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Summarizes the effects of air pollutants on the control of stomatal aperture. Effect of both high and low atmospheric concentrations of sulfur dioxide and ozone on stomatal closure; Capacity of ozone to interfere with sodium-induced stomatal closure in Aster tripolium; Interference in the control of water relations of the Fagus sylvatica by elevated concentrations of carbon dioxide.

Oxidative stress is considered a contributor to declining muscle function and mobility during aging; however, the underlying molecular mechanisms remain poorly described. We hypothesized that greater levels of cysteine (Cys) oxidation on muscle proteins are associated with decreased measures of mobility. Herein, we applied a novel redox proteomics approach to measure reversible protein Cys oxidation in vastus lateralis muscle biopsies collected from 56 subjects in the Study of Muscle, Mobility and Aging (SOMMA), a community-based cohort study of individuals aged 70 years and older. We tested whether levels of Cys oxidation on key muscle proteins involved in muscle structure and contraction were associated with muscle function (leg power and strength), walking speed, and fitness (VO 2 peak on cardiopulmonary exercise testing) using linear regression models adjusted for age, sex, and body weight. Higher oxidation levels of select nebulin Cys sites were associated with lower VO 2 peak, while greater oxidation of myomesin-1, myomesin-2, and nebulin Cys sites was associated with slower walking speed. Higher oxidation of Cys sites in key proteins such as myomesin-2, alpha-actinin-2, and skeletal muscle alpha-actin were associated with lower leg power and strength. We also observed an unexpected correlation (R = 0.48) between a higher oxidation level of eight Cys sites in alpha-actinin-3 and stronger leg power. Despite this observation, the results generally support the hypothesis that Cys oxidation of muscle proteins impairs muscle power and strength, walking speed, and cardiopulmonary fitness with aging., (© 2024 Battelle Memorial Institute and The Authors. Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

Oxidative stress is considered a contributor to declining muscle function and mobility during aging; however, the underlying molecular mechanisms remain poorly described. We hypothesized that greater levels of cysteine (Cys) oxidation on muscle proteins are associated with decreased measures of mobility. Herein, we applied a novel redox proteomics approach to measure reversible protein Cys oxidation in vastus lateralis muscle biopsies collected from 56 subjects in the Study of Muscle, Mobility and Aging (SOMMA), a community-based cohort study of individuals aged 70 years and older. We tested whether levels of Cys oxidation on key muscle proteins involved in muscle structure and contraction were associated with muscle function (leg power and strength), walking speed, and fitness (VO 2 peak on cardiopulmonary exercise testing) using linear regression models adjusted for age, sex, and body weight. Higher oxidation levels of select nebulin Cys sites were associated with lower VO 2 peak, while greater oxidation of myomesin-1, myomesin-2, and nebulin Cys sites was associated with slower walking speed. Higher oxidation of Cys sites in key proteins such as myomesin-2, alpha-actinin-2, and skeletal muscle alpha-actin were associated with lower leg power and strength. We also observed an unexpected correlation (r = 0.48) between a higher oxidation level of 8 Cys sites in alpha-actinin-3 and stronger leg power. Despite this observation, the results generally support the hypothesis that Cys oxidation of muscle proteins impair muscle power and strength, walking speed, and cardiopulmonary fitness with aging., Competing Interests: Author Disclosure S.R.C. and P.M.Ca. are consultants to Bioage Labs. All other authors declare no conflict of interest.

