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Drought causes approximately two-thirds of crop and yield loss worldwide. To sustain future generations, there is a need to develop robust crops with enhanced water use efficiency. Resurrection plants are naturally resilient and tolerate up to 95% water loss with the ability to revive upon watering. Stress is genetically encoded and resilient species may garner tolerance by tightly regulating the expression of stress-related genes. MicroRNAs (miRNAs) post-transcriptionally regulate development and other stress response processes in eukaryotes. However, their role in resurrection plant desiccation tolerance is poorly understood. In this study, small RNA sequencing and miRNA expression profiling was conducted using Tripogon loliiformis plants subjected to extreme water deficit conditions. Differentially expressed miRNA profiles, target mRNAs, and their regulatory processes were elucidated. Gene ontology enrichment analysis revealed that development, stress response, and regulation of programmed cell death biological processes; Oxidoreductase and hydrolyase molecular activities; and SPL, MYB, and WRKY transcription factors were targeted by miRNAs during dehydration stress, indicating the indispensable regulatory role of miRNAs in desiccation tolerance. This study provides insights into the molecular mechanisms of desiccation tolerance in the resurrection plant T. loliiformis. This information will be useful in devising strategies for crop improvement on enhanced drought tolerance and water use efficiency. Full article