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The control of chlorophyll levels in maturing kiwifruit

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Abstract

Chlorophyll is present in many plant organs, including immature fruit where it is usually degraded during ripening. Mature green kiwifruit (Actinidia deliciosa) are an exception, with high concentrations of chlorophyll remaining in the fruit flesh. In gold-fleshed kiwifruit (A. chinensis), chlorophyll is degraded to colourless catabolites upon fruit ripening, leaving yellow carotenoids visible. We have identified candidate genes for the control of chlorophyll degradation in kiwifruit and examined the transcript levels of these genes in maturing kiwifruit using quantitative real-time PCR. Results indicate that the biosynthesis and degradation, or turnover, of chlorophyll is transcriptionally regulated in green- and gold-fleshed kiwifruit. Both species of kiwifruit were found to have two homologues of the stay-green gene (SGR), a small protein that is postulated to aid in the dismantling of the light-harvesting complex, allowing free chlorophyll to enter the degradation pathway. However, with the exception of very mature green fruit, where degreening was observed, SGR2 was more highly expressed in gold fruit, indicating a potential regulatory step of chlorophyll degradation. When the SGR genes were over-expressed in tobacco leaves, degreening was observed. Our results show that chlorophyll degradation is differentially regulated in kiwifruit, and suggest that gold kiwifruit transcribe more degradation genes, leading to earlier and more sustained chlorophyll degradation in this fruit than in green kiwifruit.

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Abbreviations

CAO:

Chlorophyll a oxygenase

CBR:

Chlorophyll b reductase

CLH:

Chlorophyllase

CLS:

Chlorophyll synthase

DAFB:

Days after full bloom

EST:

Expressed sequence tag

GLUTR:

Glutamyl tRNA reductase

LHCB:

Light-harvesting chlorophyll a/b binding complex

NOL:

NYC-one like

NYC1:

Non-yellow colouring 1

PAO:

Pheophorbide a oxygenase

PPH:

Pheophytin pheophorbide hydrolase

RBCS:

Small subunit of ribulose-1,5-bisphosphate carboxylase

RCC:

Red chlorophyll catabolite

RCCR:

Red chlorophyll catabolite reductase

RUBISCO:

Ribulose-1,5-bisphosphate carboxylase

SGR:

Stay-green

References

  • Akhtar M, Goldschmidt E, John I, Rodoni S, Matile P, Grierson D (1999) Altered patterns of senescence and ripening in gf, a stay-green mutant of tomato (Lycopersicon esculentum mill.). J Exp Bot 50:1115–1122

    CAS  Google Scholar 

  • Alos E, Roca M, Iglesias DJ, Minguez-Mosquera MI, Damasceno CMB, Thannhauser TW, Rose JKC, Talon M, Cercos M (2008) An evaluation of the basis and consequences of a stay-green mutation in the navel negra citrus mutant using transcriptomic and proteomic profiling and metabolite analysis. Plant Physiol 147:1300–1315

    Article  PubMed  CAS  Google Scholar 

  • Ampomah-Dwamena C, McGhie T, Wibisono R, Montefiori M, Hellens RP, Allan AC (2009) The kiwifruit lycopene beta-cyclase plays a significant role in carotenoid accumulation in fruit. J Exp Bot 60:3765–3779

    Article  PubMed  CAS  Google Scholar 

  • Armstead I, Donnison I, Aubry S, Harper J, Hörtensteiner S, James C, Mani J, Moffet M, Ougham H, Roberts L, Thomas A, Weeden N, Thomas H, King I (2006) From crop to model to crop: Identifying the genetic basis of the staygreen mutation in the Lolium/Festuca forage and amenity grasses. New Phytol 172:592–597

    Article  PubMed  Google Scholar 

  • Armstead I, Donnison I, Aubry S, Harper J, Hortensteiner S, James C, Mani J, Moffet M, Ougham H, Roberts L, Thomas A, Weeden N, Thomas H, King I (2007) Cross-species identification of Mendel’s / locus. Science 315:73

    Article  PubMed  CAS  Google Scholar 

  • Ashton OBO, Wong M, McGhie TK, Vather R, Wang Y, Requejo-Jackman C, Ramankutty P, Woolf AB (2006) Pigments in avocado tissue and oil. J Agric Food Chem 54:10151–10158

