•The Arabidopsis heat-intolerant 2 (hit2) mutant was isolated on the basis of its impaired ability to withstand moderate heat stress (37°C). Determination of the genetic mutation that underlies the hit2 thermosensitive phenotype allowed better understanding of the mechanisms by which plants cope with heat stress.•Genetic analysis revealed that hit2 is a single recessive mutation. Map-based cloning was used to identify the hit2 locus. The response of hit2 to other types of heat stress was also investigated to characterize the protective role of HIT2.•hit2 was defective in basal but not in acquired thermotolerance. hit2 was sensitive to methyl viologen-induced oxidative stress, and the survival of hit2 seedlings in response to heat stress was affected by light conditions. The mutated locus was located at the EXPORTIN1A (XPO1A) gene, which encodes a nuclear transport receptor. Two T-DNA insertion lines, xpo1a-1 and xpo1a-3, exhibited the same phenotypes as hit2.•The results provide evidence that Arabidopsis XPO1A is dispensable for normal plant growth and development but is essential for thermotolerance, in part by mediating the protection of plants against heat-induced oxidative stress.