 percent oxygen and 50 percent oxygen.

“This kind of unbiased screen allows us to investigate the contribution of thousands of different pathways to hyperoxia instead of just focusing on those we already suspected were involved,” says Jain, who is also an assistant professor of biochemistry at UCSF. “It led us to molecules that have never been said before in the same sentence as oxygen toxicity.”

Four molecular pathways stood out in the screen as being involved in the effects of hyperoxia. They involved various cellular functions, including the repair of damaged DNA, the production of new DNA building blocks, and the production of cellular energy.

Common protein clusters

At first, the team couldn’t figure out what the four pathways had in common and why they were all affected by high oxygen levels. It took some molecular research to discover that each pathway had a critical protein that contained iron atoms bonded to sulfur atoms—so-called “iron-sulfur clusters”—in its molecular structure.

The researchers went on to show that at just 30 percent oxygen, the iron-sulfur clusters in the four proteins oxidize—react chemically with oxygen atoms—and this change causes the proteins to degrade. As a result, the cells stop working properly and consume even less oxygen, causing further increases in oxygen levels in the surrounding tissues.

“An important point is that hyperoxia does not affect cells and tissues only through reactive oxygen species, as many had assumed,” says Jain. “This means that the use of antioxidants—which can fight reactive oxygen species to some extent—is unlikely to be sufficient to prevent oxygen toxicity.”