    Article  PubMed  CAS  Google Scholar 

  • Aubry S, Mani J, Hörtensteiner S (2008) Stay-green protein, defective in Mendel’s green cotyledon mutant, acts independent and upstream of pheophorbide a oxygenase in the chlorophyll catabolic pathway. Plant Mol Biol 67:243–256

    Article  PubMed  CAS  Google Scholar 

  • Barry CS (2009) The stay-green revolution: recent progress in deciphering the mechanisms of chlorophyll degradation in higher plants. Plant Sci 176:325–333

    Article  CAS  Google Scholar 

  • Bedbrook JR, Smith SM, Ellis RJ (1980) Molecular cloning and sequencing of cDNA encoding the precursor to the small subunit of chloroplast ribulose-1,5-bisphosphate carboxylase. Nature 287:692–697

    Article  CAS  Google Scholar 

  • Ben-Arie R, Gross J, Sonego L (1982) Changes in ripening-parameters and pigments of the chinese gooseberry (kiwi) during ripening and storage. Sci Hortic Amst 18:65–70

    Article  Google Scholar 

  • Borovsky Y, Paran I (2008) Chlorophyll breakdown during pepper fruit ripening in the chlorophyll retainer mutation is impaired at the homolog of the senescence-inducible stay-green gene. Theor Appl Genet 117:235–240

    Article  PubMed  CAS  Google Scholar 

  • Buchanan-Wollaston V, Ainsworth C (1997) Leaf senescence in Brassica napus: cloning of senescence related genes by subtractive hybridisation. Plant Mol Biol 33:821–834

    Article  PubMed  CAS  Google Scholar 

  • Cha KW, Lee YJ, Koh HJ, Lee BM, Nam YW, Paek NC (2002) Isolation, characterization, and mapping of the stay green mutant in rice. Theor Appl Genet 104:526–532

    Article  PubMed  CAS  Google Scholar 

  • Chang S, Puryear J, Cairney J (1993) A simple and efficient method for isolating RNA from pine trees. Plant Mol Biol Rep 11:113–116

    Article  CAS  Google Scholar 

  • Cheung AY, McNellis T, Piekos B (1993) Maintenance of chloroplast components during chromoplast differentiation in the tomato mutant green flesh. Plant Physiol 101:1223–1229

    PubMed  CAS  Google Scholar 

  • Crowhurst R, Gleave A, MacRae E, Ampomah-Dwamena C, Atkinson R, Beuning L, Bulley S, Chagne D, Marsh K, Matich A, Montefiori M, Newcomb R, Schaffer R, Usadel B, Allan A, Boldingh H, Bowen J, Davy M, Eckloff R, Ferguson AR, Fraser L, Gera E, Hellens R, Janssen B, Klages K, Lo K, MacDiarmid R, Nain B, McNeilage M, Rassam M, Richardson A, Rikkerink E, Ross G, Schroder R, Snowden K, Souleyre E, Templeton M, Walton E, Wang D, Wang M, Wang Y, Wood M, Wu R, Yauk Y-K, Laing W (2008) Analysis of expressed sequence tags from Actinidia: Applications of a cross species EST database for gene discovery in the areas of flavor, health, color and ripening. BMC Genomics 9:351

    Article  PubMed  Google Scholar 

  • Eckhardt U, Grimm B, Hörtensteiner S (2004) Recent advances in chlorophyll biosynthesis and breakdown in higher plants. Plant Mol Biol 56:1–14

    Article  PubMed  CAS  Google Scholar 

  • Efrati A, Eyal Y, Paran I (2005) Molecular mapping of the chlorophyll retainer (cl) mutation in pepper (Capsicum spp.) and screening for candidate genes using tomato ESTs homologous to structural genes of the chlorophyll catabolism pathway. Genome 48:347–351

    Article  PubMed  CAS  Google Scholar 

  • Guiamet JJ, Schwartz E, Pichersky E, Nooden LD (1991) Characterization of cytoplasmic and nuclear mutations affecting chlorophyll and chlorophyll-binding proteins during senescence in soybean. Plant Physiol 96:227–231

