, Volume 250, Issue 1, pp 317–323 | Cite as

An antibody against a conserved C-terminal consensus motif from plant alternative oxidase (AOX) isoforms 1 and 2 label plastids in the explosive dwarf mistletoe (Arceuthobium americanum, Santalaceae) fruit exocarp

  • Cynthia Ross Friedman
  • Bradford N. Ross
  • Garnet D. Martens
Original Article


Dwarf mistletoes, genus Arceuthobium (Santalaceae), are parasitic angiosperms that spread their seeds by an explosive process. As gentle heating triggers discharge in the lab, we wondered if thermogenesis (endogenous heat production) is associated with dispersal. Thermogenesis occurs in many plants and is enabled by mitochondrial alternative oxidase (AOX) activity. The purpose of this study was to probe Arceuthobium americanum fruit (including seed tissues) collected over a 10-week period with an anti-AOX antibody/gold-labeled secondary antibody to determine if AOX could be localized in situ, and if so, quantitatively assess whether label distribution changed during development; immunochemical results were evaluated with Western blotting. No label could be detected in the mitochondria of any fruit or seed tissue, but was observed in fruit exocarp plastids of samples collected in the last 2 weeks of study; plastids collected in week 10 had significantly more label than week 9 (p = 0.002). Western blotting of whole fruit and mitochondrial proteins revealed a signal at 30–36 kD, suggestive of AOX, while blots of whole fruit (but not mitochondrial fraction) proteins showed a second band at 40–45 kD, in agreement with plastid terminal oxidases (PTOXs). AOX enzymes are likely present in the A. americanum fruit, even though they were not labeled in mitochondria. The results strongly indicate that the anti-AOX antibody was labeling PTOX in plastids, probably at a C-terminal region conserved in both enzymes. PTOX in plastids may be involved in fruit ripening, although a role for PTOX in thermogenesis cannot be eliminated.


Alternative oxidase (AOX) Arceuthobium americanum Dwarf mistletoe Fruit Immunocytochemistry Plastid terminal oxidase (PTOX) 