Objective: To evaluate the 104-week safety of dapagliflozin in older patients with type 2 diabetes mellitus., Methods: Pooled analysis assessing general safety (nine phase III studies ≤104 weeks) and cardiovascular safety (21 phase IIb/III studies ≤208 weeks) by age (<65; ≥65; ≥75 years). Patients with type 2 diabetes mellitus (±background glucose-lowering therapy) received: dapagliflozin 10 mg (n = 2026) vs. placebo (n = 1956) (nine-study pool); or dapagliflozin (2.5-50 mg; n = 5936) vs. control (placebo/comparator) (n = 3403) (21-study pool)., Results: Adverse events (AEs) and discontinuations owing to AEs were more common in older vs. younger patients, and were more frequent with dapagliflozin than placebo (AEs: <65 years: 73.1 vs. 70.7 %; ≥65 years: 77.4 vs. 73.1 %; ≥75 years: 80.4 vs. 75.3 %, respectively; discontinuations: <65 years: 5.9 vs. 5.0 %; ≥65 years: 14.4 vs. 12.2 %; ≥75 years: 26.8 vs. 22.1 %, respectively); serious AE (SAE) frequency was similar (<65 years: 11.0 vs. 11.8 %; ≥65 years: 20.0 vs. 20.2 %; ≥75 years: 19.6 vs. 18.2 %, respectively). Hypoglycaemia frequency was similar across age groups and was higher with dapagliflozin than placebo (<65 years: 18.0 vs. 13.4 %; ≥65 years: 20.2 vs. 17.7 %; ≥75 years: 17.5 vs. 16.9 %, respectively); major episodes were rare. Urinary tract infection frequency was similar between treatment groups in older patients, with no increase vs. younger patients (<65 years: 8.8 vs. 5.5 %; ≥65 years: 8.1 vs. 7.6 %; ≥75 years: 8.2 vs. 9.1 %, respectively); urinary tract infection SAEs were rare. Genital infection AEs were more common with dapagliflozin, with no increase in older patients (<65 years: 8.2 vs. 1.0 %; ≥65 years: 6.6 vs. 0.9 %; ≥75 years: 7.2 vs. 0.0 %, respectively) and no SAEs. Volume reduction AEs were uncommon, with a higher frequency with dapagliflozin vs. placebo and in patients ≥75 years (<65 years: 1.7 vs. 1.2 %; ≥65 years: 2.3 vs. 1.7 %; ≥75 years: 3.1 vs. 2.6 %, respectively). Dapagliflozin did not increase the risk of fractures (<65 years: 1.1 vs. 1.1 %; ≥65 years: 1.1 vs. 2.7 %; ≥75 years: 1.0 vs. 2.6 %, respectively) or falls (<65 years: 0.7 vs. 0.7 %; ≥65 years: 0.6 vs. 2.1 %; ≥75 years: 0.0 vs. 1.3 %, respectively), regardless of age. AEs of renal function were more common with dapagliflozin than placebo and increased with age (<65 years: 3.5 vs. 2.3 %; ≥65 years: 14.0 vs. 7.9 %; ≥75 years: 29.9 vs. 20.8 %, respectively). Most were non-serious small transient increases in serum creatinine. Dapagliflozin did not increase cardiovascular risk regardless of age [hazard ratio (95 % confidence interval) vs., Control: <65 years: 0.726 (0.473, 1.114); ≥65 years: 0.879 (0.565, 1.366); ≥75 years: 0.950 (0.345, 2.617), respectively]., Conclusion: Dapagliflozin treatment up to 104 weeks was well tolerated in older patients. Older dapagliflozin-treated patients had more renal AEs than placebo-treated patients; the majority of which were non-serious small transient changes in serum creatinine.

Objective: Dapagliflozin reduces hyperglycemia in type 2 diabetes mellitus (T2DM) and lowers blood pressure, at least in part, secondary to mild diuresis consequent to dapagliflozin-induced glucosuria. While blood-pressure lowering may contribute to cardiovascular risk reduction, dapagliflozin-induced diuresis may potentially contribute to adverse events (AEs) of volume reduction. The present analysis compared the frequency of AEs of volume reduction between dapagliflozin and placebo., Methods: Pooled data were assessed from 13 placebo-controlled dapagliflozin clinical trials ≤24 weeks in patients with T2DM, overall, and in those at risk (aged ≥65y, estimated glomerular filtration rate [eGFR] <60 mL/min/1.73 m(2), or on antihypertensive therapy). Longer-term (≤104 weeks) data were available for 9 of these trials., Results: The frequency of patients experiencing ≥1 AE of volume reduction over 24 weeks was low overall; 27/2360 (1.1%) with dapagliflozin 10 mg and 17/2295 (0.7%) with placebo; and slightly more frequent in patients ≥65 years (11/665 [1.7%] and 6/711 [0.8%], respectively) and in patients receiving loop diuretics (6/236 [2.5%] and 4/267 [1.5%], respectively). Over 104 weeks, AEs of volume reduction occurred in 38/2026 (1.9%) with dapagliflozin 10 mg and in 27/1956 (1.4%) with placebo; serious AEs of volume reduction in 4/2026 (0.2%) and 6/1956 (0.3%), respectively; and 2 patients in each group discontinued therapy due to these AEs. Dapagliflozin versus placebo incidence rate ratios did not suggest any meaningful increase in frequency of these AEs with dapagliflozin 10 mg, either overall or in those at risk. Although mean eGFR declined by 4.2 ml/min/1.73 m(2) within the first week of dapagliflozin therapy, thereafter eGFR gradually recovered to baseline levels by 104 weeks (mean change from baseline +0.02 mL/min/1.73 m(2); 95%CI: -0.9, 1.0)., Conclusion: No meaningful increase in frequency of AEs of volume reduction occurred with dapagliflozin 10 mg in patients with T2DM, either overall, or in those at increased risk of these events. However, caution should nevertheless be exercised when prescribing dapagliflozin to elderly patients, those with reduced eGFR, and those receiving antihypertensive medication.

