Plant Molecular Biology

, Volume 78, Issue 4–5, pp 447–460 | Cite as

Arabidopsis light-dependent protochlorophyllide oxidoreductase A (PORA) is essential for normal plant growth and development

  • Troy Paddock
  • Daniel Lima
  • Mary E. Mason
  • Klaus Apel
  • Gregory A. Armstrong
Article

Abstract

During skotomorphogenesis in angiosperms, NADPH:protochlorophyllide oxidoreductase (POR) forms an aggregate of photolabile NADPH-POR-protochlorophyllide (Pchlide) ternary complexes localized to the prolamellar bodies within etioplasts. During photomorphogenesis, POR catalyzes the light-dependent reduction of Pchlide a to chlorophyllide (Chlide) a, which is subsequently converted to chlorophyll (Chl). In Arabidopsis there are three structurally related POR genes, denoted PORA, PORB and PORC. The PORA and PORB proteins accumulate during skotomorphogenesis. During illumination, PORA is only transiently expressed, whereas PORB and PORC persist and are responsible for bulk Chl synthesis throughout plant development. Here we have tested whether PORA is important for skotomorphogenesis by assisting in etioplast development, and normal photomorphogenic development. Using reverse genetic approaches, we have identified the porA-1 null mutant, which contains an insertion of the maize Dissociation transposable element in the PORA gene. Additionally, we have characterized PORA RNAi lines. The porA-1 and PORA RNAi lines display severe photoautotrophic growth defects, which can be partially rescued on sucrose-supplemented growth media. Elimination of PORA during skotomorphogenesis results in reductions in the volume and frequency of prolamellar bodies, and in photoactive Pchlide conversion. The porA-1 mutant characterization thus establishes a quantitative requirement for PORA in etioplast development by demonstrating significant membrane ultrastructural and biochemical defects, in addition to suggesting PORA-specific functions in photomorphogenesis and plant development.

Keywords

Protochlorophyllide Oxidoreductase PORA Chlorophyll biosynthesis Etioplast Arabidopsis 

Supplementary material

11103_2012_9873_MOESM1_ESM.ppt (158 kb)
Supplementary material 1 (PPT 158 kb)

