The Peroxisome

by Robert T Mullen

Department of Molecular & Cellular Biology, College of Biological Sciences,
University of Guelph. ON N1G 2W1 Canada.

Peroxisomes targeted using YFP-PTS1 moving along actin filaments (GFP-mTalin) in hypocotyl cells of an Arabidopsis seedling. Chloroplasts exhibit Red auto fluorescence.

Peroxisomes are bound by a single membrane and contain a coarsely granular interior (matrix), sometimes with large crystalline inclusions [1]. They typically vary in size (~0.2 um to 4 um) depending upon the cell or tissue type and display remarkable structural variability, ranging from nearly spherical to ovoid and frequently pleomorphic with large invaginations that are dependent upon their placement among adjacent organelles, particularly chloroplasts and oil bodies [1,2].
Peroxisomes also display a wide range of intracellular movements that, depending on the cell/tissue type, include both slow and fast (up to ~6.5 um/sec) unidirectional and bidirectional movements, stop-and-go movements, and more subtle apparent random oscillations [3]. All of these different types of motility appear to be interchangeable, and are mediated by the interactions of peroxisomes with actin microfilaments and myosin motors [3,4].
Peroxisomes are categorized into different classes based on their complement of enzymes that correspond to the specific cell or tissue type and/or stage of plant growth and development [5]. Examples of such specialized plant peroxisomes are 1) glyoxysomes using fatty acid -oxidation and glyoxylate cycle enzymes to cooperatively convert storage lipids to sugars during post-germinative seedling growth, 2) leaf (or leaf-type) peroxisomes catalyzing essential reactions of photorespiration within photosynthetically active tissues, 3) gerontosomes using re-acquired glyoxysomal enzymes to catabolize lipids in senescing tissues, 4) root nodule peroxisomes participating in ureide biosynthesis in uninfected nodule cells, and 5) “unspecialized” peroxisomes, which are relatively undifferentiated peroxisomes found throughout the plant body. A unifying feature of all of these peroxisomes is the presence of catalase and essential oxidases that participate in detoxifications of reactive oxygen species generated within the organelle in a multitude of ways [6]. Peroxisomes can also uniquely convert from one class of specialized peroxisome to another. Such interconversions can occur several times within cells of the same tissue in response to changes in internal and/or external cue and are most often achieved through changes in enzyme content within a steady state number of pre-existing peroxisomes per cell [5].
Peroxisomes do not possess their own genome and lack ribosomes. As a consequence, matrix and membrane destined peroxisomal proteins are nuclear encoded, synthesized on free cytosolic ribosomes, and targeted post-translationally to the organelle in a regulated manner [7]. At least two types of evolutionarily conserved peroxisomal targeting signals (PTSs) are capable of directing proteins to the peroxisomal matrix – including passenger proteins such as an autofluorescent protein (e.g., GFP) and thus serve as a convenient marker for peroxisomes in transformed plant cells [8,9]. The type 1 PTS (PTS1) is an uncleaved C-terminal tripeptide motif, i.e., small-basic-hydrophobic residues, or variants thereof, that is found in most peroxisomal matrix-destined proteins. The type 2 PTS (PTS2) is a nonapeptide motif (-R/K-X6-H/Q-A/L/F-) [where X indicates any amino acid) located in the N-terminus of another set of matrix proteins that are proteolytically processed after import into peroxisomes.
Studies of the PTS1- and PTS2-mediated protein import pathways including the identification and characterization of proteinaceous receptors and several components of the translocation apparatus have revealed at least two distinct pathways that converge at peroxisomal boundary membrane [7]. The proteins involved in these import processes as well as other proteins involved in different aspects of peroxisomal biogenesis including protein import, organelle maturation (maintenance/assembly/differentiation), replication and inheritance are termed peroxins (PEX). Overall, much progress has been made recently through Arabidopisis thaliana genome searches for genes that code for peroxins, with at least 23 different plant PEX genes having been identified [10].
Peroxisomal membrane proteins (PMPs) do not possess a PTS1 or PTS2 and thus the targeting pathway(s) for the import of PMPs is considered to be distinct from the targeting pathways used by matrix-destined proteins [11]. Indeed, while some nascent PMPs are sorted to peroxisomes directly from the cytosol, other PMPs are sorted indirectly to peroxisomes by way of the endoplasmic reticulum (ER) and ER-derived vesicles [12]. The discovery of this indirect sorting pathway for some PMPs has helped lead to the resurgence of the idea that the ER serves as the membrane source of new or differentiating peroxisomes.
Membrane peroxisomal targeting signals have been described for several PMPs, most of which do not appear to exhibit a consensus in terms of their general features [11]. Nonetheless, these varied signals can be linked to a passenger protein of choice (e.g., GFP) and serve as convenient markers for PMPs sorting either directly to peroxisomes or indirectly via the ER.

