Hsp105alpha and Hsp105beta are mammalian members of the Hsp105/110 family, a divergent subgroup of the Hsp70 family. Hsp105alpha is expressed constitutively and induced by various forms of stress, whereas Hsp105beta is an alternatively spliced form of Hsp105alpha that is expressed specifically during mild heat shock. In a report, it was shown that Hsp105alpha and Hsp105beta localize to the cytoplasm and of nucleus of cells, respectively, and that Hsp105beta, but not Hsp105alpha, induces the expression of Hsp70 in mammalian cells. Here, we examined the mechanism by which Hsp105beta induces the expression of Hsp70. Using a series of deletion mutants of Hsp105beta, it was revealed that the region between amino acids 642 and 662 of Hsp105beta is necessary for the activation of the hsp70 promoter by Hsp105beta. Furthermore, it was shown that signal transducer and activator of transcription (STAT)-3 bound to the sequence of the hsp70 promoter between -206 and -187 bp, and that mutations of this sequence abrogated the activation of the hsp70 promoter by Hsp105beta. In addition, overexpression of Hsp105beta stimulated the phosphorylation of STAT3 at Tyr705 and its translocation to the nucleus. Downregulation of STAT3 expression resulted in reduction of the activation of the hsp70 promoter by Hsp105beta. Furthermore, downregulation of Hsp105beta reduced the expression of Hsp70 in heat-shocked cells. On the basis of these findings, it is suggested that Hsp105beta induces Hsp70 expression markedly through the STAT3 pathway in heat-shocked cells. This may represent the mechanism that connects the heat shock protein and STAT families for cell defense against deleterious stress.