Plant Molecular Biology

, Volume 86, Issue 1–2, pp 201–214 | Cite as

A viable Arabidopsis pex13 missense allele confers severe peroxisomal defects and decreases PEX5 association with peroxisomes

  • Andrew W. Woodward
  • Wendell A. Fleming
  • Sarah E. Burkhart
  • Sarah E. Ratzel
  • Marta Bjornson
  • Bonnie Bartel


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.


Peroxisome Organelle biogenesis Subcellular targeting 



We thank Steven Smith (University of Western Australia) for the PMDH2 antibody, Monique Gill for assistance with the mutant screen, and Kim Gonzalez, Yun-Ting Kao, Mauro Rinaldi, and Pierce Young for critical comments on the manuscript. This research was supported by the National Science Foundation (MCB-1244182) and the Robert A. Welch Foundation (C-1309). Confocal microscopy was performed on equipment obtained through a Shared Instrumentation Grant from the National Institutes of Health (S10RR026399). A. W. W. was partially supported by a UMHB Faculty Development Grant, and M. B. was partially supported by a Howard Hughes Medical Institute Professors Grant (52005717 to B. B.).


  1. Adham AR, Zolman BK, Millius A, Bartel B (2005) Mutations in Arabidopsis acyl-CoA oxidase genes reveal distinct and overlapping roles in β-oxidation. Plant J 41:859–874PubMedCrossRefGoogle Scholar
  2. Ausubel F, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (1999) Current protocols in molecular biology. Greene Publishing Associates and Wiley-Interscience, New YorkGoogle Scholar
  3. Azevedo JE, Schliebs W (2006) Pex14p, more than just a docking protein. Biochim Biophys Acta 1763:1574–1584PubMedCrossRefGoogle Scholar
  4. Boisson-Dernier A, Frietsch S, Kim T-H, Dizon MB, Schroeder JI (2008) The peroxin loss-of-function mutation abstinence by mutual consent disrupts recognition between male and female gametophytes. Curr Biol 18:63–68PubMedCentralPubMedCrossRefGoogle Scholar
  5. Burkhart SE, Lingard MJ, Bartel B (2013) Genetic dissection of peroxisome-associated matrix protein degradation in Arabidopsis thaliana. Genetics 193:125–141PubMedCentralPubMedCrossRefGoogle Scholar
  6. Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743PubMedCrossRefGoogle Scholar
  7. Curtis MD, Grossniklaus U (2003) A gateway cloning vector set for high-throughput functional analysis of genes in planta. Plant Physiol 133:462–469PubMedCentralPubMedCrossRefGoogle Scholar
  8. De Rybel B, Audenaert D, Xuan W, Overvoorde P, Strader LC, Kepinski S, Hoye R, Brisbois R, Parizot B, Vanneste S, Liu X, Gilday A, Graham IA, Nguyen L, Jansen L, Njo MF, Inze D, Bartel B, Beeckman T (2012) A role for the root cap in root branching revealed by the non-auxin probe naxillin. Nat Chem Biol 8:798–805PubMedCentralPubMedCrossRefGoogle Scholar
  9. Douangamath A, Filipp FV, Klein AT, Barnett P, Zou P, Voorn-Brouwer T, Vega MC, Mayans OM, Sattler M, Distel B, Wilmanns M (2002) Topography for independent binding of alpha-helical and PPII-helical ligands to a peroxisomal SH3 domain. Mol Cell 10:1007–1017PubMedCrossRefGoogle Scholar
  10. Eastmond PJ (2006) SUGAR-DEPENDENT1 encodes a patatin domain triacylglycerol lipase that initiates storage oil breakdown in germinating Arabidopsis seeds. Plant Cell 18:665–675PubMedCentralPubMedCrossRefGoogle Scholar
  11. Eastmond PJ (2007) MONODEHYROASCORBATE REDUCTASE4 is required for seed storage oil hydrolysis and postgerminative growth in Arabidopsis. Plant Cell 19:1376–1387PubMedCentralPubMedCrossRefGoogle Scholar
  12. Elgersma Y, van den Berg M, Tabak HF, Distel B (1993) An efficient positive selection procedure for the isolation of peroxisomal import and peroxisome assembly mutants of Saccharomyces cerevisiae. Genetics 135:731–740PubMedCentralPubMedGoogle Scholar
  13. Girzalsky W, Rehling P, Stein K, Kipper J, Blank L, Kunau W-H, Erdmann R (1999) Involvement of Pex13p in Pex14p localization and peroxisomal targeting signal 2-dependent protein import into peroxisomes. J Cell Biol 144:1151–1162PubMedCentralPubMedCrossRefGoogle Scholar
