Olfaction in Parkin single and compound heterozygotes in a cohort of young onset Parkinson's disease patients.

BACKGROUND: Parkin related Parkinson's disease (PD) is differentiated from idiopathic PD by absent or sparse Lewy bodies, and preserved olfaction. The significance of single Parkin mutations in the pathogenesis of PD is debated. OBJECTIVES: To assess olfaction results according to Parkin mutation status. To compare the prevalence of Parkin single heterozygous mutations in patients diagnosed with PD to the rate in healthy controls in order to establish whether these single mutations could be a risk factor for developing PD. METHODS: Parkin gene mutation testing was performed in young onset PD (diagnosed <50 years old) to identify three groups: Parkin homozygous or compound heterozygote mutation carriers, Parkin single heterozygote mutation carriers, and non-carriers of Parkin mutations. Olfaction was tested using the 40-item British version of the University of Pennsylvania smell identification test (UPSIT). RESULTS: Of 344 young onset PD cases tested, 8 (2.3%) were Parkin compound heterozygotes and 13 (3.8%) were Parkin single heterozygotes. Olfaction results were available in 282 cases (eight compound heterozygotes, nine single heterozygotes, and 265 non-carriers). In Parkin compound heterozygotes, the median UPSIT score was 33, interquartile range (IQR) 28.5-36.5, which was significantly better than in single Parkin heterozygotes (median 19, IQR 18-28) and non-carriers (median score 22, IQR 16-28) (ANOVA P < 0.001). These differences persisted after adjusting for age, disease duration, gender, and smoking (P < 0.001). There was no significant difference in UPSIT scores between single heterozygotes and non-carriers (P = 0.90). CONCLUSIONS: Patients with Parkin compound heterozygous mutations have relatively preserved olfaction compared to Parkin single heterozygotes and non-carriers. The prevalence of Parkin single heterozygosity is similar to the 3.7% rate reported in healthy controls.

Transcriptional activation of plant defense genes by fungal elicitor, wounding, and infection.

Activation of plant defense genes was investigated by analysis of transcripts completed in vitro by isolated nuclei. Elicitor treatment of suspension-cultured bean (Phaseolus vulgaris L.) cells caused marked transient stimulation of transcription of genes encoding apoproteins of cell wall hydroxyproline-rich glycoproteins (HRGP) and the phenylpropanoid biosynthetic enzymes phenylalanine ammonia-lyase (PAL) and chalcone synthase (CHS), concomitant with the onset of rapid accumulation of the respective mRNAs and hence expression of the phytoalexin (PAL, CHS), lignin (PAL), and HRGP defense responses. While there was a lag of 2 h prior to stimulation of HRGP gene transcription, induction of the transcription of PAL and CHS genes occurred within 5 min of elicitor treatment. Induction of transcription of PAL, CHS, and HRGP genes was also observed in wounded hypocotyls and in infected hypocotyls during race-cultivar-specific interactions with the fungus Colletotrichum lindemuthianum, the causal agent of anthracnose. Transcriptional activation occurred not only in directly infected tissue but also in distant, hitherto uninfected tissue, indicating intercellular transmission of an endogenous signal for defense gene activation. It is concluded that transcriptional activation of defense genes characteristically underlies induction of the corresponding defense responses and expression of disease resistance.

Density labelling characterisation of the effects of cordycepin and cycloheximide on the turnover of phenylalanine ammonia-lyase.

L-Phenylalanine ammonia-lyase (EC 4.3.1.5) undergoes a transient increase in activity in illuminated disc of Solanum tuberosum tuber parenchyme. Cycloheximide and cordycepin inhibit the initial increase in enzyme activity, but if addition of these anti-metabolites is delayed until the time of maximum enzyme levels, the subsequent decline in enzyme activity is inhibited (Lamb, C.J. (1977) Planta, 135, 169-175). The effect of delayed treatment with cycloheximide or cordycepin on the turnover of phenylalanine ammonia-lyase has been analysed by density labelling with 2H from 2H2O. Delayed introduction of cycloheximide or cordycepin reduces the rate of labelling of phenylalanine ammonia-lyase whilst preventing the decay in enzyme activity observed in controls not treated with inhibitor, and this labelling pattern cannot be accounted for by effects of cycloheximide or cordecypin on the labelling of amino acid pools. It is concluded that delayed treatment with cycloheximide or cordycepin leads to the maintenance of high levels of phenylalanine ammonia-lyase by inhibition of the removal of active enzyme rather than by maintenance of high rates of enzyme synthesis.

