Browsing by Author "Ratzel, Sarah E."

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    A pex1 missense mutation improves peroxisome function in a subset of Arabidopsis pex6 mutants without restoring PEX5 recycling
    (National Academy of Sciences, 2018) Gonzalez, Kim L.; Ratzel, Sarah E.; Burks, Kendall H.; Danan, Charles H.; Wages, Jeanne M.; Zolman, Bethany K.; Bartel, Bonnie
    Peroxisomes are eukaryotic organelles critical for plant and human development because they house essential metabolic functions, such as fatty acid β-oxidation. The interacting ATPases PEX1 and PEX6 contribute to peroxisome function by recycling PEX5, a cytosolic receptor needed to import proteins targeted to the peroxisomal matrix. Arabidopsis pex6 mutants exhibit low PEX5 levels and defects in peroxisomal matrix protein import, oil body utilization, peroxisomal metabolism, and seedling growth. These defects are hypothesized to stem from impaired PEX5 retrotranslocation leading to PEX5 polyubiquitination and consequent degradation of PEX5 via the proteasome or of the entire organelle via autophagy. We recovered a pex1 missense mutation in a screen for second-site suppressors that restore growth to the pex6-1 mutant. Surprisingly, this pex1-1 mutation ameliorated the metabolic and physiological defects of pex6-1 without restoring PEX5 levels. Similarly, preventing autophagy by introducing an atg7-null allele partially rescued pex6-1 physiological defects without restoring PEX5 levels. atg7 synergistically improved matrix protein import in pex1-1 pex6-1, implying that pex1-1 improves peroxisome function in pex6-1 without impeding autophagy of peroxisomes (i.e., pexophagy). pex1-1 differentially improved peroxisome function in various pex6 alleles but worsened the physiological and molecular defects of a pex26 mutant, which is defective in the tether anchoring the PEX1–PEX6 hexamer to the peroxisome. Our results support the hypothesis that, beyond PEX5 recycling, PEX1 and PEX6 have additional functions in peroxisome homeostasis and perhaps in oil body utilization.
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    A viable Arabidopsis pex13 missense allele confers severe peroxisomal defects and decreases PEX5 association with peroxisomes
    (Springer, 2014) Woodward, Andrew W.; Fleming, Wendell A.; Burkhart, Sarah E.; Ratzel, Sarah E.; Bjornson, Marta; Bartel, Bonnie
    Peroxisomes are organelles that catabolize fatty acids and compartmentalize other oxidative metabolic processes in eukaryotes. Using a forward-genetic screen designed to recover severe peroxisome-defective mutants, we isolated a viable allele of the peroxisome biogenesis gene PEX13 with striking peroxisomal defects. The pex13-4 mutant requires an exogenous source of fixed carbon for pre-photosynthetic development and is resistant to the protoauxin indole-3-butyric acid. Delivery of peroxisome-targeted matrix proteins depends on the PEX5 receptor docking with PEX13 at the peroxisomal membrane, and we found severely reduced import of matrix proteins and less organelle-associated PEX5 in pex13-4 seedlings. Moreover, pex13-4 physiological and molecular defects were partially ameliorated when PEX5 was overexpressed, suggesting that PEX5 docking is partially compromised in this mutant and can be improved by increasing PEX5 levels. Because previously described Arabidopsis pex13 alleles either are lethal or confer only subtle defects, the pex13-4 mutant provides valuable insight into plant peroxisome receptor docking and matrix protein import.
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    Influence of Peroxisomal Import and Receptor Recycling of Peroxisomal Function
    (2011) Ratzel, Sarah E.; Bartel, Bonnie
    Peroxisomes compartmentalize a variety of important metabolic reactions including fatty acid β-oxidation and the related process of IBA β-oxidation. Peroxisomal proteins are encoded by nuclear genes and must be post-translationally imported. A dynamic import process is vital for proper matrix protein localization and is dependent on the family of peroxin (PEX) proteins. The delivery and peroxisomal import of cargo from a loaded receptor, PEX5 or PEX7, is carried out by the early-acting peroxins, including PEX13 and PEX14, and receptor recycling is carried out by the late-acting peroxins, including PEX4 and PEX6. In this thesis, I describe the use of double mutant analysis to differentiate early-acting and late-acting pex mutants by phenotypic and molecular analysis. I found that double mutants made with two early-acting or two late-acting pex mutants showed enhanced phenotypes in β-oxidation and import defects. In contrast, defects of double mutants made with a weak early-acting mutant and a late-acting mutant were suppressed. Additionally, I found that receptor localization is central to proper peroxisomal function. My results suggest that when the receptor is not removed from the peroxisome, stabilized peroxisomal pores may be formed, perhaps impairing peroxisomal function due to leaching of peroxisomal contents. Together my data suggest that balance between import and receptor recycling is fundamental for peroxisomal function. In humans, peroxisomal biogenesis disorders are most often caused by defects in late-acting peroxins. Peroxisomal defects occur in plants and humans as a result of the same lesions in PEX proteins. The understanding of how these late-acting defects can be ameliorated in plants, may inspire new approaches to human therapeutics.
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    Matrix proteins are inefficiently imported into Arabidopsis peroxisomes lacking the receptor-docking peroxin PEX14
    (Springer, 2011) Monroe-Augustus, Melanie; Ramón, Naxhiely Martínez; Ratzel, Sarah E.; Lingard, Matthew J.; Christensen, Sarah E.; Murali, Chaya; Bartel, Bonnie
    Mutations in peroxisome biogenesis proteins (peroxins) can lead to developmental deficiencies in various eukaryotes. PEX14 and PEX13 are peroxins involved in docking cargo-receptor complexes at the peroxisomal membrane, thus aiding in the transport of the cargo into the peroxisomal matrix. Genetic screens have revealed numerous Arabidopsis thaliana peroxins acting in peroxisomal matrix protein import; the viable alleles isolated through these screens are generally partial loss-of-function alleles, whereas null mutations that disrupt delivery of matrix proteins to peroxisomes can confer embryonic lethality. In this study, we used forward and reverse genetics in Arabidopsis to isolate four pex14 alleles. We found that all four alleles conferred reduced PEX14 mRNA levels and displayed physiological and molecular defects suggesting reduced but not abolished peroxisomal matrix protein import. The least severe pex14 allele, pex14-3, accumulated low levels of a C-terminally truncated PEX14 product that retained partial function. Surprisingly, even the severe pex14-2 allele, which lacked detectable PEX14 mRNA and PEX14 protein, was viable, fertile, and displayed residual peroxisome matrix protein import. As pex14 plants matured, import improved. Together, our data indicate that PEX14 facilitates, but is not essential for peroxisomal matrix protein import in plants.