Little is known concerning promoter structure in the filarial parasites. Recently, transient transfection methods have been developed for the human filarial parasite Brugia malayi. These methods have been employed to localize the promoter for the 70 kDa heat shock protein (BmHSP70) to a region extending 394 nt upstream from the initiating codon of the BmHSP70 open reading frame. Replacement mutagenesis was used to define the elements necessary for BmHSP70 promoter activity in detail. Four domains, ranging in size from six to 22 nucleotides, were found to be necessary for full promoter activity. The two most distal domains encoded a binding site for the heat shock transcription factor and a putative binding site for the GAGA transcription factor, motifs that are found in many other HSP70 promoters. However, none of the essential domains contained sequences typical of cis elements that are usually found in the core domain of a eukaryotic promoter. The largest essential domain was located at positions -53 to -32, and included the splice leader addition site. These data suggest that the regulatory domains of the BmHSP70 promoter were similar to those found in other eukaryotes, but that the core promoter domain exhibited features that appeared to be distinct from those found in most other well-characterized eukaryotic promoters. An analysis of two additional promoters of B. malayi highly transcribed genes suggests that they also lack features commonly found in most eukaryotic core promoters, suggesting that the unique features of the BmHSP70 core promoter are not confined to this gene. © 2005 Elsevier Ltd. All rights reserved.