Plant paralog to viral movement protein that potentiates transport of mRNA into the phloem

Science

Volumn 283, Issue 5398, 1999, Pages 94-98

  Xoconostle Cázares, Beatriz ^a   Xiang, Yu ^a   Ruiz Medrano, Roberto ^a   Wang, Hong Li ^a   Monzer, Jan ^a   Yoo, Byung Chun ^a   McFarland, K C ^a   Franceschi, Vincent R ^a   Lucas, William J ^a  

^a NONE

Author keywords

[No Author keywords available]

Indexed keywords

MESSENGER RNA; VEGETABLE PROTEIN;

AMINO ACID SEQUENCE; ARTICLE; CUCUMBER; MICROINJECTION; NONHUMAN; NUCLEOTIDE SEQUENCE; PLANT GROWTH; PRIORITY JOURNAL; PROTEIN TRANSPORT; RNA METABOLISM; SIGNAL TRANSDUCTION;

AMINO ACID SEQUENCE; BIOLOGICAL TRANSPORT; CLONING, MOLECULAR; CUCUMIS SATIVUS; CUCURBITACEAE; MICROINJECTIONS; MOLECULAR SEQUENCE DATA; PLANT LEAVES; PLANT PROTEINS; PLANT ROOTS; PLANT STEMS; PLANT VIRAL MOVEMENT PROTEINS; REVERSE TRANSCRIPTASE POLYMERASE CHAIN REACTION; RNA, ANTISENSE; RNA, MESSENGER; RNA, PLANT; SIGNAL TRANSDUCTION; VIRAL PROTEINS;

CUCUMIS SATIVUS; CUCURBITA MAXIMA;

EID: 0038931833     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.283.5398.94     Document Type: Article

References (36)