Elicitor modulation of the turnover of L-phenylalanine ammonia-lyase in French bean cell suspension cultures.

(1) The mechanisms underlying the transient increase in phenylalanine ammonia-lyase activity during phaseollin accumulation in cell suspension cultures of Dwarf French bean (Phaseolus volgaris) have been investigated using density labelling with 3H from 2H2O coupled with residual analysis of the equilibrium distribution of enzyme activity in high-resolution KBr density gradients. (2) The resolution achieved in this system is sufficient to allow quantitative analysis of the relative proportions of light, unlabelled, pre-existing enzyme and heavy, labelled, newly synthesised enzyme. (3) Elicitor released by heat treatment of cell walls of Colletotrichum lindemuthianum, the causal agent of anthracnose disease of French bean, caused a marked but transient increase in phenylalanine ammonia-lyase activity concomitant with the onset of phaseollin accumulation in the bean cultures. The induction of enzyme activity was highly dependent on elicitor concentration, with maximum induction occurring in two discrete concentration ranges; at an intermediate elicitor concentration, or at supra-optimal elicitor concentrations, no enzyme induction was observed. (4) At low concentrations of elicitor the induction of enzyme was entirely a result of elicitor stimulation of the rate of de novo enzyme production. In contrast, at higher elicitor concentrations the increase in enzyme activity was accompanied by a marked apparent stabilization of the enzyme in vivo, and the rapid but transient increase in enzyme activity was achieved by a programme of reciprocal changes in the rate constant for de novo enzyme production and the rate constant for removal of enzyme activity. Such reciprocal control of the rates of enzyme production and removal may be crucial in determining the magnitude and duration of the phytoalexin defense response. (5) Information on the specific activity of 2H label in the amino acid pools was obtained from analysis of the equilibrium distribution of residual, labelled activity. This showed directly that the turnover of the amino acid pools was fast relative to the turnover of phenylalanine ammonia-lyase and, hence, the rate of enzyme labelling was not limited by the rate of labelling of the amino acid pools. Elicitor treatment had no effect on the specific activity of label in the amino acid pools from which phenylalanine ammonia-lyase is synthesised.

Hydrogen Peroxide Stimulates Salicylic Acid Biosynthesis in Tobacco.

Hydrogen peroxide induced the accumulation of free benzoic acid (BA) and salicylic acid (SA) in tobacco (Nicotiana tabacum L. cv Xanthi-nc) leaves. Six hours after infiltration with 300 mM H2O2, the levels of BA and SA in leaves increased 5-fold over the levels detected in control leaves. The accumulation of BA and SA was preceded by the rapid activation of benzoic acid 2-hydroxylase (BA2H) in the H2O2-infiltrated tissues. This enzyme catalyzes the formation of SA from BA. Enzyme activation could be reproduced in vitro by addition of H2O2 or cumene hydroperoxide to the assay mixture. H2O2 was most effective in vitro when applied at 6 mM. In vitro activation of BA2H by peroxides was inhibited by the catalase inhibitor 3-amino-1,2,4-triazole. We suggest that H2O2 activates SA biosynthesis via two mechanisms. First, H2O2 stimulates BA2H activity directly or via the formation of its substrate, molecular oxygen, in a catalase-mediated reaction. Second, higher BA levels induce the accumulation of BA2H protein in the cells and provide more substrate for this enzyme.

Pathway of Salicylic Acid Biosynthesis in Healthy and Virus-Inoculated Tobacco.

