44001:

An acid matrix is required for receptor-mediated protein import into plant peroxisomes.

Authors:	Brickner, Donna, G.(A)Olsen, Laura, J.(A)
Affiliations:	(A): Department of Biology, University of Michigan
Presenter:	Olsen, Laura J., ljo@umich.edu

Peroxisomes are vital organelles responsible for numerous metabolic functions in plant and animal cells. Peroxisomal matrix proteins are nuclear-encoded, synthesized on cytosolic ribosomes, and posttranslationally targeted to the organelle matrix via a carboxyl-terminal tripeptide known as a Peroxisomal Targeting Signal type 1 (PTS1). To study protein import into glyoxysomes, a class of peroxisomes abundant in oilseed plants, we cloned the PTS1 receptor (PEX5) from Arabidopsis. AtPex5p is an 84 kD protein that is 28% identical to human Pex5p. The sequence identity is most striking in the carboxyl-terminal tetratricopeptide repeat (TPR) domain, a protein-protein interaction domain. Southern blots indicate there is one copy of PEX5 in the Arabidopsis genome. PEX5 is expressed at low levels in all tissues tested. Anti-AtPex5p antibodies specifically immunoprecipitated PTS1 proteins. It is thought that the PTS1 signal binds to the TPR domain of Pex5p in the cytosol. The exact subcellular location of the dissociation of the receptor and PTS1 protein is unknown. We are interested in studying this pathway of peroxisome biogenesis in plants. The pH of membrane-bound compartments is an important biochemical cue that provides directionality or asymmetry for many protein trafficking events. We have directly and indirectly examined the importance of pH for protein import into plant peroxisomes. We propose that the relatively low pH of the peroxisomal matrix may facilitate the dissociation of a PTS1 protein and its receptor complex, similar to the release of receptor-ligand complexes during endocytosis. To test this ‘acid matrix hypothesis’ we used a combination of approaches. First, when ionophores or weak bases were added to in vitro protein import reactions the level of import was significantly reduced. Second, we have shown that ionophores and weak bases deacidify the matrix of peroxisomes. These results taken together indicate that the slightly acidic pH of the peroxisomal matrix is required for proper import of peroxisomal proteins. In addition, we found that the glyoxysomal protein isocitrate lyase readily binds to AtPex5p, the PTS1 receptor, at pH 7, typical of cytosol, but less well at pH 6, the proposed peroxisomal matrix pH. Finally, we provide some evidence that an F1FO-type ATPase and/or a V-class ATPase may be responsible for the acidification of the peroxisomal matrix. This research is funded by the USDA.