A geothermal amoeba sets a new upper temperature limit for eukaryotes

The study of temperature limits has transformed our understanding of life's boundaries but has focused on bacteria and archaea. For decades after their discovery, no eukaryote was shown to replicate above 60°C. We isolated a geothermal amoeba, Incendiamoeba cascadensis, that divides at 63°C and is motile up to 64°C. We identified an enrichment of genes related to proteostasis and environmental sensing relative to mesophilic amoebae. Comparative RNA sequencing of I. cascadensis revealed upregulation of pathways related to proteostasis, DNA repair, and membrane trafficking at high temperatures. Predicted proteins of I. cascadensis have distinct biophysical properties, including enrichment of positively charged surface residues similar to bacterial and archaeal hyperthermophiles, which suggests a convergent mechanism for protein stability under temperature stress. Together, our findings expand our understanding of where and how life can persist.