Background: Hypertension is a common comorbidity in patients with type 2 diabetes mellitus and a major risk factor for microvascular and macrovascular disease. Although the blood pressure-lowering effects of sodium-glucose cotransporter 2 (SGLT2) inhibitors are already established, guidance is needed on how to use these drugs in patients already receiving antihypertensive therapy. We aimed to compare blood pressure and glycaemic effects of the SGLT2 inhibitor dapagliflozin with placebo in patients with inadequately controlled type 2 diabetes mellitus and hypertension., Methods: In this double-blind, placebo-controlled, phase 3 study we enrolled patients from 311 centres in 16 countries across five continents. Patients had uncontrolled type 2 diabetes (HbA1c 7·0%-10·5%; 53-91 mmol/mol) and hypertension (systolic 140-165 mm Hg and diastolic 85-105 mm Hg at both enrolment and randomisation, and a mean 24 h blood pressure of ≥130/80 mm Hg by ambulatory monitoring within 1 week of randomisation) and were receiving oral antihyperglycaemic drugs, insulin, or both, plus a renin-angiotensin system blocker and an additional antihypertensive drug. Using an interactive voice-response system, we randomly assigned (1:1) patients to dapagliflozin 10 mg once a day or to placebo, with randomisation stratified by additional antihypertensive drug use and insulin use at baseline, in a block size of two. The co-primary endpoints were changes in seated systolic blood pressure and HbA1c measured in the full analysis set, which included all patients who received at least one dose of study drug and had both a baseline and at least one post-baseline measurement of efficacy. This trial is registered with ClinicalTrials.gov, number NCT01195662., Findings: Between Oct 29, 2010, and Oct 4, 2012, we randomly assigned 225 patients to dapagliflozin and 224 to placebo. Seated systolic blood pressure was significantly reduced in the group assigned to dapagliflozin (adjusted mean change from baseline -11·90 mm Hg [95% CI -13·97 to -9·82]) compared with those assigned to placebo (-7·62 mm Hg [-9·72 to -5·51]; placebo-adjusted difference for dapagliflozin -4·28 mm Hg [-6·54 to -2·02]; p=0·0002). Reductions in HbA1c concentrations were also significantly greater in patients assigned to dapagliflozin (adjusted mean change from baseline -0·63% [95% CI -0·76 to -0·50]) than in those assigned to placebo (-0·02% [-0·15 to 0·12]; placebo-adjusted difference -0·61% [-0·76 to -0·46,]; p<0·0001). In a post-hoc analysis, we found difference in blood pressure versus placebo was greater in patients receiving a β blocker (-5·76 mm Hg [95% CI -10·28 to -1·23]) or a calcium-channel blocker (-5·13 mm Hg, [-9·47 to -0·79]) as their additional antihypertensive drug than in those receiving a thiazide diuretic (-2·38 mm Hg [-6·16 to 1·40]). Adverse events were similar in the dapagliflozin and placebo groups (98 [44%] patients vs 93 [42%], respectively, had at least one adverse event), with few adverse events related to renal function (1% vs <1%) or volume depletion (<1% vs 0%)., Interpretation: Dapagliflozin 10 mg significantly improved blood pressure and HbA1c and was tolerated similarly to placebo. Its blood pressure-lowering properties were particularly favourable in patients already receiving a β blocker or calcium-channel blocker. Dapagliflozin could benefit patients with type 2 diabetes who need a diuretic-like effect to optimise control of blood pressure, adding meaningful efficacy to antihypertensive drug regimens., Funding: Bristol-Myers Squibb, AstraZeneca., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Introduction: Hyperkalemia risk is increased in diabetes, particularly in patients with renal impairment or those receiving angiotensin-converting enzyme (ACE) inhibitors, angiotensin receptor blockers (ARBs) or potassium-sparing diuretics. Conversely, other diuretics can increase hypokalemia risk. We assessed the effects of the sodium glucose co-transporter 2 (SGLT2) inhibitor dapagliflozin on serum potassium levels in a pooled analysis of clinical trials in patients with type 2 diabetes mellitus (T2DM)., Methods: Fourteen randomized, placebo-controlled, double-blind T2DM studies were analyzed: pooled data from 13 studies of ≤24 weeks' duration (dapagliflozin 10 mg, N = 2360; placebo, N = 2295); and one 52-week moderate renal impairment study in patients with baseline eGFR ≥30 to <60 mL/min/1.73 m(2) (dapagliflozin 10 mg, N = 85; placebo, N = 84). Central laboratory serum potassium levels were determined at each study visit., Results: No clinically relevant mean changes from baseline in serum potassium ≤24 weeks were reported for dapagliflozin 10 mg [-0.05 mmol/L; 95% confidence interval (CI) -0.07, -0.03] versus placebo (-0.02 mmol/L; 95% CI -0.04, 0.00) in the pooled population or in the renal impairment study (-0.03 mmol/L; 95% CI -0.14, 0.08 vs. -0.02 mmol/L; 95% CI -0.13, 0.09, respectively). The incidence rate ratio for serum potassium ≥5.5 mmol/L over 24 weeks for dapagliflozin 10 mg versus placebo was 0.90 (95% CI 0.74, 1.10) in the pooled population; with no increased risk in patients receiving ARBs/ACE inhibitors, or potassium-sparing diuretics, or in those with moderate renal impairment. Slightly more patients receiving dapagliflozin 10 mg had serum potassium ≤3.5 mmol/L versus placebo (5.2% vs. 3.6%); however, no instances of serum potassium ≤2.5 mmol/L were reported., Conclusion: Dapagliflozin is not associated with an increased risk of hyperkalemia or severe hypokalemia in patients with T2DM., Funding: Bristol-Myers Squibb and AstraZeneca.

