Peroxisomes are subcellular organelles with important functions in lipid metabolism that are found in virtually all eucaryotic cells. The peroxisomal membrane contains a number of integral and peripheral membrane proteins involved in the import of peroxisomal matrix proteins and the transport of metabolites across the membrane. The most abundant peroxisomal membrane protein (Pmp) in rat peroxisomes is Pmp22, a 22 kDa protein of unknown function that is encoded by the Pxmp2 gene. To investigate the function of the Pxmp2 gene, we have initiated mouse knockout studies. The expression level of the Pxmp2 mRNA in mice was investigated by Northern blot analysis. Pxmp2 RNA was shown to be differentially expressed with highest expression levels in liver, kidney and in heart tissue. Comparison with other peroxisomal marker genes revealed that the expression of Pxmp2, Pmp70 (Pxmp1) and catalase was regulated independently. Using 5' and 3' RACE we have cloned the full-length cDNA of murine Pxmp2 which comprises 863 nucleotides and have isolated a genomic clone containing the entire murine Pxmp2. We have analyzed the complete intron/exon structure of the Pxmp2 gene which contains five exons spanning about 11 kb on the genomic clone. All intron/exon splice junctions conform to the GT/AG rule. Sequence analysis of the Pxmp2 5' flanking region revealed that it was devoid of a TATA box, but characteristic promoter elements were identified within 250 base pairs upstream of the transcriptional start site. Using a mouse/hamster radiation hybrid panel, Pxmp2 was localized on mouse chromosome 5 at 59 cM.