Peroxisomes are single-membrane-bound organelles that have important functions in cell metabolism. They are almost ubiquitous in eukaryotic cells and biochemical findings over the past 10 years have shown that they are involved in cell respiration, and also play a significant role in lipid metabolism (for reviews, see [ l , 21). In recent years, studies have been initiated in a variety o f systems aimed at defining the mechanisms involved in peroxisome biogenesis. These include investigations in yeasts and plants, as well as in higher eukaryotic cells. Results from these studies indicate that the biogenesis of peroxisomes is a Peroxisomes contain no DNA, therefore, all s are encoded in the nucleus. It is now well established that the development of peroxisomes requires the post-translational import of proteins that are synthesized o n cytosolic ribosomes. This is true o f both pcroxisomal matrix and membrane proteins. New peroxisomes arc thought to form by division of preexisting peroxisomes. In this respect, the biogenesis of peroxisomes resembles that of mitochondria. Mitochondria contain DNA but this genomc encodes less than 10% of mitochondrial proteins. Thc majority of mitochondrial proteins arc nuclearly encoded, many as larger precursors, and enter the organelle posttranslationally. This step is often accompanied by proteolytic cleavage of a leader peptide (for a review, see [3]) . Most peroxisomal proteins are synthesized at their mature size and lack cleavable topogenic sequences. Thus, much interest has been generated as to the type of signal that acts to direct peroxisomal proteins to peroxisomes and whether the mechanism of import resembles or differs from that in other organelles. In mitochondrial studies, these questions have becn addressed by establishing both in vitro and in vivo import assays t o follow the fate of individual proteins. Similar types of experiments have been initiated with peroxisdmal proteins to define and compare the mechanisms involved in transport of these proteins to peroxisomes. The post-translational import of peroxisomal proteins was first accomplished in vitro with rat liver peroxisomes [4]. Using a modification of this system, hydrolysis of ATP was found to be a requirement for the import of acyl-CoA oxidase (AOx; the first enzyme in the peroxisomal /?-oxidation cycle) [5]. A potential across the peroxisomal membrane was not necessary for this import to occur. An import assay was established in vitro with peroxisomes from the yeast Cundidu tropiculis, for which cDNAs were available [ 61. Peroxisomes