  14. Glover JR, Andrews DW, Rachubinski RA (1994) Saccharomyces cerevisiae peroxisomal thiolase is imported as a dimer. Proc Natl Acad Sci USA 91:10541–10545PubMedCentralPubMedCrossRefGoogle Scholar
  15. Goto S, Mano S, Nakamori C, Nishimura M (2011) Arabidopsis ABERRANT PEROXISOME MORPHOLOGY9 is a peroxin that recruits the PEX1–PEX6 complex to peroxisomes. Plant Cell 23:1573–1587PubMedCentralPubMedCrossRefGoogle Scholar
  16. Gould SJ, McCollum D, Spong AP, Heyman JA, Subramani S (1992) Development of the yeast Pichia pastoris as a model organism for a genetic and molecular analysis of peroxisome assembly. Yeast 8:613–628PubMedCrossRefGoogle Scholar
  17. Graham IA (2008) Seed storage oil mobilization. Annu Rev Plant Biol 59:115–142PubMedCrossRefGoogle Scholar
  18. Haughn GW, Somerville C (1986) Sulfonylurea-resistant mutants of Arabidopsis thaliana. Mol Gen Genet 204:430–434CrossRefGoogle Scholar
  19. Hayashi M, Toriyama K, Kondo M, Nishimura M (1998) 2,4-Dichlorophenoxybutyric acid-resistant mutants of Arabidopsis have defects in glyoxysomal fatty acid β-oxidation. Plant Cell 10:183–195PubMedCentralPubMedGoogle Scholar
  20. Hayashi M, Nito K, Toriyama-Kato K, Kondo M, Yamaya T, Nishimura M (2000) AtPex14p maintains peroxisomal functions by determining protein targeting to three kinds of plant peroxisomes. EMBO J 19:5701–5710PubMedCentralPubMedCrossRefGoogle Scholar
  21. Helm M, Lück C, Prestele J, Hierl G, Huesgen PF, Frohlich T, Arnold GJ, Adamska I, Görg A, Lottspeich F, Gietl C (2007) Dual specificities of the glyoxysomal/peroxisomal processing protease DEG15 in higher plants. Proc Natl Acad Sci USA 104:11501–11506PubMedCentralPubMedCrossRefGoogle Scholar
  22. Hu J, Baker A, Bartel B, Linka N, Mullen RT, Reumann S, Zolman BK (2012) Plant peroxisomes: biogenesis and function. Plant Cell 24:2279–2303PubMedCentralPubMedCrossRefGoogle Scholar
  23. Islinger M, Grille S, Fahimi HD, Schrader M (2012) The peroxisome: an update on mysteries. Histochem Cell Biol 137:547–574PubMedCrossRefGoogle Scholar
  24. Koncz C, Schell J (1986) The promoter of the TL-DNA gene 5 controls the tissue-specific expression of chimaeric genes carried by a novel type of Agrobacterium binary vector. Mol Gen Genet 204:383–396CrossRefGoogle Scholar
  25. Krause C, Rosewich H, Woehler A, Gartner J (2013) Functional analysis of PEX13 mutation in a Zellweger syndrome spectrum patient reveals novel homooligomerization of PEX13 and its role in human peroxisome biogenesis. Hum Mol Genet 22:3844–3857PubMedCrossRefGoogle Scholar
  26. Lee MS, Mullen RT, Trelease RN (1997) Oilseed isocitrate lyases lacking their essential type 1 peroxisomal targeting signal are piggybacked to glyoxysomes. Plant Cell 9:185–197PubMedCentralPubMedCrossRefGoogle Scholar
  27. León J (2013) Role of plant peroxisomes in the production of jasmonic acid-based signals. Subcell Biochem 69:299–313PubMedCrossRefGoogle Scholar
  28. Li XR, Li HJ, Yuan L, Liu M, Shi DQ, Liu J, Yang WC (2014) Arabidopsis DAYU/ABERRANT PEROXISOME MORPHOLOGY9 is a key regulator of peroxisome biogenesis and plays critical roles during pollen maturation and germination in planta. Plant Cell 26:619–635PubMedCrossRefGoogle Scholar
  29. Lingard MJ, Bartel B (2009) Arabidopsis LON2 is necessary for peroxisomal function and sustained matrix protein import. Plant Physiol 151:1354–1365PubMedCentralPubMedCrossRefGoogle Scholar
  30. Lingard MJ, Monroe-Augustus M, Bartel B (2009) Peroxisome-associated matrix protein degradation in Arabidopsis. Proc Natl Acad Sci USA 106:4561–4566PubMedCentralPubMedCrossRefGoogle Scholar
  31. Liu Y, Björkman J, Urquhart A, Wanders RJA, Crane DI, Gould SJ (1999) PEX13 is mutated in complementation group 13 of the peroxisome-biogenesis disorders. Am J Hum Genet 65:621–634PubMedCentralPubMedCrossRefGoogle Scholar