Hypertension and type 2 diabetes mellitus (T2DM) are risk factors for cardiovascular disease. Dapagliflozin improves glycemic control and systolic blood pressure (SBP) in T2DM patients. This double-blind phase III study evaluated the effects of dapagliflozin on glycemic control and blood pressure in patients with inadequately controlled T2DM and hypertension, despite ongoing therapy with a renin-angiotensin system blocker. Patients were randomized to receive dapagliflozin 10 mg (n = 302) or placebo (n = 311) once daily for 12 weeks. Endpoints were change from baseline to week 12 in seated SBP and glycosylated hemoglobin (HbA1c); longitudinal repeated-measures analysis was performed. Additional endpoints included other hemodynamic measures, serum uric acid, fasting plasma glucose, body weight, blood lipids and heart rate. After 12 weeks, dapagliflozin-treated versus placebo-treated patients showed significant reductions in HbA1c (-0.6% vs -0.1%, p < 0.0001), mean seated SBP (-10.4 vs -7.3 mmHg, p = 0.0010) and mean 24 h ambulatory SBP (-9.6 vs -6.7 mmHg, p = 0.0043). Dapagliflozin also reduced body weight compared with placebo (-1.0 vs -0.3 kg). Dapagliflozin was well tolerated, with adverse events consistent with previous studies. Dapagliflozin improved glycemic control, and reduced SBP as well as body weight in patients with poorly controlled T2DM and hypertension.

Objective: Dapagliflozin is a highly selective, orally active inhibitor of renal sodium-glucose cotransporter 2 that reduces hyperglycemia by increasing urinary glucose excretion. The goal of this study was to evaluate dapagliflozin as monotherapy in drug-naive Asian patients with type 2 diabetes whose disease was inadequately controlled with diet and exercise., Methods: In this Phase III, multicenter, parallel-group, double-blind study, drug-naive patients with glycosylated hemoglobin (HbA1c) levels ≥7.0% to ≤10.5% (≥53-≤91 mmol/mol) were randomized (by using an interactive voice response system) to receive placebo (n = 132), dapagliflozin 5 mg (n = 128), or dapagliflozin 10 mg (n = 133). The primary end point was mean change from baseline in HbA1c level at week 24 (last-observation-carried-forward). Secondary end points included changes in fasting plasma glucose, 2-hour postprandial glucose, body weight, and other glycemic parameters., Results: Baseline characteristics were balanced across groups. Most patients (89%) were Chinese, median disease duration was 0.2 year, and mean HbA1c level was 8.26%. Most patients (87%) completed the study. At week 24, mean reductions in HbA1c were -0.29% for placebo versus -1.04% and -1.11% for dapagliflozin 5 and 10 mg, respectively (P < 0.0001 for both doses). Changes in fasting plasma glucose were 2.5, -25.1, and -31.6 mg/dL (0.14, -1.39, and -1.75 mmol/L) for placebo, dapagliflozin 5 mg, and dapagliflozin 10 mg. Changes in 2-hour postprandial glucose were 1.1, -46.8, and -54.9 mg/dL (0.06, -2.60, and -3.05 mmol/L). Reductions in body weight were -0.27, -1.64, and -2.25 kg. Proportions of patients achieving HbA1c levels <7.0% (53 mmol/mol) were 21.3%, 42.6%, and 49.8%. Adverse events (AEs) occurred in 63.6%, 61.7%, and 60.9% of patients, and serious AEs occurred in 1.5%, 3.9%, and 3.0% of patients. No deaths occurred. Hypoglycemia was uncommon (1.5%, 0.8%, and 0.8%); no hypoglycemic event led to discontinuation. Genital infections occurred in 0.8%, 3.1%, and 4.5% of patients and urinary tract infections in 3.0%, 3.9%, and 5.3% of patients. No AEs of renal infection or pyelonephritis were reported. No changes in renal function or AEs of renal failure occurred., Conclusions: Compared with placebo, dapagliflozin 5 and 10 mg demonstrated clinically and statistically significant improvements in HbA1c levels after 24 weeks of treatment. Dose-dependent, statistically significant reductions in fasting plasma glucose, postprandial glucose, and weight were also observed for both doses compared with placebo. AEs and serious AEs were balanced across groups, with low rates of hypoglycemia and no increase in renal events. Genital infections and urinary tract infections were more common with dapagliflozin. Dapagliflozin as monotherapy in these drug-naive Asian patients was well tolerated, significantly improving glycemic control with the additional benefit of weight loss., (© 2014 The Authors. Published by Elsevier HS Journals, Inc. All rights reserved.)