Peroxisomes in plants

Micrographs of peroxisomes in plant cells. (A) Electron micrograph showing a peroxisome (glyoxysome [g]) and several adjacent oil bodies (ob) in cotyledon cells of dark-grown cucumber seedlings. Abbreviations: cw; cell wall; m, mitochondrion; v, vacuole. (B) Immunfluorescence micrograph illustrating the localization of peroxisomal matrix catalase in several tobacco BY-2 suspension-cultured cells. (C) Differential interference contrast image of the same group of BY-2 cells shown in B. (A-E) Representative confocal projection images (overlays) of peroxisomes (green) and chloroplasts (red) in leaf mesophyll cells (A), guard cells (B), petals (C), roots (D), and a leaf secretory trichome (E) of a light-grown (non-infected) tobacco (N. benthamiana) plant stably expressing the peroxisomal matrix fusion protein GFP-SKL. The insets in (D-H) show higher magnified images illustrating the wide range of shapes, sizes and distributions of peroxisomes in the same organs of a GFP-SKL-transformed tobacco plant (F) High-magnification, three-dimensional and computer-rendered reconstructions of an aggregate of peroxisomes (green) and plastids (red) in a secretory trichome of a GFP-SKL-transformed (non-infected) tobacco plant; compare with the peroxisomal/plastid aggregates shown in (E). Scale bars = 0.5 um (A), 10 um (C), 5 mm (D), 7 mm (E), 50 mm (F and G), 25 mm (H) and 2.5 um (I). Scale bars in inserts for (D-H) = 1 mm (D), 3.5 mm (E-G) and 10 mm (H).

Images courtesy of Prof. R. N. Trelease, Arizona State University (A), P. Dhanoa, University of Guelph (B-C), and from Mullen et al. 2006. Vol 84, pg. 555 [Ref. 2] (D-H).
Peroxule

Peroxisomal extensions named 'peroxules' have been described by Scott et al. It has been shown that peroxules are produced within seconds of exposure to hydrogen peroxide and hydroxyl radicals. For details see Peroxule extension over ER defined paths constitutes a rapid subcellular response to hydroxyl stress. Sinclair et al. Plant J. 2009 Mar 9.PMID: 19292761. 

References

1. Huang AHC, RN Trelease, TS Moore. 1983. Plant Peroxisomes. Academic Press

2. Mullen RT, AW McCartney, CR Flynn, GST Smith. 2006. Canadian Journal of Botany 84: 551-564

3. Muench DG, RT Mullen. 2003. Plant Science 164: 307-315

4. Hashimoto K, H Igarashi, S Mano, M Nishimura, T Shimmen, E Yokota. 2005. Plant Cell Physiology 46: 782-789.

5. Hayashi M, M Nishimura. 2003. Current Opinions in Plant Biology 6: 577-582

6. del Rio LA, FJ Corpas, LM Sandalio, JM Palma, JB Barroso. 2003. IUBMB Life. 55: 71-81.