1. K. Esau, The Phloem, vol. 5/2, Encyclopedia of Plant Anatomy (Borntraeger, Berlin, 1969).
2. D. B. Fisher, Y. Yu, M. S. B. Ku, Plant Physiol. 100, 1433 (1992).
3. W. J. Lucas, B. Ding, C. Van der Schoot, New Phytol. 125, 435 (1993).
4. S. Balachandran, Y. Xiang, C. Schobert, G. A. Thompson, W. J. Lucas, Proc. Natl. Acad. Sci. U.S.A. 94, 14150 (1997).
5. R. Tiedemann and U. Carstens-Behrens, J. Plant Physiol. 143, 189 (1994).
6. C. Kühn, V. R. Franceschi, A. Schultz, R. Lemoine, W. B. Frommer, Science 275, 1298 (1997).
7. W. J. Lucas and R. L. Gilbertson, Annu. Rev. Phytopathol. 32, 387 (1994); J. C. Carrington, K. D. Kasschau, S. K. Mahajan, M. C. Schaad, Plant Cell 8, 1669 (1996).
8. W. J. Lucas and R. L. Gilbertson, Annu. Rev. Phytopathol. 32, 387 (1994); J. C. Carrington, K. D. Kasschau, S. K. Mahajan, M. C. Schaad, Plant Cell 8, 1669 (1996).
9. T. Fujiwara, D. Giesman-Cookmeyer, B. Ding, S. A. Lommel, W. J. Lucas, Plant Cell 5, 1783 (1993).
10. A. Noueiry, W. J. Lucas, R. L. Gilbertson, Cell 76, 925 (1994); B. Ding, Q. Lin, L Nguyen, P. Palukaitis, W. J. Lucas, Virology 207, 345 (1995); M. R. Rojas, F. M. Zerbini, R. F. Allison, R. L. Gilbertson, W. J. Lucas, ibid. 237, 283 (1997).
11. A. Noueiry, W. J. Lucas, R. L. Gilbertson, Cell 76, 925 (1994); B. Ding, Q. Lin, L Nguyen, P. Palukaitis, W. J. Lucas, Virology 207, 345 (1995); M. R. Rojas, F. M. Zerbini, R. F. Allison, R. L. Gilbertson, W. J. Lucas, ibid. 237, 283 (1997).
12. A. Noueiry, W. J. Lucas, R. L. Gilbertson, Cell 76, 925 (1994); B. Ding, Q. Lin, L Nguyen, P. Palukaitis, W. J. Lucas, Virology 207, 345 (1995); M. R. Rojas, F. M. Zerbini, R. F. Allison, R. L. Gilbertson, W. J. Lucas, ibid. 237, 283 (1997).
13. W. J. Lucas et al., Science 270, 1980 (1995); T. J. Lough et al., Mol. Plant-Microbe Interact. 11, 801 (1998).
14. W. J. Lucas et al., Science 270, 1980 (1995); T. J. Lough et al., Mol. Plant-Microbe Interact. 11, 801 (1998).
15. R. A. Jorgensen, R. G. Atkinson, R. L. S. Forster, W. J. Lucas, Science 279, 1486 (1998).
16. S. A. Lommel, M. Weston-Fina, Z. Xiong, G. P. Lomonossoff, Nucleic Acids Res. 16, 8587 (1988).
17. 2-terminus. To clone CmPP16 by 3′ rapid amplification of cDNA ends (RACE)-PCR, a degenerate primer was designed -16A1 [5′-ATGGGIATGATGGARGTICA-3′ (where R is A or G)] - and oligo(dT) was used as the reverse primer. A pumpkin cDNA library was prepared from stem tissue excised from 2-month-old plants (17). Pumpkin stem cDNA was synthesized (Lambda ZAP II cDNA synthesis kit, Stratagene, La Jolla, CA) and used as a template for PCR. The first amplification was done with primer 16A1 and oligo(dT) under the following conditions: 1 min at 94°C, 1 min at 59°C, 2 min at 72°C (30 cycles). The resultant PCR fragments were subcloned into the pCR2.1 vector (Invitrogen, Carlsbad, CA) and sequenced. The clone corresponding to CmPP16 was used to screen the stem cDNA library.
18. J. Sambrook, E. F. Fritsch, T. Maniatis, Molecular Cloning (Cold Spring Harbor Laboratory Press, Plainview, NY, 1989).
19. See supplementary material (Fig. 1) at www. sciencemag.org/feature/data/982968.shl.
20. H. Hug and T. F. Sarre, Biochem. J. 291, 329 (1993); J. Kopka, C. Pical, A. M. Hetherington, B. Mueller- Rober, Plant Mol. Biol. 36, 627 (1998).
21. H. Hug and T. F. Sarre, Biochem. J. 291, 329 (1993); J. Kopka, C. Pical, A. M. Hetherington, B. Mueller-Rober, Plant Mol. Biol. 36, 627 (1998).
22. We used the side-grafting technique to graft the apical 10-cm region of 4-week-old cucumber plants onto 4-week-old pumpkin plants (5). Ten days later, we severed these scions (cut was 3 cm from graft union) and collected phloem sap from the basal end (4). We also collected phloem sap from the equivalent position on control cucumber plants. Aliquots (5 μl) of sap were mixed with an equal volume of 8 M guanidinium HCl and extracted twice with a 25:24:1 mixture of phenol/chloroform/isoamyl alcohol [J. Logemann, J. Schell, L. Willmitzer, Anal. Biochem. 