    Article  PubMed  CAS  Google Scholar 

  • Harman JE, McDonald B (1989) Controlled atmosphere storage of kiwifruit. Effect on fruit quality and composition. Sci Hortic Amst 37:303–315

    Article  Google Scholar 

  • Hellens R, Allan A, Friel E, Bolitho K, Grafton K, Templeton M, Karunairetnam S, Gleave A, Laing W (2005) Transient expression vectors for functional genomics, quantification of promoter activity and RNA silencing in plants. Plant Methods 1:13

    Article  PubMed  Google Scholar 

  • Hörtensteiner S (2006) Chlorophyll degradation during senescence. Annu Rev Plant Biol 57:55–77

    Article  PubMed  Google Scholar 

  • Hörtensteiner S (2009) Stay-green regulates chlorophyll and chlorophyll-binding protein degradation during senescence. Trends Plant Sci 14:155–162

    Article  PubMed  Google Scholar 

  • Kleber-Janke T, Krupinska K (1997) Isolation of cDNA clones for genes showing enhanced expression in barley leaves during dark-induced senescence as well as during senescence under field conditions. Planta 203:332–340

    Article  PubMed  CAS  Google Scholar 

  • Kusaba M, Ito H, Morita R, Iida S, Sato Y, Fujimoto M, Kawasaki S, Tanaka R, Hirochika H, Nishimura M, Tanaka A (2007) Rice NON-YELLOW COLORING1 is involved in light-harvesting complex II and grana degradation during leaf senescence. Plant Cell 19:1362–1375

    Article  PubMed  CAS  Google Scholar 

  • McGhie TK, Ainge GD (2002) Color in fruit of the genus Actinidia: carotenoid and chlorophyll compositions. J Agric Food Chem 50:117–121

    Article  PubMed  CAS  Google Scholar 

  • Montefiori M, McGhie TK, Hallett IC, Costa G (2009) Changes in pigments and plastid ultrastructure during ripening of green-fleshed and yellow-fleshed kiwifruit. Sci Hortic Amst 119:377–387

    Article  CAS  Google Scholar 

  • Morita R, Sato Y, Masuda Y, Nishimura M, Kusaba M (2009) Defect in non-yellow coloring 3, an α/β hydrolase-fold family protein, causes a stay-green phenotype during leaf senescence in rice. Plant J 59:940–952

    Article  PubMed  CAS  Google Scholar 

  • Oster U, Tanaka R, Tanaka A, Rüdiger W (2000) Cloning and functional expression of the gene encoding the key enzyme for chlorophyll b biosynthesis (CAO) from Arabidopsis thaliana. Plant J 21:305–310

    Article  PubMed  CAS  Google Scholar 

  • Park S-Y, Yu J-W, Park J-S, Li J, Yoo S-C, Lee N-Y, Lee S-K, Jeong S-W, Seo HS, Koh H-J, Jeon J-S, Park Y-I, Paek N-C (2007) The senescence-induced staygreen protein regulates chlorophyll degradation. Plant Cell 19:1649–1664

    Article  PubMed  CAS  Google Scholar 

  • Porra RJ, Thompson WA, Kriedemann PE (1989) Determination of accurate extinction coefficients and simultaneous equations for assaying chlorophylls a and b extracted with four different solvents: Verification of the concentration of chlorophyll standards by atomic absorption spectroscopy. BBA Bioenerg 975:384–394

    Article  CAS  Google Scholar 

  • Pružinská A, Tanner G, Anders I, Roca M, Hörtensteiner S (2003) Chlorophyll breakdown: pheophorbide a oxygenase is a Rieske-type iron–sulfur protein, encoded by the accelerated cell death 1 gene. Proc Natl Acad Sci USA 100:15259–15264

    Article  PubMed  Google Scholar 

  • Rodoni S, Muhlecker W, Anderl M, Krautler B, Moser D, Thomas H, Matile P, Hörtensteiner S (1997) Chlorophyll breakdown in senescent chloroplasts (cleavage of pheophorbide a in two enzymic steps). Plant Physiol 115:669–676

