Heat shock proteins (Hsps) and other molecular chaperones perform diverse physiological roles. One is to facilitate, in part, organismal thermotolerance, of which the functional consequences depend on Hsp70 concentration and developmental stage in Drosophila melanogaster. To test whether an Hsp70-thermotolerance relationship is a general phenomenon within Drosophila, I assayed Hsp70 concentration at a range of temperatures in intact larvae and adults of three species, D. melanogaster, D. simulans, and D. mojavensis, and compared those results to the increase in survival to heat shock that occurs after an Hsp70 inducing pretreatment. Larvae of D. melanogaster and D. simulans responded similarly to heat; they expressed Hsp70 maximally at 36-37 degrees C, and their tolerance of 1 h heat shocks increased by 1.5-2 degrees C. By contrast, D. mojavensis, which tolerates higher temperatures than do D. melanogaster and D. simulans, expressed Hsp70 only at higher temperatures, although the 36 degrees C pretreatment still increased thermotolerance. Critically, the temperature that maximally induced Hsp70 was a poor inducer of thermotolerance in D. mojavensis and may have harmed larvae. Results for Drosophila adults, which tolerated heat poorly compared to larvae, likewise suggest that a close link between peak Hsp70 expression and maximal induction of thermotolerance is a feature of D. melanogaster, and not of the other species. Neither D. simulans nor D. mojavensis adults increased tolerance after exposure to the temperatures that maximally induced Hsp70.