Depletion of the isocitrate dehydrogenase 3 subunit IDHA-1 licenses germ cell-to-neuron direct reprogramming

Direct reprogramming (DR) converts one differentiated cell identity into another and is a promising strategy for regenerative approaches when cells are lost because of injury or disease. Transcription factors (TFs) that specify a given cell type can drive DR, but inhibitory mechanisms restrict this conversion in most cell types. To identify such barriers in vivo , we use Caenorhabditis elegans and the zinc finger TF CHE-1, which is required to specify the glutamatergic taste neuron fate of ASE neurons. Ectopic CHE-1 expression can induce DR of different cell types to ASE neurons upon RNAi-mediated depletion of barrier genes. Here we characterize the α subunit of the mitochondrial isocitrate dehydrogenase 3 complex (IDH3), IDHA-1, as a barrier to DR of germ cells into neurons. RNAi against idha-1 produced consistent ASE fate reporter expression, accompanied by morphological changes to neuron-like structures in the germline upon ectopic CHE-1 expression. Using different neuronal gene expression reporters, single-molecule FISH, and antibody staining, we confirm that this germ cell conversion (GeCo) produces neuron-like cells. We found that loss of IDH3 activity causes metabolic perturbations that result in a variety of direct and indirect effects on DR of germ cells. One arm implicates the hypoxia-inducible factor HIF-1, which promotes GeCo. Its loss causes a significant decrease, while the vhl-1 mutant background, in which HIF-1 protein is stabilized, leads to enhanced GeCo. Another arm implicates epigenetic changes leading to a detectable loss of the repressive histone marks H3K27me3 and H3K9me3 upon idha-1 depletion. Furthermore, stable-isotope-resolved metabolomics shows strong citrate accumulation without a noteworthy drop in α-Ketoglutarate (αKG) levels, indicating that compensatory pathways maintain αKG levels. Genetics and metabolomics analyses confirmed that Glutamate anaplerosis contributes to compensating for the loss of IDHA-1. Additionally, we found that depletion of glucose transporter FDGT-2 nearly doubles GeCo efficiency. Overall, our findings define the TCA cycle as an in vivo safeguard of germ cell identity and show that multiple metabolic inputs converge to keep germ cells refractory to TF-induced DR to neuronal cells.