Pyruvate dehydrogenase kinase 4 (PDK4) is a key regulatory enzyme involved in switching the energy source from glucose to fatty acids in response to physiological conditions. Transcription of the PDK4 gene is activated by fasting or by the administration of a PPARalpha ligand in a tissue-specific manner. Here, we show that the two mechanisms are independent, and that ERRalpha is directly involved in PPARalpha-independent transcriptional activation of the PDK4 gene with PGC-1alpha as a specific partner. This conclusion is based on the following evidence. First, detailed mutation analyses of the cloned PDK4 gene promoter sequence identified a possible ERRalpha-binding motif as the PGC-1alpha responsive element. Second, overexpression of ERRalpha by cotransfection enhanced, and the knockout of it by shRNAs diminished, PGC-1alpha-dependent activation. Third, specific binding of ERRalpha to the identified PGC-1alpha responsive sequence was confirmed by the electrophoresis mobility shift assay. Finally, cell-type-specific responsiveness to PGC-1alpha was observed and this could be explained by differences in the expression levels of ERRalpha, however, ectopic expression of ERRalpha in poorly responsive cells did not restore PGC-1alpha responsiveness, indicating that ERRalpha is necessary, but not sufficient for the response.