Salicylic acid (SA) is a likely endogenous regulator of localized and systemic disease resistance in plants. During the hypersensitive response of Nicotiana tabacum L. cv Xanthi-nc to tobacco mosaic virus (TMV), SA levels rise dramatically. We studied SA biosynthesis in healthy and TMV-inoculated tobacco by monitoring the levels of SA and its likely precursors in extracts of leaves and cell suspensions. In TMV-inoculated leaves, stimulation of SA accumulation is accompanied by a corresponding increase in the levels of benzoic acid. 14C-Tracer studies with cell suspensions and mock-or TMV-inoculated leaves indicate that the label moves from trans-cinnamic acid to SA via benzoic acid. In healthy and TMV-inoculated tobacco leaves, benzoic acid induced SA accumulation. o-Coumaric acid, which was previously reported as a possible precursor of SA in other species, did not increase SA levels in tobacco. In healthy tobacco tissue, the specific activity of newly formed SA was equal to that of the supplied [14C]benzoic acid, whereas in TMV-inoculated leaves some isotope dilution was observed, presumably because of the increase in the pool of endogenous benzoic acid. We observed accumulation of pathogen-esis-related-1 proteins and increased resistance to TMV in benzoic acid- but not in o-coumaric acid-treated tobacco leaves. This is consistent with benzoic acid being the immediate precursor of SA. We conclude that in healthy and virus-inoculated tobacco, SA is formed from cinnamic acid via benzoic acid.

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.

Radiofrequency ablation for the fast pathway of atrioventricular node reentry. Evidence for partial damage to the fast and lower common pathways.

Radiofrequency ablation was attempted in a 30-year-old woman with atrioventricular (AV) node reentry of the common variety. Energy was delivered to ablate a fast pathway. After energy delivery, the atrio-His interval prolonged following transient AV node block. Retrograde conduction was no longer present. However, dual AV nodal physiology and AV node reentry with similar retrograde atrial activation sequence could be shown post-ablation. These observations suggest that both fast and lower common pathways were partially damaged by ablation.

Direct in vivo visualization of right cardiac anatomy by fiberoptic endoscopy. Hemodynamic effects and image validation.

The authors tested the ability of a balloon-tipped fiberoptic endoscope to accurately visualize and identify right-heart anatomy in 7 anesthetized dogs. A 3.6-mm-diameter fiberoptic endoscope with a latex balloon covering the distal tip was inserted into the right atrium, where the balloon was inflated with air in 5 mL increments. Heart rate did not show changes. Mean arterial pressure and cardiac output started to show significant decreases with a balloon volume at 25 and 20 mL, respectively (n = 7). Visual image quality was excellent with a balloon volume of 10 mL or greater. With a balloon volume of 7-10 mL, the visual field was 15-20 mm in diameter. Right-heart anatomy including the right free wall, ostium of the coronary sinus, atrioventricular node area, tricuspid valve, right ventricular structures, and pulmonary arteries was clearly and accurately identified. Additionally, spatial relationships among these structures could be established. Furthermore, there was an excellent concordance between endoscopically observed images and postmortem findings. In conclusion, balloon-tipped fiberoptic endoscopy can accurately visualize normal intracardiac structures with no or minimal hemodynamic compromise.

Glutathione causes a massive and selective induction of plant defense genes.

The reduced form of glutathione (GSH), when supplied to suspension cultured cells of bean (Phaseolus vulgaris L.) at concentrations in the range 0.01 to 1.0 millimolar, stimulates transcription of defense genes including those that encode cell wall hydroxyproline-rich glycoproteins and the phenylpropanoid biosynthetic enzymes phenylalanine ammonialyase (PAL) and chalcone synthase (CHS) involved in lignin (PAL) and phytoalexin (PAL, CHS) production. Transcriptional activation of these genes leads to marked accumulation of the corresponding transcripts, contributing to a massive change in the overall pattern of protein synthesis which closely resembles that previously observed in response to fungal elicitor. GSH causes a marked increase in extractable PAL activity, whereas the oxidized form of glutathione, constituent amino acids, or other reducing agents are inactive. Possible roles of GSH in signaling biological stress are discussed.

Glutathione-elicited changes in chromatin structure within the promoter of the defense gene chalcone synthase.

Sites hypersensitive to digestion by DNase I have been identified within the 5'-flanking and 3'-coding sequences of genes encoding the defense enzyme chalcone synthase in bean (Phaseolus vulgaris L.). Two of the 5'-flanking hypersensitive sites are markedly induced upon elicitation of cells with glutathione and delineate sequence elements that are also present in the promoters of coordinately regulated genes. In contrast, other hypersensitive sites within the 5'-flanking sequences are expressed constitutively and one maps within an element that is also present in the promoters of coordinately regulated genes. These results suggest that the transcriptional activation of chalcone synthase genes is accompanied by structural changes in the chromatin associated with the proximal region of the promoter and that these probably reflect the binding of transcription factors tocis-regulatory elements.

