The Protein Composition of the Digestive Fluid from the Venus Flytrap Sheds Light on Prey Digestion Mechanisms*

The Venus flytrap (Dionaea muscipula) is one of the most well-known carnivorous plants because of its unique ability to capture small animals, usually insects or spiders, through a unique snap-trapping mechanism. The animals are subsequently killed and digested so that the plants can assimilate nutrients, as they grow in mineral-deficient soils. We deep sequenced the cDNA from Dionaea traps to obtain transcript libraries, which were used in the mass spectrometry-based identification of the proteins secreted during digestion. The identified proteins consisted of peroxidases, nucleases, phosphatases, phospholipases, a glucanase, chitinases, and proteolytic enzymes, including four cysteine proteases, two aspartic proteases, and a serine carboxypeptidase. The majority of the most abundant proteins were categorized as pathogenesis-related proteins, suggesting that the plant's digestive system evolved from defense-related processes. This in-depth characterization of a highly specialized secreted fluid from a carnivorous plant provides new information about the plant's prey digestion mechanism and the evolutionary processes driving its defense pathways and nutrient acquisition.

[1]  Thomas F Dyrlund,et al.  MS Data Miner: A web‐based software tool to analyze, compare, and share mass spectrometry protein identifications , 2012, Proteomics.

[2]  R. Hedrich,et al.  Poplar Extrafloral Nectaries: Two Types, Two Strategies of Indirect Defenses against Herbivores1[C][W] , 2012, Plant Physiology.

[3]  Martin Vingron,et al.  Oases: robust de novo RNA-seq assembly across the dynamic range of expression levels , 2012, Bioinform..

[4]  W. Schulze,et al.  Nitrate and ammonium lead to distinct global dynamic phosphorylation patterns when resupplied to nitrogen-starved Arabidopsis seedlings , 2012, The Plant journal : for cell and molecular biology.

[5]  Jan C. Refsgaard,et al.  CrossWork: software-assisted identification of cross-linked peptides. , 2011, Journal of proteomics.

[6]  Axel Mithöfer,et al.  Carnivorous pitcher plants: insights in an old topic. , 2011, Phytochemistry.

[7]  Rainer Hedrich,et al.  A special pair of phytohormones controls excitability, slow closure, and external stomach formation in the Venus flytrap , 2011, Proceedings of the National Academy of Sciences.

[8]  M. Selbach,et al.  Global quantification of mammalian gene expression control , 2011, Nature.

[9]  A. Weber,et al.  Critical assessment of assembly strategies for non-model species mRNA-Seq data and application of next-generation sequencing to the comparison of C(3) and C(4) species. , 2011, Journal of experimental botany.

[10]  T. Isobe,et al.  A Cysteine Endopeptidase (“Dionain”) Is Involved in the Digestive Fluid of Dionaea muscipula (Venus’s Fly-trap) , 2011, Bioscience, biotechnology, and biochemistry.

[11]  D. Knox Proteases in blood-feeding nematodes and their potential as vaccine candidates. , 2011, Advances in experimental medicine and biology.

[12]  L. Leinwand,et al.  Whole transcriptome analysis of the fasting and fed Burmese python heart: insights into extreme physiological cardiac adaptation. , 2011, Physiological genomics.

[13]  T. Tokunaga,et al.  Trap‐Closing Chemical Factors of the Venus Flytrap (Dionaea muscipulla Ellis) , 2010, Chembiochem : a European journal of chemical biology.

[14]  Neil D. Rawlings,et al.  MEROPS: the peptidase database , 2009, Nucleic Acids Res..

[15]  Koji Matsumoto,et al.  Comparative studies on the acid proteinase activities in the digestive fluids of Nepenthes, Cephalotus, Dionaea, and Drosera , 2009, Carnivorous Plant Newsletter.

[16]  Michael K. Udvardi,et al.  Dissection of Symbiosis and Organ Development by Integrated Transcriptome Analysis of Lotus japonicus Mutant and Wild-Type Plants , 2009, PloS one.

[17]  D. Waller,et al.  Evolving Darwin's 'most wonderful' plant: ecological steps to a snap-trap. , 2009, The New phytologist.

[18]  A. Volkov,et al.  Electrical memory in Venus flytrap. , 2009, Bioelectrochemistry.

[19]  M. Gerstein,et al.  RNA-Seq: a revolutionary tool for transcriptomics , 2009, Nature Reviews Genetics.

[20]  W. Bielawski,et al.  Isolation and characterization of carboxypeptidase III from germinating triticale grains. , 2009, Acta biochimica et biophysica Sinica.

[21]  M. Mann,et al.  MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification , 2008, Nature Biotechnology.

[22]  Hanlee P. Ji,et al.  Next-generation DNA sequencing , 2008, Nature Biotechnology.

[23]  A. Weber,et al.  Low-coverage massively parallel pyrosequencing of cDNAs enables proteomics in non-model species: comparison of a species-specific database generated by pyrosequencing with databases from related species for proteome analysis of pea chloroplast envelopes. , 2008, Journal of biotechnology.

