Calcium transport into and out of the mitochondria – the powerhouses of cells – is central to cellular energy production and cell death. To maintain calcium balance in these powerhouses, cells rely on a protein known as the mitochondrial sodium-calcium exchanger, or NCLX. In a new research paper, scientists at the Lewis Katz School of Medicine at Temple University have now discovered a novel regulator of NCLX activity, a protein called TMEM65, which helps remove calcium from mitochondria, protecting against harmful calcium overload.
Increasing NCLX Activity Against Heart Disease and Alzheimer’s Disease
The discovery, described online in the journal Nature Metabolism, is the first to characterize the interaction of TMEM65 with NCLX in mitochondria. “TMEM65 is the first protein identified as a true interactor and regulator of NCLX,” said Dr. John W. Elrod, holder of the W.W. Smith Chair in Cardiovascular Medicine and founding director of the Aging + Cardiovascular Discovery Center at the Lewis Katz School of Medicine and lead investigator of the new study. The discovery could help scientists develop new therapeutics to combat mitochondrial calcium overload in diseases such as heart failure and Alzheimer’s disease.
Mitochondrial calcium exchange plays a crucial role in regulating cell survival and pro-energetic signaling pathways. When mitochondria take up too much calcium, which can be the case in certain disease states, energy metabolism is disrupted and cells die. This is most evident in the heart, where calcium overload contributes to the permanent loss of heart muscle cells in heart attacks and heart failure. It can also lead to the loss of brain cells in Alzheimer’s disease and other neurodegenerative diseases.
Dr. Elrod and his colleagues have already identified NCLX as a key factor in the removal of calcium from mitochondria in the heart and brain. Research has also shown that an increase in NCLX activity can limit the progression of not only heart failure and Alzheimer’s disease, but also cancer. Despite these promising results, the understanding of the mechanisms underlying NCLX regulation remains unclear. “NCLX has a very complex structure, which has hindered the study of its regulation and progress in therapeutic development,” said Dr. Elrod. For their latest study, the researchers opted for a different approach, using biotin labels that allowed them to track the interactions of NCLX with other proteins in intact cells.
Possible Therapeutic Strategy
Under the direction of postdoctoral fellow Joanne F. Garbincius, PhD, Dr. Elrod’s team generated a fusion of NCLX and a biotinylation protein. The fusion protein was then reintroduced into cells, and other proteins in its vicinity were biotinylated or biochemically labeled. The biotinylated molecules were then easily isolated and identified using mass spectrometry. In this way, the researchers finally discovered TMEM65 as a prime suspect in NCLX regulation.
In subsequent experiments, it was found that calcium levels in the mitochondria accumulate when TMEM65 is removed from the cells. This led to the realization that TMEM65 is required for NCLX activity. Its role in the regulation of NCLX was confirmed in a mouse model in which TMEM65 levels were significantly reduced. As the animals matured, there was a progressive loss of neuromuscular function, such that they were barely able to walk in adulthood. The methods used to identify TMEM65 and to elucidate NCLX regulation are groundbreaking in the field of basic cardiovascular research.
In 2024, Dr. Garbincius was awarded the American Heart Association’s Louis N. and Arnold M. Katz Basic Science Research Prize for Early Career Investigators for her research. The work has also inspired ongoing investigations of TMEM65. Dr. Elrod and her colleagues next plan to explore the possibility of modulating TMEM65 activity as a therapeutic strategy. If the experts can figure out how to enhance or otherwise alter its interaction with NCLX, this could be an important treatment option for patients affected by diseases that cause pathogenic calcium deposition in the mitochondria.