References

  1. Armstrong GA, Runge S, Frick G, Sperling U, Apel K (1995) Identification of NADPH:protochlorophyllide oxidoreductases A and B: a branched pathway for light-dependent chlorophyll biosynthesis in Arabidopsis thaliana. Plant Physiol 108:1505–1517PubMedCrossRefGoogle Scholar
  2. Czechowski T, Stitt M, Altmann T, Udvardi MK, Scheible WR (2005) Genome-wide identification and testing of superior reference genes for transcript normalization in Arabidopsis. Plant Physiol 139:5–17PubMedCrossRefGoogle Scholar
  3. Eguchi S, Takano H, Ono K, Takio S (2002) Photosynthetic electron transport regulates the stability of the transcript for the protochlorophyllide oxidoreductase gene in the liverwort, Marchantia paleacea var. diptera. Plant Cell Physiol 43:573–577PubMedCrossRefGoogle Scholar
  4. Forreiter C, Cleve B, Schmidt A, Apel K (1991) Evidence for a general light-dependent negative control of NADPH-protochlorophyllide oxidoreductase in angiosperms. Planta 183:126–132CrossRefGoogle Scholar
  5. Franck F, Sperling U, Frick G, Pochert B, van Cleve B, Apel K, Armstrong GA (2000) Regulation of etioplast pigment-protein complexes, inner membrane architecture, and protochlorophyllide a chemical heterogeneity by light-dependent NADPH:protochlorophyllide oxidoreductases A and B. Plant Physiol 124:1678–1696PubMedCrossRefGoogle Scholar
  6. Frick G, Su Q, Apel K, Armstrong GA (2003) An Arabidopsis porB porC double mutant lacking light-dependent NADPH:protochlorophyllide oxidoreductases B and C is highly chlorophyll-deficient and developmentally arrested. Plant J 35:141–153PubMedCrossRefGoogle Scholar
  7. Fusada N, Masuda T, Kuroda H, Shiraishi T, Shimada H, Ohta H, Takamiya K (2000) NADPH-protochlorophyllide oxidoreductase in cucumber is encoded by a single gene and its expression is transcriptionally enhanced by illumination. Photosynth Res 64:147–154PubMedCrossRefGoogle Scholar
  8. Heyes DJ, Hunter CN (2005) Making light work of enzyme catalysis: protochlorophyllide oxidoreductase. Trends Biochem Sci 30:642–649PubMedCrossRefGoogle Scholar
  9. Hilson P, Allemeersch J, Altmann T, Aubourg S, Avon A, Beynon J, Bhalerao RP, Bitton F, Caboche M, Cannoot B, Chardakov V, Cognet-Holliger C, Colot V, Crowe M, Darimont C, Durinck S, Eickhoff H, de Longevialle AF, Farmer EE, Grant M, Kuiper MTR, Lehrach H, Leon C, Leyva A, Lundeberg J, Lurin C, Moreau Y, Nietfeld W, Paz-Ares J, Reymond P, Rouze P, Sandberg G, Segura MD, Serizet C, Tabrett A, Taconnat L, Thareau V, Van Hummelen P, Vercruysse S, Vuylsteke M, Weingartner M, Weisbeek PJ, Wirta V, Wittink FRA, Zabeau M, Small I (2004) Versatile gene-specific sequence tags for Arabidopsis functional genomics: transcript profiling and reverse genetics applications. Genome Res 14:2176–2189PubMedCrossRefGoogle Scholar
  10. Holtorf H, Reinbothe S, Reinbothe C, Bereza B, Apel K (1995) Two routes of chlorophyllide synthesis that are differentially regulated by light in barley (Hordeum vulgare L.). Proc Natl Acad Sci USA 92:3254–3258PubMedCrossRefGoogle Scholar
  11. Iwamoto K, Fukuda H, Sugiyama M (2001) Elimination of POR expression correlates with red leaf formation in Amaranthus tricolor. Plant J 27:275–284PubMedCrossRefGoogle Scholar
  12. Masuda T, Takamiya K (2004) Novel insights into the enzymology, regulation and physiological functions of light-dependent protochlorophyllide oxidoreductase in angiosperms. Photosynth Res 81:1–29PubMedCrossRefGoogle Scholar
  13. Masuda T, Fusada N, Shiraishi T, Kuroda H, Awai K, Shimada H, Ohta H, Takamiya K (2002) Identification of two differentially regulated isoforms of protochlorophyllide oxidoreductase (POR) from tobacco revealed a wide variety of light- and development-dependent regulations of POR gene expression among angiosperms. Photosynth Res 74:165–172PubMedCrossRefGoogle Scholar
  14. Masuda S, Ikeda R, Masuda T, Hashimoto H, Tsuchiya T, Kojima H, Nomata J, Fujita Y, Mimuro M, Ohta H, Takamiya K (2009) Prolamellar bodies formed by cyanobacterial protochlorophyllide oxidoreductase in Arabidopsis. Plant J 58:952–960PubMedCrossRefGoogle Scholar
  15. Matsumoto F, Obayashi T, Sasaki-Sekimoto Y, Ohta H, Takamiya K, Masuda T (2004) Gene expression profiling of the tetrapyrrole metabolic pathway in Arabidopsis with a mini-array system. Plant Physiol 135:2379–2391Google Scholar
  16. Oosawa N, Masuda T, Awai K, Fusada N, Shimada H, Ohta H, Takamiya K (2000) Identification and light-induced expression of a novel gene of NADPH-protochlorophyllide oxidoreductase isoform in Arabidopsis thaliana. FEBS Lett 474:133–136PubMedCrossRefGoogle Scholar
  17. op den Camp RGL, Przybyla D, Ochsenbein C, Laloi C, Kim C, Danon A, Wagner D, Hideg E, Gobel C, Feussner I, Nater M, Apel K (2003) Rapid induction of distinct stress responses after the release of singlet oxygen in Arabidopsis. Plant Cell 15:2320–2332PubMedCrossRefGoogle Scholar
