Identification, purification, and characterization of pathogenesis-related proteins from virus-infected Samsun NN tobacco leaves

Abstract

Ten pH-3 soluble, low-molecular-weight pathogenesis-related proteins (PRs) were found to accumulate in leaves of tobacco cv. Samsun NN reacting hypersensitively to tobacco mosaic virus. Besides the previously characterized PRs 1a, 1b, 1c and 2, these proteins were provisionally designated N, O, P, Q, R, and S in order of decreasing electrophoretic mobility in native polyacrylamide gels. Two-dimensional gel electrophoresis indicated that the PRs consist of single polypeptides, except for R, which is composed of two components with slightly different molecular weights. Estimated molecular weights in SDS-containing gels were: PRs 1a and 1b 17 kD, 1c 16.5 kD, 2 31 kD, N 33 kD, O 35 kD, P 27 kD, Q 28 kD, R 13 and 15 kD, and S 25 kD. However, based on their elution from gel filtration columns and relative moblities in native gels of different acrylamide concentrations, P and Q appeared to have molecular weights similar to those of the PR 1 group. Upon chromatofocusing no additional components were resolved. The PRs were eluted between pH 7 and 4; except for R, their pIs, as judged from isoelectric focusing, appeared to lie in the range from pH 4 to 5.2. In the presence of 6 M urea PR 1a was split into two components, one of which was strongly retarded on gels, as were P and Q. None of the PRs was detected when gels were stained for glycoproteins.By combinations of gel filtration, DEAE-cellulose chromatography, and chromatofocusing, PRs 1a, 1b, 1c, 2 and N were purified, their amino acid compositions determined, and antisera raised against each of these components. By Western blotting, antisera against either PR 1a, 1b, or 1c reacted with each of the components of the PR 1 group, as well as with PR S. Similarly, the antisera against either PR 2 or N reacted with both 2 and N, as well as with O and R. On the basis of major similarities in molecular weight characteristics, amino acid compositions, and serological relationships, it is proposed to classify tobacco PRs into five groups: 1: PRs 1a, 1b, and 1c; 2: 2a (formerly 2), 2b (N), and 2c (O); 3: 3a (P), and 3b (Q); 4: 4a and 4b (the two components of R); and 5: PR 5 (S).

Purification and Serological Characterization of Three Basic 15-Kilodalton Pathogenesis-Related Proteins from Tomato

“…In leaves of TMV-infected 'Samsun NN' and 'Xanthi-nc' tobacco, three serologically related, extracellular, acidic PR proteins (PR-la, -lb, and -lc) with similar molecular mass (15 kD) but different pl values have been identified (15,24,27). The primary structures of the three PR-1 proteins show a homology of approximately 90% (4).…”

The role of vacuolar and secreted pathogenesis-related ß (1–3)-glucanases and chitinases in the defence response of plants

“…For this, wc purificd and eharacterizcd the PR protcins obtaincd aftcr salieylic acid treatment of tobacco plants. As it has becomc clcar that within the PH protein fraction, both intra-and extracellular isoforms arc present, wc initially analyzed total protcin extracts, enriched for PR protcins hy autodigcstion of the leaf homogcnate (van Loon et al, 1987). The respective steps in the purification and charaetcrization of the PH proteins are outlined in figure 2. ln summary, the autodigestcd totalleaf protein extraets ar·c scparated by anion exchange ehromatography and the different fractions arc analyzed enzymatically for 6 (l-3)-glucanases, chitinases, and pcroxidasc aetivity and immunologically with an tisera directcd against 13 ( 1-3)-glueanases and ehitinases.…”

Cloning plant genes differentially during colonization of roots of Hordeum vulgare by the vesicular—arbuscular mycorrhizal fungus Glomus intraradices

“…GianinazziPearson ei al. (1990) compared the expression of the pathogenesis related protein gene, PR-b 1 (Van Loon, Gerritsen & Ritter, 1987) in tobacco roots infected by either Glomns mosseae or the root pathogen Chalara elegans. The results indicated that the accumulation of PR-b 1 mRNA transcripts was higher in mycorrhizal roots than in the nonmycorrhizal controls, but very low in comparison with the pathogen infected roots.…”