Free Salicylic Acid Levels Research Articles

Salicylic Acid Accumulation in Barley Is Pathogen Specific but Not Required for Defense-Gene Activation

Barley (Hordeum vulgare) seedlings were inoculated with the biotrophic pathogen Erysiphe graminis f. sp. hordei, the biotrophic nonhost pathogen E. graminis f. sp. tritici, and the necrotrophic nonhost pathogen Pseudomonas syringae pv. syringae. The levels of free salicylic acid and of salicylic-acid conjugates remained low after infection with E. graminis f. sp. hordei or E. graminis f. sp. tritici while they increased after inoculation with P. syringae pv. syringae. Pathogenesis-related proteins PR1, PR3 (chitinase), PR5 (thaumatin-like), and PR9 (peroxidase) accumulated after inoculation with all three pathogens.

Salicylic acid has a dual role in the activation of defence‐related genes in parsley

Systemic acquired resistance is an inducible plant defence state, the activation of which depends mostly on the accumulation of salicylic acid (SA). During the past several years, it has been demonstrated that pretreatment of cultured parsley cells with SA potentiates the elicitation of several defence responses that are local in whole plants, including the accumulation of phenylpropanoid products. Here it is reported that while anionic peroxidase and mannitol dehydrogenase encoding genes are directly responsive to SA, pretreating parsley cells with SA not only enhances elicitation of the phenylpropanoid genes phenylalanine ammonia-lyase and 4-coumarate:CoA ligase but also of genes for PR-10 and a hydroxyproline-rich glycoprotein. Enhanced induction of these genes was seen at low levels of endogenous free SA. Enhancement of phenylalanine ammonia-lyase gene activation was proportional to the length of SA pretreatment. Furthermore, the ability of SA analogues to both potentiate elicited and directly induce defence gene activation correlated with their biological activity to promote plant disease resistance. In summary, these results emphasize that SA has at least a dual role in plant defence gene activation.

Activation of Host Defense Mechanisms by Elevated Production of H2O2 in Transgenic Plants.

Active oxygen species have been postulated to perform multiple functions in plant defense, but their exact role in plant resistance to diseases is not fully understood. We have recently demonstrated H2O2-mediated disease resistance in transgenic potato (Solanum tuberosum) plants expressing a foreign gene encoding glucose oxidase. In this study we provide further evidence that the H2O2-mediated disease resistance in potato is effective against a broad range of plant pathogens. We have investigated mechanisms underlying the H2O2-mediated disease resistance in transgenic potato plants. The constitutively elevated levels of H2O2 induced the accumulation of total salicylic acid severalfold in the leaf tissue of transgenic plants, although no significant change was detected in the level of free salicylic acid. The mRNAs of two defense-related genes encoding the anionic peroxidase and acidic chitinase were also induced. In addition, an increased accumulation of several isoforms of extracellular peroxidase, including a newly induced one, was observed. This was accompanied by a significant increase in the lignin content of stem and root tissues of the transgenic plants. The results suggest that constitutively elevated sublethal levels of H2O2 are sufficient to activate an array of host defense mechanisms, and these defense mechanisms may be a major contributing factor to the H2O2-mediated disease resistance in transgenic plants.

Differential Accumulation of Salicylic Acid and Salicylic Acid-Sensitive Catalase in Different Rice Tissues

We previously proposed that salicylic acid (SA)-sensitive catalases serve as biological targets of SA in plant defense responses. To further examine the role of SA-sensitive catalases, we have analyzed the relationship between SA levels and SA sensitivity of catalases in different rice (Oryza sativa) tissues. We show here that, whereas rice shoots contain extremely high levels of free SA, as previously reported (I. Raskin, H. Skubatz, W. Tang, B.J.D. Meeuse [1990] Ann Bot 66: 369-373; P. Silverman, M. Seskar, D. Kanter, P. Schweizer, J.-P. Metraux, I. Raskin [1995] Plant Physiol 108: 633-639), rice roots and cell-suspension cultures have very low SA levels. Catalases from different rice tissues also exhibit differences in sensitivity to SA. Catalase from rice shoots is insensitive to SA, but roots and cell-suspension cultures contain SA-sensitive catalase. The difference in SA sensitivity of catalases from these different tissues correlates with the tissue-specific expression of two catalase genes, CatA and CatB, which encode highly distinctive catalase proteins. CatA, which encodes a catalase with relatively low sequence homology to the tobacco SA-sensitive catalases, is expressed at high levels exclusively in the shoots. On the other hand, in roots and cell-suspension cultures, with northern analysis we detected expression of only the CatB gene, which encodes a catalase with higher sequence homology to tobacco catalases. The role of catalases in mediating some of the SA-induced responses is discussed in light of these results and the recently defined mechanisms of catalase inhibition by SA.

