Study reveals potential target for treatment

Mitochondrial DNA mutations

Image: Mitochondria are engulfed by autophagy
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Credit: Júlio CB Ferreira / USP

A study by Brazilian scientists suggests a possible biological mechanism that modulates the accumulation of mutated mitochondrial DNA in cells during aging, particularly in the liver. The mechanism is called autophagy, and it is a cellular cleansing process that destroys organelles, including mitochondria, and recycles their components. The findings set a paradigm in the field and posit a hypothesis for the emergence of mitochondrial DNA-related diseases, paving the way for the development of therapeutics.

The study was supported by FAPESP and reported in an article published in the magazine autophagy.

Mitochondria are responsible for cellular respiration, processing substrates to provide energy for the organism. They have their own genome inherited only from the mother, unlike the DNA found in the cell nucleus, which is inherited from both parents. Mitochondrial DNA also undergoes mutations, some of which can lead to disease. It is estimated that disorders caused by mutations in mitochondrial DNA affect at least 1 in 5,000 people worldwide. One such disorder is MELAS syndrome (mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes), which can cause seizures, pain, altered consciousness, and focal neurological deficits, among other symptoms.

“Besides disorders directly caused by mutations in mitochondrial DNA, there are other disorders in which mutations contribute at least partly.” Marcos Kerati, the last author of the article. These include diabetes, Parkinson’s disease and Alzheimer’s disease, he added, noting that the study could pave the way for treatments that include autophagy in the context of various diseases.

Chiarati is Professor in the Department of Genetics and Evolution at the Federal University of São Carlos (UFSCar) in the state of São Paulo. The other authors of the article are researchers affiliated with UFSCar and the University of São Paulo (USP) in Brazil, and with Cedars-Sinai Medical Center and Stanford University in the United States. The study was led by Chiaratti and supported by FAPESP across seven projects (16 / 07868-4And the 17 / 05899-2And the 17 / 04372-0And the 18/20028-0And the 20 / 15412-6And the 19 / 25049-9And the 12 / 50231-6).

aging

Research has shown that mitochondrial DNA mutations accumulate as part of aging. “One of the oldest questions in biomedicine is why,” Chiarati said. Previous studies have ruled out theories that blame mitochondrial DNA replication or cell proliferation. Given the importance of autophagy in the life cycle of cells and mitochondrial recycling, Kirati and colleagues decided to see if it plays a role in mitochondrial DNA mutations. “Autophagy is in vogue. The scientific literature on this topic is growing,” he said. The 2016 Nobel Prize in Physiology or Medicine was awarded to Japanese cell biologist Yoshinori Ohsumi for his discoveries regarding the mechanisms of autophagy.

methodology

Practically all humans have a small amount of mutated DNA. The coexistence of mutant and wild-type (non-mutant) mitochondrial DNA in a cell is called heterochromatin. In this study, Chiaratti et al. Mice with 30% heterozygous plasma were used: a mutant strain known as NZB/BINJ (or NZB) represented 30% of the mitochondrial genome, while the rest was C57BL/6N (or BL6), considered ‘normal’.

The researchers crossed these mice with one that lacked the gene atg7 Specifically in the liver. Without this gene, the mechanism required to make autophagy possible disappears. “We spent almost a year reproducing,” Chiarati said.

After this period, they had two groups of mice: a control group containing mitochondrial DNA NZB and the . gene atg7; And group with NZB but without the gene atg7 in hepatocytes, so autophagy in the liver was impossible (liver specific atg7 Kassa).

Experiments were performed to see if autophagy affected the accumulation of mutant mitochondrial DNA over time. Ten days after birth, similar levels of NZB were found in the tissues of the two groups of mice, but after 21 days NZB began to accumulate in the livers of control animals, while the knockout group had the same percentage as before. After 100 days, when mice are considered adults, controls had higher levels of NZB while the knockouts had virtually the same levels as when they were born, making the difference between the two groups more pronounced and indicating that autophagy plays a role in age-related mitochondrial DNA accumulation. .

The researchers then set out to validate this finding by conducting an experiment on mice lacking the gene Prkn, which is responsible for a certain stage of autophagy. in Prkn For knockout mice, autophagy occurred but was defective. “We once again compared these mice to controls,” Chiarati said. although Prkn Ultimately, mice that knocked out NZB mitochondrial DNA knockdown collected, and the accumulation was smaller than that of the control group. “This reinforced the theory that autophagy was involved since manipulation of another related gene led to changes in the data.”

The researchers also found that autophagy mostly removed BL6 mitochondrial DNA in controls, which likely explains the higher levels of NZB. When exploring this possibility further, they discovered that the presence of NZB enhances the respiratory capacity of the mitochondria in hepatocytes. “Therefore, we hypothesized that autophagy might help eliminate the least potent mitochondria while preserving the fittest, which is a positive of course,” Chiarati said.

Another phase of the study involved isolating fibroblasts, a type of connective tissue cell, from mice, and observing in the lab that their levels of mitochondrial NZB DNA decreased over time. However, when autophagy-inducing drugs were used, the decrease was reversed. “This was further evidence of the importance of autophagy in regulating mitochondrial DNA,” Chiarati said.

Next steps

While NZB was considered mutated in the mice participating in these experiments, the results showed that it is not harmful to the liver. On the contrary, it enhances the respiratory capacity of mitochondria in hepatocytes, as has already been noted. “Now we need to investigate this phenomenon using various mitochondrial DNAs that are known to be pathological,” Chiarati said.

If autophagy also proves appropriate in this context, researchers can begin to test treatments that stimulate the cell’s clearing mechanism against diseases that affect mitochondrial function. Chiarati added that other possibilities for future research will include the role of autophagy in the accumulation of mitochondrial DNA in tissues elsewhere in the body beyond the liver and isolated fibroblasts.

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About the São Paulo Research Foundation (FAPESP)

The São Paulo Research Foundation (FAPESP) is a public institution whose mission is to support scientific research in all fields of knowledge by offering scholarships, fellowships and grants to researchers associated with institutions of higher education and research in the state of São Paulo, Brazil. FAPESP understands that only the best research can be done by working with the best researchers in the world. Therefore, it has established partnerships with funding agencies, higher education, private companies, and research organizations in other countries known for the quality of its research and encourages the scholars funded by its grants to further develop their international collaborations. You can learn more about FAPESP at www.fapesp.br/en And visit FAPESP news agency at www.agencia.fapesp.br/en To keep abreast of the latest scientific achievements that FAPESP helps to achieve through many programs, awards and research centers. You can also subscribe to FAPESP news agency at http://agencia.fapesp.br/subscribe.