7. Baker A, IA Sparkes. 2005 Current Opinions in Plant Biology 8: 640-647.

8. Mullen RT. 2002. In Plant Peroxisomes. Kluwer Academic Press. Pp. 339-384.

9. Dhanoa PK, AM Sinclair, RT Mullen, J Mathur. 2006. Canadian Journal of Botany 84: 515-522.

10. Hayashi M, M Nishimura. 2006. Biochimica et Biophysica Acta 1763: 1383-1391.

11. Trelease. 2002. In Plant Peroxisomes. Kluwer Academic Press. Pp. 305-338.

12. Mullen RT, RN Trelease. 2006. Biochimica et Biophysica Acta 1763: 1655-1668

____________________________________________________________________________________________
Fluorescent Protein Probes for Peroxisomes
compiled by Alison Sinclair
Department of Molecular & Cellular Biology, University of Guelph. Canada.

A large number of protein fusions target to peroxisomes [23]. The Table presents some of the commonly used probes. Note, that Peroxin16 [9] has been shown to have a dual localization as it transits through the ER [11].


Peroxisome FP-Probes

References for the Table

1. Carrie, C., Murcha, M.W., Millar, A.H., Smith, S.M. and Whelan, J. 2007. Nine 3-ketoacyl-CoA thiolases (KATs) and acetoacetyl-CoA thiolases (ACATs) encoded by five genes in arabidopsis thaliana are targeted either to peroxisomes or cytosol but not to mitochondria. Plant Molecular Biology. 63(1): 97-108.

2. Dammann, C., Ichida, A., Hong, B., Romanowsky, S.M., Hrabak, E.M., Harmon, A.C., Pickard, B.G. and Harper, J.F. 2003. Subcellular targeting of nine calcium-dependent protein kinase isoforms from arabidopsis. Plant Physiol. 132(4): 1840-1848.

3. Fulda, M., Shockey, J., Werber, M., Wolter, F.P. and Heinz, E. 2002. Two long-chain acyl-CoA synthetases from arabidopsis thaliana involved in peroxisomal fatty acid beta-oxidation. The Plant Journal. 32(1): 93-103.

4. Goepfert, S., Vidoudez, C., Rezzonico, E., Hiltunen, J.K. and Poirier, Y. 2005. Molecular identification and characterization of the arabidopsis {delta} 3, 5,{delta} 2, 4-dienoyl-coenzyme A isomerase, a peroxisomal enzyme participating in the {beta}-oxidation cycle of unsaturated fatty acids. Plant Physiology. 138: 1947-1956.

5. Hu, J., Aguirre, M., Peto, C., Alonso, J., Ecker, J. and Chory, J. 2002. A role for peroxisomes in photomorphogenesis and development of Arabidopsis. Science. 297(5580): 405-409.

6. Igarashi, D., Miwa, T., Seki, M., Kobayashi, M., Kato, T., Tabata, S., Shinozaki, K. and Ohsumi, C. 2003. Identification of photorespiratory glutamate:Glyoxylate aminotransferase (GGAT) gene in arabidopsis. The Plant Journal. 33(6): 975-987.

7. Jedd, G and Chua N.H.2002.Visualization of peroxisomes in living plant cells reveals acto-myosin-dependent cytoplasmic streaming and peroxisome budding. Plant Cell Physiology. 43(4):384-392.

8. Koo, A. J. K., Chung, H.S., Kobayashi, Y. and Howe, G.A. 2006. Identification of a peroxisomal acyl-activating enzyme involved in the biosynthesis of jasmonic acid in arabidopsis. The Journal of Biological Chemistry. 281(44): 33511-33520.

9. Lin, Y., Cluette-Brown, J.E. and Goodman, H.M. 2004. The peroxisome deficient arabidopsis mutant sse1 exhibits impaired fatty acid synthesis. Plant Physiology. 135(2): 814-827.