Tobacco plants epigenetically suppressed in phenylalanine ammonia‐lyase expression do not develop systemic acquired resistance in response to infection by tobacco mosaic virus

The response of tobacco (Nicotiana tabacum L. cv. Xanthinc) plants, epigenetically suppressed for phenylalanine ammonia‐lyase (PAL) activity, was studied following infection by tobacco mosaic virus (TMV). These plants contain a bean PAL2 transgene in the sense orientation, and have reduced endogenous tobacco PAL mRNA and suppressed production of phenylpropanoid products. Lesions induced by TMV infection of PAL‐suppressed plants are markedly different in appearance from those induced on control plants that have lost the bean transgene through segregation, with a reduced deposition of phenofics. However, they develop at the same rate as on control tobacco, and pathogenesis‐related (PR) proteins are induced normally upon primary infection. The levels of free salicylic acid (SA) produced in primary inoculated leaves of PAL‐suppressed plants are approximately fourfold lower than in control plants after 84 h, and a similar reduction is observed in systemic leaves. PR proteins are not induced in systemic leaves of PAL‐suppressed plants, and secondary infection with TMV does not result in the restriction of lesion size and number seen in control plants undergoing systemic acquired resistance (SAR). In grafting experiments between wild‐type and PAL‐suppressed tobacco, the SAR response can be transmitted from a PAL‐suppressed root‐stock, but SAR is not observed if the scion is PAL‐suppressed. This indicates that, even if SA is the systemic signal for establishment of SAR, the amount of pre‐existing phenylpropanoid compounds in systemic leaves, or the ability to synthesize further phenylpropanoids in response to the systemic signal, may be important for the establishment of SAR. Treatment of PAL‐suppressed plants with dichloro‐isonicotinic acid (INA) induces PR protein expression and SAR against subsequent TMV infection. However, treatment with SA, while inducing PR proteins, only partially restores SAR, further suggesting that de novo synthesis of SA, and/or the presence or synthesis of other phenylpropanoids, is required for expression of resistance in systemic leaves.

Arachidonic Acid Induces Local but not Systemic Synthesis of Salicylic Acid and Confers Systemic Resistance in Potato Plants toPhytophthora infestansandAlternaria solani

When the lower leaves of young potato plants (cv. Bintje) were treated with an aqueous suspension of the polyunsaturated fatty acid arachidonic acid (AA), a threefold increase in the amount of free salicylic acid (SA) in the treated leaves was observed 3 h after treatment. The highest level of free SA was measured 6 h after treatment, reaching 3.21 g/g fresh weight compared to 0.15 g/g fresh weight in controls. The levels of conjugated SA in the treated leaves increased up to 51 g/g fresh weight 3 days after treatment compared to 0.83 g/g fresh weight in controls. Four days after treatment with AA the upper leaves became resistant to the potato late blight pathogen, Phytophthora infestans, but no accumulation of SA was observed. This systemic protection was observed in plants treated with AA at concentrations as low as 10 ppm and is unlikely to have arisen from AA translocated in the plant. In addition, treatment with AA protected potato plants against the early blight pathogen, Alternaria solani, indicating a broader spectrum of protection. Local but not systemic accumulation of a pathogenesis-related (PR)1-like protein was observed 24 h after treatment with AA. Application of SA on potato leaves did not confer resistance against P. infestans, but a local accumulation of a PR1-like protein was observed 24 h after treatment. Younger leaves of healthy susceptible and resistant potato plants contained higher levels of SA than older leaves, and this gradient correlated with their natural resistance to P. infestans. In general, cultivars showing field resistance contained higher amounts of conjugated SA than did susceptible cultivars.

Ortho-Anisic Acid as Internal Standard for the Simultaneous Quantitation of Salicylic Acid and Its Putative Biosynthetic Precursors in Cucumber Leaves

Salicylic acid (SA) acts as an endogenous signal for the induction of systemic acquired resistance (SAR) in plants infected by pathogens. In order to study SA biosynthesis in plants, a quantitative method using an internal standard was developed for the simultaneous measurement of SA and three of its putative bioprecursors, namely trans-cinnamic acid (CA), trans-ortho-hydroxycinnamic acid (oHCA), and benzoic acid (BA). ortho-Anisic acid (oANI) was found to be a suitable internal standard for all four compounds. It was not present as endogenous compound in cucumber leaves, no substance interfered with its detection, and it followed identical losses as SA, CA, oHCA, and BA during the extraction procedure. Baseline separation of these four compounds and of eight other plant phenolics was achieved in 35 min using deactivated reversed-phase HPLC. On-line detection was performed fluorimetrically for SA, oANI, and oHCA (with the excitation and emission wavelengths optimized for each compound) and with uv detection for CA and BA. Correlation equations for SA, CA, oHCA, and BA using oANI as internal standard were linear for a broad range of concentrations. The limits of detection (in ng/g fresh weight, FW) obtained during routine analyses were as follows: SA, 4; oHCA, 100; CA, 150; and BA, 500. The method was used to quantify levels of free and acid-hydrolyzed bound SA in cucumber plants either 5 days postinoculation with Pseudomonas lachrymans or after mock-inoculation with water on their first leaf. Free SA increased 33-fold locally (to 382.0 ng/g FW) in the infected leaf and 4.2-fold systemically (to 21.3 ng/g FW) in the second leaf compared to controls. Bound SA reached 4399 and 287.3 ng/g FW in the first and second leaves of infected plants, respectively (not detectable in controls).