  1. Andersson ME, Nordlund P (1999) A revised model of the active site of alternative oxidase. FEBS Lett 449:17–22PubMedCrossRefGoogle Scholar
  2. Antonsson B, Montessuit S, Sanchez B, Martinou JC (2001) Bax is present as a high molecular weight oligomer/complex in the mitochondrial membrane of apoptotic cells. J Biol Chem 276:11615–11623PubMedCrossRefGoogle Scholar
  3. Bots M, Feron R, Uehlein N, Weterings K, Kaldenhoff R, Mariani T (2005) PIP1 and PIP2 aquaporins are differentially expressed during tobacco anther and stigma development. J Exp Bot 56:113–121PubMedGoogle Scholar
  4. Carol P, Kuntz M (2001) A plastid terminal oxidase comes to light: implications for carotenoid biosynthesis and chlororespiration. Trends Plant Sci 6:31–36PubMedCrossRefGoogle Scholar
  5. Cournac L, Redding K, Ravenel J, Rumeau D, Josse E-M, Kuntz M, Peltier G (2000) Electron flow between photosystem II and oxygen in chloroplasts of photosystem I-deficient algae is mediated by a quinol oxidase involved in chlororespiration. J Biol Chem 275:17256–17262PubMedCrossRefGoogle Scholar
  6. Day DA, Wiskich JT (1995) Regulation of alternative oxidase activity in higher plants. J Bioenerg Biomembr 27:379–385PubMedCrossRefGoogle Scholar
  7. Hawksworth FG (1959) Ballistics of dwarf mistletoe seeds. Science 130:504CrossRefGoogle Scholar
  8. Hawksworth FG, Wiens D (1996) Dwarf mistletoes: biology, pathology, and systematics: USDA For Serv Hndbk No 709. US Government Printing Office, WashingtonGoogle Scholar
  9. Hinds TE, Hawksworth FG (1965) Seed dispersal velocity in four dwarf mistletoes. Science 148:517–519PubMedCrossRefGoogle Scholar
  10. Hoefnagel MHN, Wiskich JT (1998) Activation of the plant alternative oxidase by high reduction levels of the Q-pool and pyruvate. Arch Biochem Biophys 355:262–270PubMedCrossRefGoogle Scholar
  11. Ip DW (1992) Dwarf mistletoe. Nat Resour Can, Can For Serv, North For Cent, Edmonton, Alberta For Leafl No. 18Google Scholar
  12. Josse EM, Simkin AJ, Gaffé J, Labouré AM, Kuntz M, Carol P (2000) A plastid terminal oxidase associated with carotenoid desaturation during chromoplast differentiation. Plant Physiol 123:1427–1436PubMedCrossRefGoogle Scholar
  13. Kamata T, Matsukawa K, Kakizaki Y, Ito K (2009) In vivo redox state of the ubiquinone pool in the spadices of the thermogenic skunk cabbage, Symplocarpus renifolius. J Plant Res 122:645–649PubMedCrossRefGoogle Scholar
  14. Rezaul K, Wu L, Mayya V, Hwang SI, Han D (2005) A systematic characterization of mitochondrial proteome from human T leukemia cells. Mol Cell Proteomics 4:169–181PubMedGoogle Scholar
  15. Ross CM (2006) Viscin cells in the dwarf mistletoe Arceuthobium americanum: ‘green springs’ with potential roles in explosive seed discharge and seed adhesion. Davidsonia 17:75–86Google Scholar
  16. Ross Friedman CM, Ross BN, Martens GD (2010) Antibodies raised against tobacco aquaporins of the PIP2 class label the viscin tissue of the explosive dwarf mistletoe fruit. Plant Biol 12:229–233PubMedCrossRefGoogle Scholar
  17. Seymour RS (2004) Dynamics and precision of thermoregulatory responses of eastern skunk cabbage Symplocarpus foetidus. Plant Cell Environ 27:1014–1022CrossRefGoogle Scholar
  18. Seymour RS, Schultze-Motel P (1998) Physiological temperature regulation by flowers of the sacred lotus. Philos Trans R Soc Lond B Biol Sci 353:935–943CrossRefGoogle Scholar
  19. Seymour RS, Gibernau M, Ito K (2003) Thermogenesis and respiration of inflorescences of the dead horse arum, Helicodiceros muscivorous, a pseudo-thermoregulatory aroid associated with fly pollination. Funct Ecol 17:886–894CrossRefGoogle Scholar
  20. Seymour RS, Terry I, Roemer RB (2004) Respiration and thermogenesis by cones of the Australian cycad Macrozamia machinii. Funct Ecol 18:925–930CrossRefGoogle Scholar
  21. Shahbazi M, Gilbert M, Labouré A-M, Kuntz M (2007) Dual role of the plastid terminal oxidase in tomato. Plant Physiol 145:691–702PubMedCrossRefGoogle Scholar
  22. Skubatz H, Williamson PS, Schneider EL, Meeuse BJD (1990) Cyanide-insensitive respiration in thermogenic flowers of Victoria and Nelumbo. J Exp Bot 41:1335–1339CrossRefGoogle Scholar
  23. Tang W (1987) Heat production in cycad cones. Bot Gaz 148:165–174CrossRefGoogle Scholar
  24. Umbach AL, Siedow JN (1993) Covalent and noncovalent dimers of the cyanide-resistant alternative oxidase protein in higher plant mitochondria and their relationship to enzyme activity. Plant Physiol 103:845–854PubMedGoogle Scholar
  25. Van der Pijl L (1966) Ecological aspects of fruit evolution: a functional study of dispersal organs. Proc K Ned Akad Wet 69:597–640Google Scholar
  26. Wagner AM, Krab K, Wagner MJ, Moore AL (2008) Regulation of thermogenesis in flowering Araceae: the role of the alternative oxidase. Biochim Biophys Acta 1777:993–1000PubMedCrossRefGoogle Scholar
  27. Watling JR, Robinson SA, Seymour S (2006) Contribution of the alternative pathway to respiration during thermogenesis in flowers of the sacred lotus. Plant Physiol 140:1367–1373PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • Cynthia Ross Friedman
    • 1
  • Bradford N. Ross
    • 2
  • Garnet D. Martens
    • 2
  1. 1.Department of Biological SciencesThompson Rivers UniversityKamloopsCanada
  2. 2.BioImaging FacilityUniversity of British ColumbiaVancouverCanada

Personalised recommendations