10. Lingard, M.J., and Trelease, R.N. 2006. Five arabidopsis peroxin 11 homologs individually promote peroxisome elongation, duplication or aggregation. Journal of Cell Science. 119(9): 1961-1972.

11. Lisenbee, C.S., Karnik, S.K. and Trelease, R.N. 2003. Overexpression and mislocalization of a tail-anchored GFP redefines the identity of peroxisomal ER. Traffic. 4(7): 491-501.

12. Ma, C., Haslbeck, M., Babujee, L., Jahn, O. and Reumann, S. 2006. Identification and characterization of a stress-inducible and a constitutive small heat-shock protein targeted to the matrix of plant peroxisomes. Plant Physiol. 141(1): 47-60.

13. Mano, S., Hayashi, M. and Nishimura, M. 1999. Light regulates alternative splicing of hydroxypyruvate reductase in pumpkin. The Plant Journal. 17(3): 309.

14. Mano, S., Nakamori, C., Nito, K., Kondo, M. and Nishimura, M. 2006. The arabidopsis pex12 and pex13 mutants are defective in both PTS1- and PTS2-dependent protein transport to peroxisomes. The Plant Journal. 47(4): 604-618.

15. Mano, S., Nakamori, C., Hayashi, M., Kato, A., Kondo, M. and Nishimura, M. 2002. Distribution and characterization of peroxisomes in arabidopsis by visualization with GFP: Dynamic morphology and actin-dependent movement. Plant and Cell Physiology. 43(3): 331-341.

16. Mathur, J, Mathur, N,and Hülskamp, M.2002. Simultaneous visualization of peroxisomes and cytoskeletal elements reveals actin and not microtubule-based peroxisome motility in plants. Plant Physiology 128(3):1031-1045.

17.  Orth, T., Reumann, S., Zhang, X., Fan, J., Wenzel, D., Quan, S. and Hu, J. 2007. The PEROXIN11 protein family controls peroxisome proliferation in arabidopsis. The Plant Cell. 19(1): 333-350.

18.  Pracharoenwattana, I., Cornah, J.E. and Smith, S.M. 2005. Arabidopsis peroxisomal citrate synthase is required for fatty acid respiration and seed germination. The Plant Journal. 17(7): 2037-2048.

19.  Reisen, D., and Hanson, M.R. 2007. Association of six YFP-myosin XI-tail fusions with mobile plant cell organelles. BMC Plant Biology. 7(6):

20.  Schneider, K., Kienow, L., Schmelzer, E., Colby, T., Bartsch, M., Miersch, O., Wasternack, C., Kombrink, E. and Stuible, H.-. 2005. A new type of peroxisomal acyl-coenzyme A synthetase from arabidopsis thaliana has the catalytic capacity to activate biosynthetic precursors of jasmonic acid. J. Biol. Chem. 280(14): 13962-13972.

21.  Sparkes, I.A., Brandizzi, F., Slocombe, S.P., El-Shami, M., Hawes, C. and Baker, A. 2003. An arabidopsis pex10 null mutant is embryo lethal, implicating peroxisomes in an essential role during plant embryogenesis. Plant Physiology. 133(4): 1809-1819.

22.  Strassner, J., Schaller, F., rick, U.B., Howe, G.A., Weiler, E.W., Amrhein, N., Macheroux, P. and Schaller, A. 2002. Characterization and cDNA-microarray expression analysis of 12-oxophytodienoate reductases reveals differential roles for octadecanoid biosynthesis in the local versus the systemic wound response. The Plant Journal. 34(4): 585-601.

23.  Cutler SR, Ehrhardt DW, Griffitts JS, Somerville CR. 2000.Random GFP::cDNA fusions enable visualization of subcellular structures in cells of Arabidopsis at a high frequency.Proc Natl Acad Sci U S A. 28;97(7):3718-3723.