[24]  Tatsuro Hamada,et al.  Proteome analysis of pitcher fluid of the carnivorous plant Nepenthes alata. , 2008, Journal of proteome research.

[25]  L. Adamec Mineral nutrition of carnivorous plants: A review , 1997, The Botanical Review.

[26]  M. Mann,et al.  In-gel digestion for mass spectrometric characterization of proteins and proteomes , 2006, Nature Protocols.

[27]  Jan Dvorák,et al.  A Multienzyme Network Functions in Intestinal Protein Digestion by a Platyhelminth Parasite* , 2006, Journal of Biological Chemistry.

[28]  C. Pieterse,et al.  Significance of inducible defense-related proteins in infected plants. , 2006, Annual review of phytopathology.

[29]  A. Iwamatsu,et al.  Nepenthesin, a unique member of a novel subfamily of aspartic proteinases: enzymatic and structural characteristics. , 2005, Current protein & peptide science.

[30]  M. Mann,et al.  Exponentially Modified Protein Abundance Index (emPAI) for Estimation of Absolute Protein Amount in Proteomics by the Number of Sequenced Peptides per Protein*S , 2005, Molecular & Cellular Proteomics.

[31]  J. Enghild,et al.  The TSG-6 and IαI Interaction Promotes a Transesterification Cleaving the Protein-Glycosaminoglycan-Protein (PGP) Cross-link*[boxs] , 2005, Journal of Biological Chemistry.

[32]  L. Mahadevan,et al.  How the Venus flytrap snaps , 2005, Nature.

[33]  N. Goh,et al.  Carnivorous pitcher plant uses free radicals in the digestion of prey , 2004, Redox report : communications in free radical research.

[34]  T. Tokunaga,et al.  Mechanism of antifeedant activity of plumbagin, a compound concerning the chemical defense in carnivorous plant , 2004 .

[35]  Jan A. Delcour,et al.  Structural Basis for Inhibition of Aspergillus niger Xylanase by Triticum aestivum Xylanase Inhibitor-I* , 2004, Journal of Biological Chemistry.

[36]  A. Iwamatsu,et al.  Enzymic and structural characterization of nepenthesin, a unique member of a novel subfamily of aspartic proteinases. , 2004, The Biochemical journal.

[37]  Michael T. Stewart,et al.  Cathepsin L1, the Major Protease Involved in Liver Fluke (Fasciola hepatica) Virulence , 2004, Journal of Biological Chemistry.

[38]  A. Loukas,et al.  Digestive proteases of blood-feeding nematodes. , 2003, Trends in parasitology.

[39]  Ruey-Hua Lee,et al.  Molecular characterization of a senescence-associated gene encoding cysteine proteinase and its gene expression during leaf senescence in sweet potato. , 2002, Plant & cell physiology.

[40]  T. Kageyama Pepsinogens, progastricsins, and prochymosins: structure, function, evolution, and development , 2002, Cellular and Molecular Life Sciences CMLS.

[41]  E. Schulze,et al.  Quantification of insect nitrogen utilization by the venus fly trap Dionaea muscipula catching prey with highly variable isotope signatures. , 2001, Journal of experimental botany.

[42]  R. Laursen,et al.  Amino-acid sequence and glycan structures of cysteine proteases with proline specificity from ginger rhizome Zingiber officinale. , 2000, European journal of biochemistry.

[43]  D. N. Perkins,et al.  Probability‐based protein identification by searching sequence databases using mass spectrometry data , 1999, Electrophoresis.

[44]  T. Tranbarger,et al.  Structure and expression of a developmentally regulated cDNA encoding a cysteine protease (pseudotzain) from Douglas fir. , 1996, Gene.

[45]  A. Barrett Cellular Proteolysis An Overview , 1992, Annals of the New York Academy of Sciences.

[46]  E. Elstner,et al.  Oxidative protein modification as predigestive mechanism of the carnivorous plant Dionaea muscipula: an hypothesis based on in vitro experiments. , 1990, Free radical biology & medicine.

[47]  A. Bennett,et al.  Leaf Closure in the Venus Flytrap: An Acid Growth Response , 1982, Science.

[48]  A. F. Bury Analysis of protein and peptide mixtures , 1981 .

[49]  B. Juniper,et al.  THE SECRETORY CYCLE OF DIONAEA MUSCIPULA ELLIS IV. THE ENZYMOLOGY OF THE SECRETION , 1980 .

[50]  B. Juniper,et al.  THE SECRETORY CYCLE OF DIONAEA MUSCIPULA ELLIS , 1980 .

[51]  S. Williams,et al.  PREY CAPTURE AND FACTORS CONTROLLING TRAP NARROWING IN DIONAEA (DROSERACEAE) , 1977 .

[52]  J. Scala,et al.  Digestive Secretion of Dionaea muscipula (Venus's Flytrap). , 1969, Plant physiology.

[53]  S. Hoover,et al.  Effect of pH upon proteolysis by papain. , 1947, The Journal of biological chemistry.

[54]  Victims of the Venus Flytrap , 1935, Science.

[55]  C. Darwin Insectivorous plants, by Charles Darwin. , 1875 .