Mutation Common to ALS Affects How Lipids Are Metabolized in Motor Neurons, Study Shows

Patricia Inácio, PhD avatar

by Patricia Inácio, PhD |

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Biomarkers and ALS

Loss of C9orf72, a protein whose corresponding gene is among those most often mutated in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), causes an imbalance in the metabolism of fats (lipids) in motor neurons, a new study shows.

These findings suggest that an imbalance in energy and lipid metabolism could be particularly relevant to ALS.

“Cells with this mutation act as if they’re chronically under stress, which could underlie the pathology of diseases associated with this defect,” Jiou Wang, MD, PhD, an associate professor at Johns Hopkins Bloomberg School of Public Health and a study lead author, said in a press release.

“Our study raises the question of whether we should be looking at problems with lipid metabolism as a potential cause for these diseases,” Wang added.

The study, “A C9orf72-CARM1 Axis Regulates Lipid Metabolism Under Glucose Starvation-induced Nutrient Stress,” was published in the journal Genes & Development.

Mutations in the C9orf72 gene have been identified as the most common underlying cause of ALS and FTD, a related neurodegenerative disease. Such mutations lead to lower levels of the C9orf72 protein (whose production the gene oversees), and have also been linked with Alzheimer’s disease and bipolar disorder.

Despite the link with neurodegenerative diseases, the molecular functions of C9orf72 remained largely unknown.

Researchers at the Bloomberg School investigated how the protein works at a cellular level, or its molecular functions.  They deleted the C9orf72 gene from mouse cells (specifically, mouse embryonic fibroblasts) and compared the protein content of these mutant cells to cells with a working gene (control cells).

Experiments were done in cells growing under normal conditions, and in conditions without glucose — a sugar and a key source of energy. The absence of glucose causes what is known as nutrient starvation stress.

Results showed that loss of the C9orf72 protein altered the cell’s fat (lipid) metabolism under glucose starvation. Compared to cells with a fully functional C9orf72 gene, cells lacking the C9orf72 gene and corresponding protein had a much higher load of proteins linked to the metabolism of lipids.

Healthy cells respond to nutrient starvation stress, such as a lack of glucose, by accumulating lipid droplets, i.e., small warehouses filled with lipids.

When comparing normal and C9orf72 protein-depleted cells growing under nutrient starvation stress conditions, the researchers saw that the number of lipid droplets was significantly higher in the mutant (C9orf72-depleted) cells.

The lipid droplets in these cells were filled with a type of lipid called free fatty acids, which are stored as triglycerides — the major lipid found in lipid droplets and the main energy source under conditions of starvation.

Cells can accumulate lipid droplets in two ways: either from scratch, a pathway called de novo lipid biogenesis (or lipogenesis), or by using a recycling pathway, called autophagy, where they degrade cell components to release lipids and make the lipid droplets.

Both pathways, the researchers found, were enriched in cells lacking the C9orf72 gene — meaning that both pathways were abnormal in the absence of the C9orf72 protein.

Researchers then investigated the molecular mechanism underlying C9orf72-mediated regulation of lipid metabolism. Specifically, they aimed to identify the protein(s) interacting with C9orf72, using a technique called stable isotope labeling by amino acids in cell culture (SILAC) mass spectrometry.

Their screen identified a transcription factor (a protein capable of modulating gene expression, or protein production) called CARM1, previously known to influence autophagy and lipid metabolism.

Further experiments were performed to determine if CARM1 was a critical player, linking C9orf72 to lipid metabolism.

This work showed that C9orf72 is an important regulator of CARM1 protein degradation. Cells lacking the C9orf72 protein had higher-than-normal levels of CARM1 and, as a consequence, increased expression of lipid-related genes.

Using cells from ALS patients, researchers first confirmed that C9orf72 levels were low in their cells when compared to normal cells, especially under glucose-deprived conditions.

Motor neurons derived from induced pluripotent stem cell (iPSC) of two ALS and FTD patients with low levels of C9orf72 were also seen to have increased levels of CARM1 compared with controls. The same high levels were detected in spinal cord tissue from three C9orf72-linked ALS/FTD patients.

The motor neurons also again showed increased lipid levels, indicative of dysfunctional lipid metabolism.

These findings suggest that mutations in the C9orf72 gene affect lipid metabolism due to increased expression of CARM1.

“This metabolic imbalance of energy and lipids could be particularly relevant to the pathogenesis of ALS/FTD, which is associated with hypermetabolism [faster metabolism] and hyperlipidemia [increased lipids],” the researchers wrote.

“As we learn more about this newly discovered biological pathway,” Wang said, “it could lead to new therapeutic interventions that protect cells that carry this mutation from harm.”