Background: Management of type 2 diabetes with metformin often does not provide adequate glycemic control, thereby necessitating add-on treatment. In a 24-week clinical trial, dapagliflozin, an investigational sodium glucose cotransporter 2 inhibitor, improved glycemic control in patients inadequately controlled with metformin. The present study is an extension that was undertaken to evaluate dapagliflozin as long-term therapy in this population., Methods: This was a long-term extension (total 102 weeks) of a 24-week phase 3, multicenter, randomized, placebo-controlled, double-blind, parallel-group trial. Patients were randomly assigned (1:1:1:1) to blinded daily treatment (placebo, or dapagliflozin 2.5 to 5, or 10 mg) plus open-label metformin (≥1,500 mg). The previously published primary endpoint was change from baseline in glycated hemoglobin (HbA1c) at 24 weeks. This paper reports the follow-up to week 102, with analysis of covariance model performed at 24 weeks with last observation carried forward; a repeated measures analysis was utilized to evaluate changes from baseline in HbA1c, fasting plasma glucose (FPG), and weight., Results: A total of 546 patients were randomized to 1 of the 4 treatments. The completion rate for the 78-week double-blind extension period was lower for the placebo group (63.5%) than for the dapagliflozin groups (68.3% to 79.8%). At week 102, mean changes from baseline HbA1c (8.06%) were +0.02% for placebo compared with -0.48% (P = 0.0008), -0.58% (P <0.0001), and -0.78% (P <0.0001) for dapagliflozin 2.5 to 5, and 10 mg, respectively. In addition, all dapagliflozin groups had sustained reductions from baseline in FPG (-1.07 to -1.47 mmol/l) and body weight (-1.10 to -1.74 kg) at 102 weeks, whereas increases were noted in placebo-treated patients for both of these outcomes. Events of hypoglycemia were rare and were not severe. Evidence suggestive of genital infection was reported in 11.7% to 14.6% of dapagliflozin patients and 5.1% of placebo patients, with one related discontinuation (dapagliflozin 5 mg). Evidence suggestive of urinary tract infection was reported in 8.0% to 13.3% of dapagliflozin patients and 8.0% of placebo patients, with one related discontinuation (dapagliflozin 2.5 mg)., Conclusions: Dapagliflozin added to metformin for 102 weeks enabled sustained reductions in HbA1c, FPG, and weight without increased risk of hypoglycemia in patients with type 2 diabetes who were inadequately controlled on metformin alone., Trial Registration: ClinicalTrials.gov: NCT00528879.

Locus At5g03555 encodes a nucleobase cation symporter 1 (AtNCS1) in the Arabidopsis genome. Arabidopsis insertion mutants, AtNcs1-1 and AtNcs1-3, were used for in planta toxic nucleobase analog growth studies and radio-labeled nucleobase uptake assays to characterize solute transport specificities. These results correlate with similar growth and uptake studies of AtNCS1 expressed in Saccharomyces cerevisiae. Both in planta and heterologous expression studies in yeast revealed a unique solute transport profile for AtNCS1 in moving adenine, guanine and uracil. This is in stark contrast to the canonical transport profiles determined for the well-characterized S. cerevisiae NCS1 proteins FUR4 (uracil transport) or FCY2 (adenine, guanine, and cytosine transport)., (Copyright © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

In plants, nucleobase biochemistry is highly compartmented relying upon a well-regulated and selective membrane transport system. In Arabidopsis two proteins, AtAzg1 and AtAzg2, show substantial amino acid sequence similarity to the adenine-guanine-hypoxanthine transporter AzgA of Aspergillus nidulans. Analysis of single and double mutant lines harboring T-DNA insertion alleles AtAzg1-1 and AtAzg2-1 reveal a marked resistance to growth in the presence of 8-azaadenine and 8-azaguanine but not to other toxic nucleobase analogues. Conversely, yeast strains expressing AtAzg1 and AtAzg2 gain heightened sensitivity to growth on 8-azaadenine and 8-azaguanine. Radio-labeled purine uptake experiments in yeast and in planta confirm the function of AtAzg1 and AtAzg2 as plant adenine-guanine transporters.