163, 16 (1987)]. RNA was precipitated, centrifuged at 4°C for 45 min, and then resuspended in deionized sterile water. We isolated polyadenylated RNA from total RNA by using an oligotex mRNA extraction kit (Qiagen) and synthesized first-strand cDNA with Superscript reverse transcriptase (Gibco). We then used this cDNA as template for long-distance PCR (Clontech) according to the manufacturer's recommendations with a Robocycler PCR system (Stratagene). We used specific primers to amplify the CmPP16 cDNA according to the following protocol: 1 min at 94°C (one cycle); 30 s at 94°C, 30 s at 61°C, 70 s at 72°C (35 cycles). Preliminary RT-PCR experiments conducted on pumpkin and cucumber confirmed the specificity of the primers designed to amplify CmPP16. The amplified products were run on an agarose gel and visualized directly by ethidium bromide staining.
23. Preparation of fluorescent probes and microinjection procedures are described in (9, 10).
24. Preliminary experiments in which the biolistic bombardment method [A. Itaya et al., Plant J. 12, 1223 (1997)] was used to deliver various combinations of fluorescein isothiocyanate (FITC)-tabeled R-CmPP16-1 and RC-NMV FITC-MP with or without CF-labeled RNA yielded results consistent with our microinjection experiments.
25. J. Bollmann, R. Carpenter, E. S. Coen, Plant Cell 3, 1327 (1991); C. Künz, H. Schob, M. Stam, J. M. Kooter, F. Meins, Plant J. 10, 437 (1996); J.-C. Palauqui et al., Plant Physiol. 112, 1447 (1996); O. Voinnet and D. C. Baulcombe, Nature 389, 553 (1997); N. S. Al-Kaff et al. Science 279, 2113 (1998).
26. J. Bollmann, R. Carpenter, E. S. Coen, Plant Cell 3, 1327 (1991); C. Künz, H. Schob, M. Stam, J. M. Kooter, F. Meins, Plant J. 10, 437 (1996); J.-C. Palauqui et al., Plant Physiol. 112, 1447 (1996); O. Voinnet and D. C. Baulcombe, Nature 389, 553 (1997); N. S. Al-Kaff et al. Science 279, 2113 (1998).
27. J. Bollmann, R. Carpenter, E. S. Coen, Plant Cell 3, 1327 (1991); C. Künz, H. Schob, M. Stam, J. M. Kooter, F. Meins, Plant J. 10, 437 (1996); J.-C. Palauqui et al., Plant Physiol. 112, 1447 (1996); O. Voinnet and D. C. Baulcombe, Nature 389, 553 (1997); N. S. Al-Kaff et al. Science 279, 2113 (1998).
28. J. Bollmann, R. Carpenter, E. S. Coen, Plant Cell 3, 1327 (1991); C. Künz, H. Schob, M. Stam, J. M. Kooter, F. Meins, Plant J. 10, 437 (1996); J.-C. Palauqui et al., Plant Physiol. 112, 1447 (1996); O. Voinnet and D. C. Baulcombe, Nature 389, 553 (1997); N. S. Al-Kaff et al. Science 279, 2113 (1998).
29. J. Bollmann, R. Carpenter, E. S. Coen, Plant Cell 3, 1327 (1991); C. Künz, H. Schob, M. Stam, J. M. Kooter, F. Meins, Plant J. 10, 437 (1996); J.-C. Palauqui et al., Plant Physiol. 112, 1447 (1996); O. Voinnet and D. C. Baulcombe, Nature 389, 553 (1997); N. S. Al-Kaff et al. Science 279, 2113 (1998).
30. J.-C. Palauqui, T. Elmayan, J. M. Pollien, H. Vaucheret, EMBO J. 16, 4738 (1997).
31. A. Fire et al., Nature 391, 806 (1998).
32. Abbreviations for amino acid residues are as follows: A, Ala; C, Cys; D, Asp; E. Glu; F, Phe; C, Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q. Gln; R, Arg; S, Ser; T, Thr; V, Val; W, Trp; and Y., Tyr.
33. Polyclonal antibodies directed against recombinant CmPP16-1 and RCNMV MP were raised in rabbits; preimmune serum was tested against the antigens before the immunological scheme. Immunoglobulin G-purified antibody preparations were used in immunoblot analyses.
34. S. F. Altschul et al., Nucleic Acids Res 25, 3389 (1997).
35. Semithin sections (500 to 750 nm) of pumpkin stems and petioles were prepared and processed for CmPP16 immunolocalization by the procedures described by Wang et al. [H.-L. Wang, Y Wang, D. Giesman-Cookmeyer, S. A. Lommel, W. J. Lucas, Virology 245, 75 (1998)].
36. R.R.-M. was partially supported by a postdoctoral fellowship from CONACYT-México. Supported by Department of Energy Biosciences grant DE-FG03-94ER20134 and National Science Foundation grant IBN-94-06974 (W.J.L.). We thank the many colleagues whose comments and advice contributed to the development of this work. The RCNMV MP antibody used in initial screening studies was generously provided by S. Lommel, North Carolina State University. The plasmid containing the potato SUT1 was kindly provided by W. Frommer, University of Tübingen, Germany.