Direct in vivo visualization of right cardiac anatomy by fibreoptic endoscopy: observation of radiofrequency-induced acute lesions around the ostium of the coronary sinus.

Current mapping techniques used during electrophysiological study involve catheter placement under fluoroscopic guidance and are associated with prolonged radiation exposure. We considered that direct visualization of right heart anatomy by means of fibreoptic endoscopy could be useful in accurately localizing and guiding the ablation of arrhythmogenic substrates. Our goal was to evaluate the ability of this device to safely and accurately visualize the ostium of the coronary sinus and its vicinity as well as radiofrequency-induced acute lesions. Anaesthetized dogs (n = 4) were studied. Multipolar electrode catheters and a 3.6 mm diameter fibreoptic endoscope with a latex balloon covering the distal tip were inserted into the right atrium. The blood pressure, and surface and intracardiac electrocardiograms were recorded simultaneously. Radiofrequency energy was delivered just inside the coronary sinus ostium or in its vicinity. The acute lesions were carefully observed by endoscopy. Postmortem examination was then performed. With a balloon volume of 7-10 ml, the visual field was 15-20 mm in diameter. The blood pressure was generally stable. The ostium of the coronary sinus and its vicinity were clearly and accurately identified, and catheter placement in the coronary sinus and its vicinity could be achieved under direct vision in all four dogs. Additionally, the process by which acute lesions were created by radiofrequency was also directly visualized in great detail. There was a good concordance between endoscopically observed images and postmortem findings, and there was a strong correlation between delivered energy and the volume of the radiofrequency-induced lesions (r = 0.895).(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of a nuclear protein that binds to three elements within the silencer region of a bean chalcone synthase gene promoter.

The chalcone synthase (EC 2.3.1.74) gene promoter from the bean Phaseolus vulgaris L. contains a silencer element between positions -140 and -326 fro the transcription start site that is functional in electroporated soybean protoplasts. This element contains three binding sites for a bean nuclear factor (SBF-1) with DNA sequence recognition properties that are very similar to those of nuclear factor GT-1. By using a synthetic tetramer of one of the binding sites as probe, we have purified sequence-specific SBF-1 activity approximately 1750-fold from suspension-cell nuclei, by using a combination of ammonium sulfate precipitation, gel filtration, heparin-agarose chromatography, and sequence-specific DNA affinity chromatography. The factor exhibited an apparent molecular weight of 160,000-200,000 on the basis of gel filtration. A subunit molecular weight of approximately 95,000 was determined from SDS/polyacrylamide gel electrophoretic analysis of purified fractions, followed by Southwestern blot analysis (a protein blot probed with oligonucleotide probes), and from UV-cross-linking experiments. The factor lost DNA-binding activity on treatment with alkaline phosphatase. We discuss the properties of SBF-1 in relation to the functionality of GT-1 binding sequences in plant genes.

Glutathione and fungal elicitor regulation of a plant defense gene promoter in electroporated protoplasts.

To investigate the mechanisms underlying activation of plant defenses against microbial attack we have studied elicitor regulation of a chimeric gene comprising the 5' flanking region of a defense gene encoding the phytoalexin biosynthetic enzyme chalcone synthase fused to a bacterial chloramphenicol acetyltransferase gene. Glutathione or fungal elicitor caused a rapid, marked but transient expression of the chimeric gene electroporated into soybean protoplasts. The response closely resembled that of endogenous chalcone synthase genes in suspension cultured cells. Functional analysis of 5' deletions suggests that promoter activity is determined by an elicitor-regulated activator located between the "TATA box" and nucleotide position -173 and an upstream silencer between -173 and -326. These cis-acting elements function in the transduction of the elicitation signal to initiate elaboration of an inducible defense response.

Hydroxyproline-Rich Glycoprotein Transcripts Exhibit Different Spatial Patterns of Accumulation in Compatible and Incompatible Interactions between Phaseolus vulgaris and Colletotrichum lindemuthianum.