Purpose: To determine whether lowering oxygen saturation alarm limits for infants at risk for retinopathy of prematurity (ROP) reduces its incidence and/or severity., Methods: Oximetry alarm limits were lowered to 85% and 93% for all infants with a birth weight 1250 g or less and/or gestational age 28 weeks or less, and maintained until 32 weeks' postmenstrual age or until oxygen saturations were consistently greater than 93% in room air. The new policy was effective for infants born on or after June 1, 2003. ROP data were prospectively collected, and we compared the rate and severity of ROP in the year after the oximeter alarm policy change to the rates in the immediately preceding 3 years., Results: In the year after the oximeter alarm limit policy change, 4 of 72 infants developed prethreshold ROP compared with 44 of 251 infants in the previous 3-year epoch (17.5% vs 5.6%, p=0.01). Similarly, only 6 of 144 eyes developed prethreshold ROP in the year after the policy change, compared with 84 of 502 in the previous 3 years (16.7% vs 4.2%, p=0.001)., Conclusions: A simple change in oximeter alarm parameters in the first weeks of life for infants with a birth weight 1250 g or less may decrease the incidence of prethreshold ROP.

A fluoroorotic acid (FOA)-resistant mutant of Arabidopsis thaliana was isolated by screening M2 populations of ethyl methane sulphonate (EMS)-mutagenized Columbia seed. FOA resistance was due to a nuclear recessive gene, for1-1, which locates to a 519 kb region in chromosome 5. Assays of key regulatory enzymes in de novo pyrimidine synthesis (uridine monophosphate synthase) and salvage biochemistry (thymidine kinase) confirmed that FOA resistance in for1-1/for1-1 plants was not due to altered enzymatic activities. Uptake studies using radiolabelled purines, pyrimidines, and [14C]FOA reveal that for1-1/for1-1 plants were specifically defective in the uptake of uracil or uracil-like bases. To confirm such specificity, genetic crosses show that FOR1 is a distinct locus from FUR1 which encodes a deoxyuridine nucleoside transporter. In addition, for1-1/for1-1 plants were restored to FOA sensitivity by transformation with the Escherichia coli uracil transporter gene uraA driven by the cauliflower mosaic virus (CaMV) 35S promoter. Molecular mapping studies reveal that FOR1 does not correspond to loci belonging to any of the six known nucleobase transporter families identified in the Arabidopsis genome. Moreover, FOR1 does not appear to regulate the transcript levels of either uracil transporter-encoding loci At2g03590 or At2g03530. The above results strongly suggest that the for1-1 mutant allele affects a transport mechanism that is specific for the uptake of uracil.

Farnesoid X receptor (FXR) is a bile acid-activated transcription factor that is a member of the nuclear hormone receptor superfamily. Fxr-null mice exhibit a phenotype similar to Byler disease, an inherited cholestatic liver disorder. In the liver, activation of FXR induces transcription of transporter genes involved in promoting bile acid clearance and represses genes involved in bile acid biosynthesis. We investigated whether the synthetic FXR agonist GW4064 could protect against cholestatic liver damage in rat models of extrahepatic and intrahepatic cholestasis. In the bile duct-ligation and alpha-naphthylisothiocyanate models of cholestasis, GW4064 treatment resulted in significant reductions in serum alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase, as well as other markers of liver damage. Rats that received GW4064 treatment also had decreased incidence and extent of necrosis, decreased inflammatory cell infiltration, and decreased bile duct proliferation. Analysis of gene expression in livers from GW4064-treated cholestatic rats revealed decreased expression of bile acid biosynthetic genes and increased expression of genes involved in bile acid transport, including the phospholipid flippase MDR2. The hepatoprotection seen in these animal models by the synthetic FXR agonist suggests FXR agonists may be useful in the treatment of cholestatic liver disease.