    Article  PubMed  CAS  Google Scholar 

  • Rozen S, Skaletsky HJ (1998) Primer3. Code available at http://www-genome.wi.mit.edu/genome_software/other/primer3.html. Accessed 9 Mar 2012

  • Rüdiger W (2002) Biosynthesis of chlorophyll b and the chlorophyll cycle. Photosynth Res 74:187–193

    Article  PubMed  Google Scholar 

  • Sato Y, Morita R, Nishimura M, Yamaguchi H, Kusaba M (2007) Mendel’s green cotyledon gene encodes a positive regulator of the chlorophyll-degrading pathway. Proc Natl Acad Sci USA 104:14169–14174

    Article  PubMed  CAS  Google Scholar 

  • Sato Y, Morita R, Katsuma S, Nishimura M, Tanaka A, Kusaba M (2009) Two short-chain dehydrogenase/reductases, non-yellow coloring 1 and nyc1-like, are required for chlorophyll b and light-harvesting complex II degradation during senescence in rice. Plant J 57:120–131

    Article  PubMed  CAS  Google Scholar 

  • Schelbert S, Aubry S, Burla B, Agne B, Kessler F, Krupinska K, Hörtensteiner S (2009) Pheophytin pheophorbide hydrolase (pheophytinase) is involved in chlorophyll breakdown during leaf senescence in Arabidopsis. Plant Cell 21:767–785

    Article  PubMed  CAS  Google Scholar 

  • Shabala S, Shabala L, Van Volkenburgh E, Newman I (2005) Effect of divalent cations on ion fluxes and leaf photochemistry in salinized barley leaves. J Exp Bot 56:1369–1378

    Article  PubMed  CAS  Google Scholar 

  • Tamura K, Dudley J, Nei M, Kumar S (2007) Mega4: molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol Biol Evol 24:1596–1599

    Article  PubMed  CAS  Google Scholar 

  • Thomas H, Howarth CJ (2000) Five ways to stay green. J Exp Bot 51(Suppl 1):329–337

    Article  PubMed  CAS  Google Scholar 

  • Tobin EM, Silverthorne J (1985) Light regulation of gene expression in higher plants. Annu Rev Plant Physiol 36:569–593

    Article  CAS  Google Scholar 

  • Voinnet O, Rivas S, Mestre P, Baulcombe D (2003) An enhanced transient expression system in plants based on suppression of gene silencing by the p19 protein of tomato bushy stunt virus. Plant J 33:949–956

    Article  PubMed  CAS  Google Scholar 

  • Warrington IJ, Weston GC (1990) Kiwifruit: science and management. New Zealand Society for Horticultural Science, Palmerston North

    Google Scholar 

  • Żelisko A, García-Lorenzo M, Jackowski G, Jansson S, Funk C (2005) AtFtsH6 is involved in the degradation of the light-harvesting complex II during high-light acclimation and senescence. Proc Natl Acad Sci USA 102:13699–13704

    Article  PubMed  Google Scholar 

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Acknowledgments

This research was funded by the New Zealand Foundation for Research Science and Technology (Horticultural Genomics C06X0812). We would like to thank the Tertiary Education Commission for an Enterprise Scholarship with Plant and Food Research, and Tim Holmes for photography.

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Authors and Affiliations

  1. The New Zealand Institute for Plant and Food Research Limited, Private Bag 92169, Auckland, New Zealand

    Sarah M. Pilkington, Mirco Montefiori & Andrew C. Allan

  2. University of Canterbury, Private Bag 4800, Christchurch, 8140, New Zealand

    Sarah M. Pilkington & Paula E. Jameson

  3. School of Biological Sciences, University of Auckland, Private Bag 92019, Auckland, New Zealand

    Andrew C. Allan

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  1. Sarah M. Pilkington
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  2. Mirco Montefiori
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  3. Paula E. Jameson
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  4. Andrew C. Allan
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Correspondence to Andrew C. Allan.

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Pilkington, S.M., Montefiori, M., Jameson, P.E. et al. The control of chlorophyll levels in maturing kiwifruit. Planta 236, 1615–1628 (2012). https://doi.org/10.1007/s00425-012-1723-x

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