Sartori-Maldonado, R. | Search bioRxiv

bioRxiv2024

Finnish gyrate atrophy mutation OAT;c.1205C>T leads to accumulation of intracellular GABA

Hyperornithinaemia with gyrate atrophy of choroid and retina (HOGA) is a recessive metabolic disease caused by dysfunction of the ornithine aminotransferase (OAT) gene, leading to ornithine accumulation and a complex metabolic imbalance. This causes retinal degeneration that ultimately evolve to blindness. However, the mechanisms of this degeneration remain unknown. Here, we have conducted untargeted metabolomic analysis in patient-derived induced pluripotent stem cells and their isogenic counterparts. Mutant cells show altered levels of ornithine-related metabolites, including low creatine, proline and glutamate, and elevated arginine and citrulline. The untargeted metabolomics approach revealed changes in the urea cycle and polyamine synthesis pathways with a significant intracellular accumulation of gamma-aminobutyric acid (GABA). Hence, we propose GABA as a key player in the disease pathogenicity, potentially affecting neuronal function in the eye.

biochemistry↗

bioRxiv2023

CRISPR/Cas9 DNA synthesis disruption as a tool to control cell proliferation in vitro and in vivo

Engineered cells hold great promise for regenerative medicine and gene therapy. However, living cell products entail a fundamental biological risk of unwanted growth. Here, we describe a novel metabolic safety system to control cell proliferation without added genetic elements. We inactivated a key enzyme for nucleotide metabolism, TYMS, in several cell lines, thus obtaining cells that proliferate only when supplemented with exogenous thymidine but fail to replicate in its absence. Under supplementation, TYMS-/- pluripotent stem cells proliferate normally, produce teratomas and differentiate into potentially therapeutic cell types such as pancreatic beta cells. After differentiation, the postmitotic cells do not require thymidine to function, as seen by prolonged in vivo production of human insulin in implanted mice. Hence, this method allows robust cell culture and manufacture while mitigating the risk of uncontrolled growth of transplanted cells. One Sentence Summary: Genetic disruption of DNA synthesis prevents unwanted proliferation in cell therapies without affecting cell function.

bioengineering↗