The distribution of transcripts encoding hydroxyproline-rich glycoproteins in hypocotyls of Phaseolus vulgaris L. infected with Colletotrichum lindemuthianum was examined by in situ hybridization to tissue sections. The expression of hypersensitive resistance in an incompatible interaction was accompanied by a massive early accumulation of transcripts in the epidermal, cortical, and perivascular parenchymal tissues immediately adjacent to the inoculation site. In a compatible interaction, there was no accumulation of transcripts in the epidermal and cortical tissues even though fungal hyphae ramified throughout these tissues. However, transcripts accumulated at a later stage in the perivascular tissue directly below the site of infection and in tissue several millimeters from the inoculation site. Thus, there is a spatial and tissue-specific counterpart to the differential timing of transcript accumulation in incompatible versus compatible interactions (AM Showalter, JN Bell, CL Cramer, JA Bailey, CJ Lamb [1985] Proc Natl Acad Sci USA 82: 6551-6555). These differences in the spatial distribution and tissue specificity of transcript accumulation imply the differential induction of signaling systems involved in race:cultivar-specific interactions.

atpk1, a novel ribosomal protein kinase gene from Arabidopsis. II. Functional and biochemical analysis of the encoded protein.

The Arabidopsis Atpk1 protein expressed in insect cells and plant cells exhibited multiple sizes consisting mainly of two doublets: p70 (68 and 70 kDa) and p85 (82 and 85 kDa). Extraction of p85 from cells required the presence of SDS, suggesting that p85 is associated with less soluble subcellular components. p70 was extracted by nonionic detergent without SDS, indicating that this form is cytoplasmic. p70 expressed in either Arabidopsis or insect cells underwent serine-specific autophosphorylation, indicating that Atpk1 is a protein-serine kinase. A point mutation (lysine 163 to arginine) in the ATP-binding site of the catalytic domain substantially diminished activity when expressed in insect cells. A 14-kDa protein (p14) was co-immunoprecipitated with p70 from insect cells expressing wild-type Atpk1 and was phosphorylated in immune complex kinase assays with Atpk1, suggesting it is a homolog of a natural substrate of Atpk1. Two plant ribosomal proteins (14 and 16 kDa) can be phosphorylated by the Atpk1 protein kinase, and we propose that Atpk1 is a novel ribosomal protein kinase. A 60-kDa form of Atpk1 derived from the insect cell-expressed p70 was more highly phosphorylated than p70 in in vitro kinase assays, suggesting a negative regulatory domain can be removed by proteolysis.

atpk1, a novel ribosomal protein kinase gene from Arabidopsis. I. Isolation, characterization, and expression.

Two protein kinase genes (atpk1 and atpk2) were isolated from Arabidopsis thaliana genomic DNA with a probe generated by polymerase chain reaction (PCR) using oligonucleotide primers encoding conserved eukaryotic protein kinase sequences. atpk1 and atpk2 are organized in a head-to-tail tandem array on chromosome 3 and have about 80% nucleotide sequence identity. atpk1 encodes a hydrophilic polypeptide of 465 amino acids, M(r) = 52,554. The centrally located catalytic domain contains all the conserved residues characteristic of eukaryotic protein kinases, with greatest similarity to the catalytic domains of 70-kDa ribosomal S6 protein kinase, protein kinase C, and protein kinase A. The C-terminal 75 residues also show homology to protein kinase C and S6 protein kinase. In contrast, the N-terminal 130 residues have no homology to any known protein, and thus may represent a new class of protein kinase regulatory domain. Other motifs found in the Atpk1 protein include two putative autophosphorylation sites, a pseudosubstrate site, two acidic domains, a lysine-rich domain, and two putative PEST sequences, which may contribute to the regulation of protein kinase activity. RNA-blot hybridization showed that atpk1 encoded a 1.8-kb mRNA. Analysis of atpk1 promoter/beta-glucuronidase reporter gene fusions in transgenic plants showed that atpk1 was expressed in all tissues and at all developmental stages, with the strongest expression observed in metabolically active tissues, suggesting that atpk1 is involved in the control of plant growth and development. The first intron of atpk1 functions as an enhancer in atpk1 expression.

Molecular cloning of plant transcripts encoding protein kinase homologs.

