ERp29 is a soluble protein localized in the endoplasmic reticulum (ER) of eukaryotic cells, which is conserved in all mammalian species. The N-terminal domain of ERp29 displays sequence and structural similarity to the protein disulfide isomerase despite the lack of the characteristic double cysteine motif. Although the exact function of ERp29 is not yet known, it was hypothesized that it may facilitate folding and/or export of secretory proteins in/from the ER. ERp29 is induced by ER stress, i.e. accumulation of unfolded proteins in the ER. To gain an insight into the mechanisms regulating ERp29 expression we have cloned and characterized the rat ERp29 gene and studied in details its distribution in human tissues. Comparison with the murine and human genes and phylogenetic analysis demonstrated common origin and close ortholog relationships of these genes. Additionally, we have cloned approximately 3 kb of the 5'-flanking region of the ERp29 gene and functionally characterized its promoter. Such characteristics of the promoter as GC-rich sequence, absence of TATA-box, multiple transcription start sites taken together with the ubiquitous gene expression, reaching maximum levels in the specialized secretory tissues, indicate that ERp29 belongs to the group of the constitutively expressed housekeeping genes. A 337 bp fragment of the 5' flank was identified as a core promoter sufficient for the transcriptional activation of the gene. Gel mobility shift assay indicated interaction of the predicted GC and E box elements within the core promoter with Sp1/Sp3 and USF1/USF2 transcription factors, respectively, suggesting their key role in the basal expression of the gene.