  18. Osuna D, Usadel B, Morcuende R, Gibon Y, Blasing OE, Hohne M, Gunter M, Kamlage B, Trethewey R, Scheible W, Stitt M (2007) Temporal responses of transcripts, enzyme activities and metabolites after adding sucrose to carbon-deprived Arabidopsis seedlings. Plant J 49:463–491PubMedCrossRefGoogle Scholar
  19. Paddock T, Mason M, Lima D, Armstrong G (2010) Arabidopsis protochlorophyllide oxidoreductase A (PORA) restores bulk chlorophyll synthesis and normal development to a porB porC double mutant. Plant Mol Biol 72:445–457PubMedCrossRefGoogle Scholar
  20. Parinov S, Sevugan M, De Y, Yang W, Kumaran M, Sundaresan V (1999) Analysis of flanking sequences from dissociation insertion lines: a database for reverse genetics in Arabidopsis. Plant Cell 11:2263–2270PubMedCrossRefGoogle Scholar
  21. Parks BM (2003) The red side of photomorphogenesis. Plant Physiol 133:1437–1444PubMedCrossRefGoogle Scholar
  22. Rosinski J, Rosen WG (1972) Characterization of prolamellar chloroplast development: fine structure and chlorophyll synthesis. Q Rev Biol 47:160–191CrossRefGoogle Scholar
  23. Runge S, Sperling U, Frick G, Apel K, Armstrong GA (1996) Distinct roles for light-dependent NADPH:protochlorophyllide oxidoreductases (POR) A and B during greening in higher plants. Plant J 9:513–523PubMedCrossRefGoogle Scholar
  24. Schoefs B, Franck F (2003) Protochlorophyllide reduction: mechanisms and evolution. Photochem Photobiol 78:543–557PubMedCrossRefGoogle Scholar
  25. Selstam E, Sandelius AS (1984) A comparison between prolamellar bodies and prothylakoid membranes of etioplasts of dark-grown wheat concerning lipid and polypeptide composition. Plant Physiol 76:1036–1040PubMedCrossRefGoogle Scholar
  26. Selstam E, Schelin J, Brain T, Williams WP (2002) The effects of low pH on the properties of protochlorophyllide oxidoreductase and the organization of prolamellar bodies of maize (Zea mays). Eur J Biochem 269:2336–2346PubMedCrossRefGoogle Scholar
  27. Solymosi K, Martinez K, Kristof Z, Sundqvist C, Boddi B (2004) Plastid differentiation and chlorophyll biosynthesis in different leaf layers of white cabbage (Brassica oleracea cv. capitata). Physiol Plantarum 121:520–529CrossRefGoogle Scholar
  28. Spano AJ, He Z, Michel H, Hunt DF, Timko MP (1992) Molecular cloning, nuclear gene structure, and developmental expression of NADPH: protochlorophyllide oxidoreductase in pea (Pisum sativum L.). Plant Mol Biol 18:967–972PubMedCrossRefGoogle Scholar
  29. Sperling U, van Cleve B, Frick G, Apel K, Armstrong GA (1997) Overexpression of light-dependent PORA or PORB in plants depleted of endogenous POR by far-red light enhances seedling survival in white light and protects against photooxidative damage. Plant J 12:649–658PubMedCrossRefGoogle Scholar
  30. Sperling U, Franck F, van Cleve B, Frick G, Apel K, Armstrong GA (1998) Etioplast differentiation in Arabidopsis: both PORA and PORB restore the prolamellar body and photoactive protochlorophyllide-F655 to the cop1 photomorphogenic mutant. Plant Cell 10:283–296PubMedCrossRefGoogle Scholar
  31. Su Q, Frick G, Armstrong G, Apel K (2001) PORC of Arabidopsis thaliana: a third light- and NADPH-dependent protochlorophyllide oxidoreductase that is differentially regulated by light. Plant Mol Biol 47:805–813PubMedCrossRefGoogle Scholar
  32. Sundaresan V, Springer P, Volpe T, Haward S, Jones JD, Dean C, Ma H, Martienssen R (1995) Patterns of gene action in plant development revealed by enhancer trap and gene trap transposable elements. Genes Dev 9:1797–1810PubMedCrossRefGoogle Scholar
  33. Teakle GR, Griffiths WT (1993) Cloning, characterization and import studies on protochlorophyllide reductase from wheat (Triticum aestivum). Biochem J 296(Pt 1):225–230PubMedGoogle Scholar
  34. Thum KE, Shin MJ, Palenchar PM, Kouranov A, Coruzzi GM (2004) Genome-wide investigation of light and carbon signaling interactions in Arabidopsis. Genome Biol 5:R10PubMedCrossRefGoogle Scholar
  35. Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, De Paepe A, Speleman F (2002) Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol 3:R34CrossRefGoogle Scholar
  36. Wang T, Iyer LM, Pancholy R, Shi X, Hall TC (2005) Assessment of penetrance and expressivity of RNAi-mediated silencing of the Arabidopsis phytoene desaturase gene. New Phytol 167:751–760PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2012

Authors and Affiliations

  • Troy Paddock
    • 1
  • Daniel Lima
    • 1
  • Mary E. Mason
    • 2
  • Klaus Apel
    • 3
  • Gregory A. Armstrong
    • 1
  1. 1.Department of Plant Cellular and Molecular BiologyThe Ohio State UniversityColumbusUSA
  2. 2.Department of EntomologyOhio Agricultural Research and Development CenterWoosterUSA
  3. 3.Boyce Thompson Institute for Plant ResearchIthacaUSA

Personalised recommendations