Abstract
Levels of two wound-inducible serine proteinase inhibitors, called Inhibitor I and Inhibitor II, and their mRNAs were quantified in leaves of tomato (Lycopersicon escululentum (L.) Mill.) plants after wounding the leaves with a hemostat. A single wound on a lower leaf of 25-old tomato plants caused the accumulation of the two inhibitor proteins in wounded and non-wounded leaves beginning about 4–6 h following wounding. The rate of inhibitor accumulation was maximal in leaves for the next 4 h and then declined. By 20 h the accumulation had nearly ceased. Following a single wound, Inhibitor I mRNA [600 bases in length] and Inhibitor II mRNA (760 bases) began to accumulate in wounded leaves about 2 h before the inhibitor proteins could be detected. The levels of mRNA for both inhibitors reached a maximum at about 8 h following wounding and then decayed, both with apparent half lives of about 10 h. Four consecutive wounds, inflicted hourly, increased the levels of mRNA for both inhibitors to over twice the levels induced by a single wound. Within 4 h following multiple wounds, Inhibitor I mRNA represented about 0.5% of the total polyadenylated mRNA (poly(A+)mRNA) and Inhibitor II mRNA about 0.15% of the total. The rates of accumulation of the two inhibitor proteins varied depending upon the age of the plants and their environment during growth, and ranged between 3 and 10 μg Inhibitor I·h-1·(g tissue)-1 for Inhibitor I and about half of these rates for Inhibitor II. Nuclei were isolated from leaves of wounded and non-wounded plants, and in mRNA runoff experiments using specific inhibitor copy DNAs (cDNAs) as probes the synthesis of Inhibitor I and II mRNAs were shown to be regulated, at least in part, at the level of transcription.
Similar content being viewed by others
Abbreviations
- cDNA:
-
copy DNA
- bp:
-
base pair(s)
- Mr :
-
molecular mass
- PIIF:
-
proteinase inhibitor inducing factor
- poly (A+)mRNA:
-
polyadenylated mRNA
References
Bishop, P., Pearce, G., Bryant, J.E., Ryan, C.A. (1984) Isolation and characterization of the proteinase inhibitor inducing factor from tomato leaves: identity and activity of polyand oligogalacturonide fragments. J. Biol. Chem. 259, 13172–13177
Birnboim, H.C., Doly, J. (1979) A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucl. Acids Res. 7, 1513–1523
Graham, J., Pearce, G., Merryweather, J., Titani, K., Ericsson, L., Ryan, C.A. (1985a) Wound-induced proteinase inhibitor mRNA from tomato leaves: I. The cDNA-deduced sequence of pre-Inhibitor I and its post-translational processing. J. Biol. Chem. 260, 6555–6560
Graham, J., Pearce, G., Merryweather, J., Titani, K., Ericsson, L., Ryan, C.A. (1985b) Wound-induced proteinase inhibitors from tomato leaves: II. The cDNA-deduced primary structure of pre-Inhibitor II. J Biol. Chem. 260, 6561–6564
Green, T.R., Ryan, C.A. (1972) Wound-induced proteinase inhibitor in plant leaves: A possible defense mechanism against insects. Science 175, 776–777
Green, T.R., Ryan, C.A. (1973) Wound-induced proteinase inhibitor in tomato leaves: Some effects of light and temperature on the wound response. Plant Physiol. 51, 19–21
Gustafson, G., Ryan, C.A. (1976) The specificity of protein turnover in tomato leaves: The accumulation of proteinase inhibitors, induced with the wound hormone, PIIF. J. Biol. Chem. 251, 7004–7010
Luthe, D.S., Quatrano, R.S. (1980) Transcription in isolated wheat nuclei: II. Characterization of RNA synthesized in-vitro. Plant Physiol. 65, 309–313
Maniatis, T., Fritsch, E.F., Sambrook, J. (1982) Molecular cloning. A laboratory manual. Cold Spring Laboratory, Cold Spring Harbor, N.Y., USA
Maxwell, I.H., Maxwell, F., Hahn, W.E. (1977) Removal of RNase activity from DNase by affinity chromatography on agarose-coupled aminophenyl-phosphoryl-uridine 2′(3′)-phosphate. Nucl. Acids Res. 4, 241–246
McKnight, G.S., Palmiter, R.D. (1979) Transcriptional regulation of the ovalbumin and conalbumin genes by steroid hormones in chick oviduct. J Biol. Chem. 254, 9050–9058
Nelson, C.E., Ryan, C.A. (1980) In-vitro synthesis of pre-proteins of two vacuolar compartmented proteinase inhibitors that accumulate in leaves of wounded tomato plants. Proc. Natl. Acad. Sci. USA 77, 1975–1979
Plunkett, G., Senear, D.F., Zuroske, G., Ryan, C.A. (1982) Proteinase Inhibitor I and II from leaves of wounded tomato plants: purification and properties. Arch. Biochem. Biophys. 213, 463–472
Ryan, C.A. (1967) The quantitative determination of soluble cellular proteins by radial diffusion in agar gels containing antibodies. Anal. Biochem. 19, 434–440
Ryan, C.A. (1974) Assay and biochemical properties of the proteinase inhibitor inducing factor, a wound hormone. Plant Physiol. 54, 328–332
Ryan, C.A. (1978) Proteinase inhibitors in plant leaves: A biochemical model for pest-induced natural plant protection. Trends Biochem. Sci 5, 148–150
Tratuman, R., Cowan, K.M., Wagner, G.G. (1971) Data for processing for radial immunodiffusion. Immunochemistry 8, 901–916
Walker-Simmons, M., Ryan, C.A. (1977) Immunological identification of proteinase Inhibitors I and II in isolated tomato leaf vacuoles. Plant Physiol. 60, 61–63
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Graham, J.S., Hall, G., Pearce, G. et al. Regulation of synthesis of proteinase inhibitors I and II mRNAs in leaves of wounded tomato plants. Planta 169, 399–405 (1986). https://doi.org/10.1007/BF00392137
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00392137