Induction of Benzoic Acid 2-Hydroxylase in Virus-Inoculated Tobacco.

Salicylic acid (SA) plays an important role in the induction of plant resistance to pathogens. An accompanying article (N. Yalpani, J. Leon, M.A. Lawton, I. Raskin [1993] Plant Physiol 103: 315-321) shows that SA is synthesized via the decarboxylation of cinnamic acid to benzoic acid (BA), which is, in turn, hydroxylated to SA. Leaf extracts of tobacco (Nicotiana tabacum L. cv Xanthi-nc) catalyze the 2-hydroxylation of BA to SA. The monooxygenase catalyzing this reaction, benzoic acid 2-hydroxylase (BA2H), required NAD(P)H or reduced methyl viologen as an electron donor. BA2H activity was detected in healthy tobacco leaf extracts (1-2 nmol h-1 g-1 fresh weight) and was significantly increased upon inoculation with tobacco mosaic virus (TMV). This increase paralleled the levels of free SA in the leaves. Induction of BA2H activity was restricted to tissue expressing a hypersensitive response at 24[deg]C. TMV induction of BA2H activity and SA accumulation were inhibited when inoculated tobacco plants were incubated at 32[deg]C. However, when inoculated plants were incubated for 4 d at 32[deg]C and then transferred to 24[deg]C, they showed a 15-fold increase in BA2H activity and a 65-fold increase in free SA content compared with healthy plants incubated at 24[deg]C. Treatment of leaf tissue with the protein synthesis inhibitor cycloheximide blocked the induction of BA2H activity by TMV. The effect of TMV inoculation on BA2H could be duplicated by infiltrating leaf discs of healthy plants with BA. This response was observed even when applied levels of BA were much lower than the levels observed in vivo after virus inoculation. Feeding tobacco leaves with phenylalanine, cinnamic acid, or o-coumaric acid (putative precursors of SA) failed to trigger the induction of BA2H activity. BA2H appears to be a pathogen-inducible protein with an important regulatory role in SA accumulation during the development of induced resistance to TMV in tobacco.

Temperature-Dependent Induction of Salicylic Acid and Its Conjugates during the Resistance Response to Tobacco Mosaic Virus Infection.

Increases in endogenous salicylic acid (SA) levels and induction of several families of pathogenesis-related genes (PR-1 through PR-5) occur during the resistance response of tobacco to tobacco mosaic virus infection. We found that at temperatures that prevent the induction of PR genes and resistance, the increases in SA levels were eliminated. The addition of exogenous SA to infected plants at these temperatures was sufficient to induce the PR genes but not the hypersensitive response. However, when the resistance response was restored by shifting infected plants to permissive temperatures, SA levels increased dramatically and preceded PR-1 gene expression and necrotic lesion formation associated with resistance. SA was also found in a conjugated form whose levels increased in parallel with the free SA levels. The majority of the conjugates appeared to be SA glucosides. The same glucoside was formed when plants were supplied with exogenous SA. These results provide further evidence that endogenous SA signals the induction of certain defense responses and suggests additional complexity in the modulation of this signal.

Alteration in metabolism of acetylsalicylic acid in children with Down's syndrome: decreased plasma binding and formation of salicyluric acid.

Extract: This paper reports the differences in absorption and distribution in plasma of free and bound salicylate, and the differences in excretion of salicylic and salicyuric acid in the urine of children with Down's syndrome and in control subjects. After oral administration of a single dose of acetylsalicylic acid (35 mg/kg) to normal children, the peak level of salicylic acid in plasma occurred in 2 h and the drug level returned to the base line at approximately 8 h; in children with Down's syndrome, th peak level of salicylic acid occurred in 4 h and the drug level returned to a base line at a approximately 16 h. In children with Down's syndrome, the 24-h excretion of salicylate metabolites in the urine indicated a higher level of free salicylic acid (table I) and a lower level of salicyluric acid (tables II and III) when compared with the control group. Although the concentration of salicylate is small in erythrocytes, children with Down's syndrome showed a higher level when compared with their normal counter part (table IV). The in vitro plasma-binding studies indicated that in comparison with normal subjects, children with Down's syndrome were less capable of binding salicylate (table V). Speculation: Patients with Down's syndrome are a highly abnormal group of individuals because of their morphological, physiological and biochemical aberrations. They are directed and influenced by their abnormalities, one manifestation of which is altered pharmacogenetics. It should be expected that any therapeutic regime must take these differences into consideration and dosage must be calculated not on the basis of body weight but according to the plasma level of the drug or of its active metabolites.