The article reviews the role of transfer RNA (tRNA) modifications in cellular thermotolerance, particularly in thermophilic organisms. tRNA modifications are crucial for decoding the genetic code and stabilizing tRNA structures. In thermophiles, specific tRNA modifications are dynamically regulated in response to varying growth temperatures, enhancing their thermal stability and resistance to degradation. Key modifications include 2′-O-methylation (Nm), pseudouridine (Ψ), 5-methyluridine (m5U), 4-thiouridine (s4U), 7-methylguanosine (m7G), and 3-(3-amino-3-carboxypropyl)uridine (ac3U). These modifications stabilize tRNA structures and optimize protein synthesis at different temperatures. The article also discusses the reversible tRNA phosphorylation (U^P) and its role in maintaining tRNA stability during temperature changes. Additionally, it highlights the importance of tRNA modifications in human diseases and their potential as therapeutic targets. The review emphasizes the significance of tRNA modifications in the adaptation of thermophiles to extreme environments and the ongoing research efforts to understand and utilize these modifications.The article reviews the role of transfer RNA (tRNA) modifications in cellular thermotolerance, particularly in thermophilic organisms. tRNA modifications are crucial for decoding the genetic code and stabilizing tRNA structures. In thermophiles, specific tRNA modifications are dynamically regulated in response to varying growth temperatures, enhancing their thermal stability and resistance to degradation. Key modifications include 2′-O-methylation (Nm), pseudouridine (Ψ), 5-methyluridine (m5U), 4-thiouridine (s4U), 7-methylguanosine (m7G), and 3-(3-amino-3-carboxypropyl)uridine (ac3U). These modifications stabilize tRNA structures and optimize protein synthesis at different temperatures. The article also discusses the reversible tRNA phosphorylation (U^P) and its role in maintaining tRNA stability during temperature changes. Additionally, it highlights the importance of tRNA modifications in human diseases and their potential as therapeutic targets. The review emphasizes the significance of tRNA modifications in the adaptation of thermophiles to extreme environments and the ongoing research efforts to understand and utilize these modifications.