The nuclear bile acid receptor FXR has been proposed to play a central role in the feedback repression of the gene encoding cholesterol 7 alpha-hydroxylase (CYP7A1), the first and rate-limiting step in the biosynthesis of bile acids. We demonstrate that FXR directly regulates expression of fibroblast growth factor-19 (FGF-19), a secreted growth factor that signals through the FGFR4 cell-surface receptor tyrosine kinase. In turn, FGF-19 strongly suppresses expression of CYP7A1 in primary cultures of human hepatocytes and mouse liver through a c-Jun N-terminal kinase (JNK)-dependent pathway. This signaling cascade defines a novel mechanism for feedback repression of bile acid biosynthesis and underscores the vital role of FXR in the regulation of multiple pathways of cholesterol catabolism in the liver.

The application of advanced modern biomedical and chemical research technologies in the pharmaceutical industry has led to a significant increase in the number of potential drug targets and lead candidates. Whereas the drug discovery process is enhanced significantly, the failure rate of new compounds due to toxicity remains very high. The pharmaceutical industry is setting high hopes on the new discipline of toxicogenomics to revolutionize the process of drug toxicity assessment by reducing the bottleneck of new drug candidates and minimizing late-stage developmental failures. Toxicogenomics is expected to facilitate the efficient screening of new compounds at an early stage, resulting in significant savings of time and cost associated with new drug development. In this review, a general description of the new discipline of toxicogenomics and its potential impact on the safety assessment of new drugs in the pharmaceutical industry is provided. An overview of the key issues and questions that are confronting investigators in this field today is also given as well as a prospective view of the future of this new discipline.

•  When Aster tripolium is grown at high salinity, stomatal closure is induced by the presence of sodium ions in the apoplast surrounding the guard cells. The occurrence of this system in Aster tripolium and not in the closely related glycophyte Aster amellus suggests that it could be an important factor in the network of physiological attributes required for salt tolerance. •  Gas exchange and growth parameters were measured in Aster tripolium plants grown at different levels of salinity. A simple mechanistic model was constructed to test whether the Na-sensing feature of the guard cells was a realistic component of salinity tolerance. •  The model captured very well the behaviour of plants in terms of salt uptake and reduction of growth with increasing salinity. There was moderate variance between measured and modelled rates of decrease of conductance with increasing levels of salinity. •  No evidence was found to refute our hypothesis that stomatal closure in response to sodium plays an important role in salt tolerance of Aster tripolium.

Background: Chediak-Higashi syndrome (CHS) is an inherited immunodeficiency disease characterized by giant lysosomes and impaired leukocyte degranulation. CHS results from mutations in the lysosomal trafficking regulator (LYST) gene, which encodes a 425-kD cytoplasmic protein of unknown function. The goal of this study was to identify proteins that interact with LYST as a first step in understanding how LYST modulates lysosomal exocytosis., Materials and Methods: Fourteen cDNA fragments, covering the entire coding domain of LYST, were used as baits to screen five human cDNA libraries by a yeast two-hybrid method, modified to allow screening in the activation and the binding domain, three selectable markers, and more stringent confirmation procedures. Five of the interactions were confirmed by an in vitro binding assay., Results: Twenty-one proteins that interact with LYST were identified in yeast two-hybrid screens. Four interactions, confirmed directly, were with proteins important in vesicular transport and signal transduction (the SNARE-complex protein HRS, 14-3-3, and casein kinase II)., Conclusions: On the basis of protein interactions, LYST appears to function as an adapter protein that may juxtapose proteins that mediate intracellular membrane fusion reactions. The pathologic manifestations observed in CHS patients and in mice with the homologous mutation beige suggest that understanding the role of LYST may be relevant to the treatment of not only CHS but also of diseases such as asthma, urticaria, and lupus, as well as to the molecular dissection of the CHS-associated cancer predisposition.

Regulation of the concentration of Ca2+ in the apoplast of plants is essential in order to allow Ca(2+)-dependent processes, such as Ca(2+)-mediated signal transduction in stomatal guard cells, to function correctly. This is particularly important for plants growing with high levels of Ca2+ in the rhizosphere. Recently, we have shown that in two calcicoles, Leontodon hispidus L. and Centaurea scabiosa L., trichomes play a key role in this regulatory process. Ozone is known to have a marked effect on plant Ca2+ homeostasis. Therefore, we have examined the effect of this pollutant on the regulation by trichomes of apoplastic Ca2+ in the calcicole L. hispidus. Treatment with 100 nl l-1 ozone resulted in a reduction in stomatal conductance of approximately 25% in plants grown with 15 mM Ca2+ in the rhizosphere. Analysis of total Ca2+ levels revealed that these changes in stomatal behaviour reflect a decrease in the ability of trichomes to sequester Ca2+. The amount of Ca2+ present in the trichome tip cell was reduced by approximately 38%. This was accompanied by an increase in the levels of Ca2+ in the guard cells and other tissues of the leaf. These data suggest that ozone has a detrimental effect on the ability of trichomes to regulate the concentration of apoplastic Ca2+ in L. hispidus, resulting in altered stomatal behaviour, and hence gaseous exchange, possibly due to the disruption of guard-cell Ca(2+)-mediated signal transduction. This has important implications for the growth and survival of plants growing in Ca(2+)-rich soils.