  32. Mano S, Nakamori C, Nito K, Kondo M, Nishimura M (2006) The Arabidopsis pex12 and pex13 mutants are defective in both PTS1- and PTS2-dependent protein transport to peroxisomes. Plant J 47:604–618PubMedCrossRefGoogle Scholar
  33. McNew JA, Goodman JM (1994) An oligomeric protein is imported into peroxisomes in vivo. J Cell Biol 127:1245–1257PubMedCrossRefGoogle Scholar
  34. Michaels SD, Amasino RM (1998) A robust method for detecting single-nucleotide changes as polymorphic markers by PCR. Plant J 14:381–385PubMedCrossRefGoogle Scholar
  35. Monroe-Augustus M, Ramón NM, Ratzel SE, Lingard MJ, Christensen SE, Murali C, Bartel B (2011) Matrix proteins are inefficiently imported into Arabidopsis peroxisomes lacking the receptor-docking peroxin PEX14. Plant Mol Biol 77:1–15PubMedCentralPubMedCrossRefGoogle Scholar
  36. Mullen RT, Flynn CR, Trelease RN (2001) How are peroxisomes formed? The role of the endoplasmic reticulum and peroxins. Trends Plant Sci 6:256–261PubMedCrossRefGoogle Scholar
  37. Neff MM, Neff JD, Chory J, Pepper AE (1998) dCAPS, a simple technique for the genetic analysis of single nucleotide polymorphisms: experimental applications in Arabidopsis thaliana genetics. Plant J 14:387–392PubMedCrossRefGoogle Scholar
  38. Nito K, Hayashi M, Nishimura M (2002) Direct interaction and determination of binding domains among peroxisomal import factors in Arabidopsis thaliana. Plant Cell Physiol 43:355–366PubMedGoogle Scholar
  39. Nito K, Kamigaki A, Kondo M, Hayashi M, Nishimura M (2007) Functional classification of Arabidopsis peroxisome biogenesis factors proposed from analyses of knockdown mutants. Plant Cell Physiol 48:763–774PubMedGoogle Scholar
  40. Otera H, Setoguchi K, Hamasaki M, Kumashiro T, Shimizu N, Fujiki Y (2002) Peroxisomal targeting signal receptor Pex5p interacts with cargoes and import machinery components in a spatiotemporally differentiated manner: conserved Pex5p WXXXF/Y motifs are critical for matrix protein import. Mol Cell Biol 22:1639–1655PubMedCentralPubMedCrossRefGoogle Scholar
  41. Pires JR, Hong X, Brockmann C, Volkmer-Engert R, Schneider-Mergener J, Oschkinat H, Erdmann R (2003) The ScPex13p SH3 domain exposes two distinct binding sites for Pex5p and Pex14p. J Mol Biol 326:1427–1435PubMedCrossRefGoogle Scholar
  42. Pracharoenwattana I, Cornah JE, Smith SM (2007) Arabidopsis peroxisomal malate dehydrogenase functions in β-oxidation but not in the glyoxylate cycle. Plant J 50:381–390PubMedCrossRefGoogle Scholar
  43. Ramón NM, Bartel B (2010) Interdependence of the peroxisome-targeting receptors in Arabidopsis thaliana: PEX7 facilitates PEX5 accumulation and import of PTS1 cargo into peroxisomes. Mol Biol Cell 21:1263–1271PubMedCentralPubMedCrossRefGoogle Scholar
  44. Ratzel SE, Lingard MJ, Woodward AW, Bartel B (2011) Reducing PEX13 expression ameliorates physiological defects of late-acting peroxin mutants. Traffic 12:121–134PubMedCentralPubMedCrossRefGoogle Scholar
  45. Schumann H, Huesgen PF, Gietl C, Adamska I (2008) The DEG15 serine protease cleaves peroxisomal targeting signal 2-containing proteins in Arabidopsis. Plant J 148:1847–1856Google Scholar
  46. Shimozawa N, Suzuki Y, Zhang Z, Imamura A, Toyama R, Mukai S, Fujiki Y, Tsukamoto T, Osumi T, Orii T, Wanders RJ, Kondo N (1999) Nonsense and temperature-sensitive mutations in PEX13 are the cause of complementation group H of peroxisome biogenesis disorders. Hum Mol Genet 8:1077–1083PubMedCrossRefGoogle Scholar
  47. Stasinopoulos TC, Hangarter RP (1990) Preventing photochemistry in culture media by long-pass light filters alters growth of cultured tissues. Plant Physiol 93:1365–1369PubMedCentralPubMedCrossRefGoogle Scholar
  48. Stein K, Schell-Steven A, Erdmann R, Rottensteiner H (2002) Interactions of Pex7p and Pex18p/Pex21p with the peroxisomal docking machinery: implications for the first steps in PTS2 protein import. Mol Cell Biol 22:6056–6069PubMedCentralPubMedCrossRefGoogle Scholar
