Aminoacyl-tRNA synthetases (aaRSs) are a family of ancient enzymes. Their primary function is to build proteins by loading up the proper amino acids to charging transfer ribonucleic acids (tRNAs). In recent decades, this protein family has been increasingly studied for non-classical functions beyond protein synthesis.
In humans, this family of proteins is encoded by 37 genes, with 17 encoding the cytoplasmic aaRSs, 17 encoding the mitochondrial forms, and 3 bi-functional aaRSs functioning in both cytoplasm and mitochondria. Up until 2017, 31 out of 37 aaRS genes have been implicated in human diseases including Charcot-Marie-Tooth disease, a neuropathy, as well as a large number of other conditions associated with both neurological and non-neurological manifestations.
Phe-tRNA synthetase (FARS) is a cytoplasmic aaRS composed of two subunits: alpha and beta. These two subunits of FARS are encoded by separate genes — FARSA and FARSB, respectively. Neither gene had been implicated in human diseases when we started our study.
Our study began in Dr. Wendy Chung’s laboratory at Columbia University, when her group identified a family with a novel disease. In this family, both parents are asymptomatic, but two of their three children had an unusual condition that affected their blood vessels and lungs. Their daughter died at the age of 10 when she had a sudden, unexpected stroke. Her autopsy showed that the condition affected multiple organs including the lungs, liver, blood vessels throughout the body, and calcifications in the brain. By sequencing the exome (the part of the genome that encodes the instructions to make proteins), two mutations in the FARSB gene were identified. The parents each carry one of the mutations and are healthy and are “carriers” for the “hidden” (recessive) condition. Both children shared the two mutations identified in their parents, thereby unmasking the morbid condition.
In addition to the family identified by Dr. Chung, our collaborators in Germany and the United Kingdom, led by Drs. Matthias Griese, Robert W. Taylor, and Holger Prokisch identified three other families with patients carrying additional FARSB mutations and sharing similar features, especially similar lung disease with cholesterol deposits in the lungs. They also shared similar facial features.
We then investigated where the mutations were located in the FARSb protein. In the 3-dimensional structure of FARS, they appear to be localized in two clusters. None of these residues is known to be directly involved in protein synthesis-related activities.
To experimentally explore the disease mechanism, we went back to the first family. Dr. Chung obtained blood and skin cells from the surviving patient and both parents and from normal healthy controls. Prof. Paul Schimmel and Dr. Leslie Nangle’s team at the Hong Kong University of Science and Technology performed experiments with these cells and found that less FARSB was produced in the patients and their parents compared to controls. Despite a reduction of FARS proteins in patient’s cells, the global protein synthesis rate remained similar across the patient, the parents and control cells. These findings suggest that the reduced FARS protein level was sufficient to support protein synthesis in patient cells.
These findings were published in the American Journal of Human Genetics in July 2018. Around the same time, there were two other publications by independent groups, both reporting recessive mutations in the FARSB gene (distinct from those we identified) in patients with similar features. One reported 8 affected individuals from an extended family with the same mutation in both copies of the FARSB gene. Because the studies were independently identified and are so similar, the findings give us great confidence that the results of the three independent groups are correct.
The next question is: what causes the disease? Our study demonstrated that the reduced FARS protein level was sufficient to support protein synthesis. This is not surprising, as we know about other diseases associated with aaRSs. Other aaRS recessive mutations are associated with variable but somewhat overlapping features, and there are suggestions that other aaRS recessive disorders are not attributable to the disruption of the protein translational machinery. Accumulating evidence has demonstrated non-translational activities of aaRS proteins in various signaling pathways that affect metabolism, development, angiogenesis, tumorigenesis, neuronal function, immune function, and inflammation.
We hypothesize that symptoms in our disease may be caused by the disruption of a novel ex-protein synthesis function of FARS. Further functional studies of these FARSB genetic variants in model systems are in progress, which may shed light on the disease mechanism and the expanded functions of FARS, as well as provide insight into other aaRS recessive disorders.
These findings are described in the article entitled Bi-allelic Mutations in Phe-tRNA Synthetase Associated with Multi-system Pulmonary Disease Support Non-translational Function, recently published in the American Journal of Human Genetics.