Oligonucleotides, corresponding to conserved regions of animal protein-serine/threonine kinases, were used to isolate cDNAs encoding plant homologs in the dicot bean (Phaseolus vulgaris L.) and the monocot rice (Oryzae sativa L.). The C-terminal regions of the deduced polypeptides encoded by the bean (PVPK-1) and rice (G11A) cDNAs, prepared from mRNAs of suspension cultures and leaves, respectively, contain features characteristic of the catalytic domains of eukaryotic protein-serine/threonine kinases, indicating that these cDNAs encode plant protein kinases. The putative catalytic domains are most closely related to cyclic nucleotide-dependent protein kinases and the protein kinase C family, suggesting the plant homologs may likewise transduce extracellular signals. However, outside these domains, PVPK-1 and G11A exhibit no homology either to each other or to regulatory domains of other protein kinases, indicating the plant homologs are modulated by other signals. PVPK-1 corresponds to a 2.4-kb transcript in suspension cultured bean cells. Southern blots of genomic DNA indicate that PVPK-1 and G11A correspond to single copy genes that form part of a family of related plant sequences.

Benzoic acid 2-hydroxylase, a soluble oxygenase from tobacco, catalyzes salicylic acid biosynthesis.

Benzoic acid 2-hydroxylase (BA2H) catalyzes the biosynthesis of salicylic acid from benzoic acid. The enzyme has been partially purified and characterized as a soluble protein of 160 kDa. High-efficiency in vivo labeling of salicylic acid with 18O2 suggested that BA2H is an oxygenase that specifically hydroxylates the ortho position of benzoic acid. The enzyme was strongly induced by either tobacco mosaic virus inoculation or benzoic acid infiltration of tobacco leaves and it was inhibited by CO and other inhibitors of cytochrome P450 hydroxylases. The BA2H activity was immunodepleted by antibodies raised against SU2, a soluble cytochrome P450 from Streptomyces griseolus. The anti-SU2 antibodies immunoprecipitated a radiolabeled polypeptide of around 160 kDa from the soluble protein extracts of L-[35S]-methionine-fed tobacco leaves. Purified BA2H showed CO-difference spectra with a maximum at 457 nm. These data suggest that BA2H belongs to a novel class of soluble, high molecular weight cytochrome P450 enzymes.

Cloning and properties of a rice gene encoding phenylalanine ammonia-lyase.

Phenylalanine ammonia-lyase (PAL) genomic sequences were isolated from a rice (Oryza sativa L.) genomic library using a PCR-amplified rice PAL DNA fragment as a probe. There is a small family of PAL genes in the rice genome. The nucleotide sequence of one PAL gene, ZB8, was determined. The ZB8 gene is 4660 bp in length and consists of two exons and one intron. It encodes a polypeptide of 710 amino acids. The transcription start site was 137 bp upstream from the translation initiation site. Rice PAL transcripts accumulated to a high level in stems, with lower levels in roots and leaves. Wounding of leaf tissues induced ZB8 PAL transcripts to a high level. In rice suspension-cultured cells treated with fungal cell wall elicitors, the ZB8 PAL transcript increased within 30 min and reached maximum levels in 1-2 h. The transcription of the ZB8 gene was investigated by fusing its promoter to the reporter gene beta-glucuronidase (GUS) and transforming the construct into rice and tobacco plants, as well as rice suspension-cultured cells. High levels of GUS activity were observed in stems, moderate levels in roots and low levels in leaves of transgenic rice and tobacco plants. Histochemical analysis indicated that in transgenic rice the promoter was active in root apical tips, lateral root initiation sites, and vascular and epidermal tissues of stems and roots. In rice flowers, high GUS activity was observed in floral shoots, receptacles, anthers and filaments, occasionally GUS activity was also detected in lemma and awn tissues. In tobacco flowers, high GUS activity was detected in the pink part of petals. Consistent with the activity of endogenous PAL transcripts, wounding of rice and tobacco leaf tissues induced GUS activity from low basal levels. Tobacco mosaic virus (TMV) infection of tobacco leaves induced GUS activity to a high level. Fungal cell wall elicitors strongly induced GUS activity and GUS transcripts to high levels in transgenic rice suspension-cultured cells. We demonstrated that the promoter of ZB8 gene is both developmentally regulated and stress-inducible.