  49. Strader LC, Bartel B (2011) Transport and metabolism of the endogenous auxin precursor indole-3-butyric acid. Mol Plant 4:477–486PubMedCentralPubMedCrossRefGoogle Scholar
  50. Strader L, Culler Hendrickson A, Cohen J, Bartel B (2010) Conversion of endogenous indole-3-butyric acid to indole-3-acetic acid drives cell expansion in Arabidopsis seedlings. Plant Physiol 153:1577–1586PubMedCentralPubMedCrossRefGoogle Scholar
  51. Strader LC, Wheeler DL, Christensen SE, Berens JC, Cohen JD, Rampey RA, Bartel B (2011) Multiple facets of Arabidopsis seedling development require indole-3-butyric acid-derived auxin. Plant Cell 23:984–999PubMedCentralPubMedCrossRefGoogle Scholar
  52. Toyama R, Mukai S, Itagaki A, Tamura S, Shimozawa N, Suzuki Y, Kondo N, Wanders RJ, Fujiki Y (1999) Isolation, characterization and mutation analysis of PEX13-defective Chinese hamster ovary cell mutants. Hum Mol Genet 8:1673–1681PubMedCrossRefGoogle Scholar
  53. Walton PA, Hill PE, Subramani S (1995) Import of stably folded proteins into peroxisomes. Mol Biol Cell 6:675–683PubMedCentralPubMedCrossRefGoogle Scholar
  54. Waterham HR, Ebberink MS (2012) Genetics and molecular basis of human peroxisome biogenesis disorders. Biochim Biophys Acta 1822:1430–1441PubMedCrossRefGoogle Scholar
  55. Williams C, Distel B (2006) Pex13p: docking or cargo handling protein? Biochim Biophys Acta 1763:1585–1591PubMedCrossRefGoogle Scholar
  56. Woodward AW, Bartel B (2005) The Arabidopsis peroxisomal targeting signal type 2 receptor PEX7 is necessary for peroxisome function and dependent on PEX5. Mol Biol Cell 16:573–583PubMedCentralPubMedCrossRefGoogle Scholar
  57. Zolman BK, Bartel B (2004) An Arabidopsis indole-3-butyric acid-response mutant defective in PEROXIN6, an apparent ATPase implicated in peroxisomal function. Proc Natl Acad Sci USA 101:1786–1791PubMedCentralPubMedCrossRefGoogle Scholar
  58. Zolman BK, Yoder A, Bartel B (2000) Genetic analysis of indole-3-butyric acid responses in Arabidopsis thaliana reveals four mutant classes. Genetics 156:1323–1337PubMedCentralPubMedGoogle Scholar
  59. Zolman BK, Monroe-Augustus M, Thompson B, Hawes JW, Krukenberg KA, Matsuda SPT, Bartel B (2001a) chy1, an Arabidopsis mutant with impaired β-oxidation, is defective in a peroxisomal β-hydroxyisobutyryl-CoA hydrolase. J Biol Chem 276:31037–31046PubMedCrossRefGoogle Scholar
  60. Zolman BK, Silva ID, Bartel B (2001b) The Arabidopsis pxa1 mutant is defective in an ATP-binding cassette transporter-like protein required for peroxisomal fatty acid β-oxidation. Plant Physiol 127:1266–1278PubMedCentralPubMedCrossRefGoogle Scholar
  61. Zolman BK, Monroe-Augustus M, Silva ID, Bartel B (2005) Identification and functional characterization of Arabidopsis PEROXIN4 and the interacting protein PEROXIN22. Plant Cell 17:3422–3435PubMedCentralPubMedCrossRefGoogle Scholar
  62. Zolman BK, Nyberg M, Bartel B (2007) IBR3, a novel peroxisomal acyl-CoA dehydrogenase-like protein required for indole-3-butyric acid response. Plant Mol Biol 64:59–72PubMedCrossRefGoogle Scholar
  63. Zolman BK, Martinez N, Millius A, Adham AR, Bartel B (2008) Identification and characterization of Arabidopsis indole-3-butyric acid response mutants defective in novel peroxisomal enzymes. Genetics 180:237–251PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2014

Authors and Affiliations

  • Andrew W. Woodward
    • 1
    • 2
  • Wendell A. Fleming
    • 1
  • Sarah E. Burkhart
    • 1
  • Sarah E. Ratzel
    • 1
  • Marta Bjornson
    • 1
    • 3
  • Bonnie Bartel
    • 1
  1. 1.Department of Biochemistry and Cell BiologyRice UniversityHoustonUSA
  2. 2.Department of BiologyUniversity of Mary Hardin-BaylorBeltonUSA
  3. 3.Departments of Plant Biology and Plant SciencesUniversity of CaliforniaDavisUSA

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