We have established the nematode, Caenorhabditis elegans , as a system for studying peroxisome biogenesis and modeling the associated human peroxisomal biogenesis disorders (PBDs). Peroxisomes are present in virtually all eukaryotic cells where they carry out a variety of essential metabolic functions including many components of lipid metabolism (such as beta-oxidation and ether phospholipid synthesis). The peroxisome is also involved in amino acid and purine metabolism. Peroxin proteins (called PEX genes in humans, PRX in C. elegans ), are required for either the import of enzymes into the peroxisomal matrix, or are necessary for the division and growth of peroxisomes. Mutations in peroxin genes cause the devastating PBDs, such as neonatal adrenoleukodystrophy and Zellweger syndrome. The genome of C. elegans encodes homologs for 10 of the 14 mammalian peroxins. We used dsRNA interference to determine the biological role of the peroxisome in nematodes. We targeted 5 nematode peroxin homologs with the highest degree of homology to PEX 5, PEX6, PEX12, PEX13 and PEX19. Using a GFP reporter targeted to the peroxisome, we show that peroxisome biogenesis is impaired in
prx-5(RNAi) worms. In addition, inactivation of each of these 5 peroxin genes by RNAi results in an early larval arrest at the L1 stage. This implies that peroxisome function is necessary for the normal development of C. elegans . The L1 arrested worms are active and survive for up to 5 days, which is strikingly similar to starvation-arrested L1 worms. We hypothesize that a by-product of peroxisome metabolism may be necessary to signal the advance to post-embryonic development. Importantly, the L1 arrest phenotype suggests that genetic suppressor analysis might identify novel ways to bypass toxicity related to the PBDs. Global developmental delay is a major hallmark of the PBDs, and is thought to be at least in part caused by defects in neuroblast migration during development. We have noticed no change in neuronal position in either
mec-4::GFP;
prx-5(RNAi) (touch neurons) or
unc129:GFP;
prx-5(RNAi) (DA and DB motor neurons) worms. In order to examine post-embryonic neuroblast migration in RNAi treated nematodes, we are now examining the Q neuroblasts which normally begin migration within the first hour after hatching. To obtain a better understanding of the biochemical functions carried out by the nematode peroxisome, we took an in silico approach to identify all C. elegans proteins which contain a type I peroxisomal targeting signal (S/A/C-K/R/H-L/M/I). We identified more than 30 predicted open reading frames homologous to either human or yeast peroxisomal enzymes, including enzymes that participate in beta-oxidation and branched-chain fatty acid oxidation. We have also identified 15 novel PTS1-containing ORFs and are currently confirming the subcellular localization of these proteins with GFP fusion constructs.