Peroxisome proliferator-activated receptor gamma (PPAR gamma) agonists, including the glitazone class of drugs, are insulin sensitizers that reduce glucose and lipid levels in patients with type 2 diabetes mellitus. To more fully understand the molecular mechanisms underlying their therapeutic actions, we have characterized the effects of the potent, tyrosine-based PPAR gamma ligand GW1929 on serum glucose and lipid parameters and gene expression in Zucker diabetic fatty rats. In time-course studies, GW1929 treatment decreased circulating FFA levels before reducing glucose and triglyceride levels. We used a comprehensive and unbiased messenger RNA profiling technique to identify genes regulated either directly or indirectly by PPAR gamma in epididymal white adipose tissue, interscapular brown adipose tissue, liver, and soleus skeletal muscle. PPAR gamma activation stimulated the expression of a large number of genes involved in lipogenesis and fatty acid metabolism in both white adipose tissue and brown adipose tissue. In muscle, PPAR gamma agonist treatment decreased the expression of pyruvate dehydrogenase kinase 4, which represses oxidative glucose metabolism, and also decreased the expression of genes involved in fatty acid transport and oxidation. These changes suggest a molecular basis for PPAR gamma-mediated increases in glucose utilization in muscle. In liver, PPAR gamma activation coordinately decreased the expression of genes involved in gluconeogenesis. We conclude from these studies that the antidiabetic actions of PPAR gamma agonists are probably the consequence of 1) their effects on FFA levels, and 2), their coordinate effects on gene expression in multiple insulin-sensitive tissues.

The association of inheritance of different apolipoprotein E (APOE, gene; apoE, protein) alleles with the risk and rate of onset of Alzheimer's disease (AD) is now well established and widely confirmed. While there are now a collection of hypotheses concerning the specific relationship of APOE polymorphisms to various phenotypic manifestations of AD, no single compelling theory has been tested and universally accepted. The only clear fact emerging during the past 6 years is that differences in APOE genotype affect the average rate of disease onset as a predictable function of the inheritance of this polymorphic gene. Methods now exist to enable experimental designs to study the metabolic effects of inheriting different APOE alleles, addressing what differences that may be present for many years, perhaps over the entire lifetime, can lead to earlier or later manifestations of the disease and are therapeutically tractable. This review summarizes part of an experimental approach to identify biological pathways influenced by the different APOE polymorphisms that are relevant to the pathogenesis of AD.

Two large-scale yeast two-hybrid screens were undertaken to identify protein-protein interactions between full-length open reading frames predicted from the Saccharomyces cerevisiae genome sequence. In one approach, we constructed a protein array of about 6,000 yeast transformants, with each transformant expressing one of the open reading frames as a fusion to an activation domain. This array was screened by a simple and automated procedure for 192 yeast proteins, with positive responses identified by their positions in the array. In a second approach, we pooled cells expressing one of about 6,000 activation domain fusions to generate a library. We used a high-throughput screening procedure to screen nearly all of the 6,000 predicted yeast proteins, expressed as Gal4 DNA-binding domain fusion proteins, against the library, and characterized positives by sequence analysis. These approaches resulted in the detection of 957 putative interactions involving 1,004 S. cerevisiae proteins. These data reveal interactions that place functionally unclassified proteins in a biological context, interactions between proteins involved in the same biological function, and interactions that link biological functions together into larger cellular processes. The results of these screens are shown here.

Small changes in the gaseous composition of the atmosphere have many different impacts on terrestrial plants. Some of the most important involve changes in stomatal control of leaf conductance. Evolution has provided highly complex mechanisms by which stomata respond to a wide range of environmental factors to balance the conflicting priorities of carbon gain for photosynthesis and water conservation. These mechanisms involve direct responses of the guard cells to aspects of the aerial environment, and hormonal communication within the plant enabling conductance to be adjusted according to soil moisture status. Various aspects of these delicately balanced mechanisms can be disturbed by air pollutants. Impairment of the regulation of plant water use by SO2 and O3 has been known for some years, but there are still many obstacles to our understanding of the variations in response between species, or even between genotypes of the same species. A surprising outcome of some recent studies is the suggestion that CO2 pollution may disrupt the control of water relations in some species because their stomata do not close sufficiently in CO2-enriched air. It has often been taken for granted that the elevation of atmospheric CO2 would lead to economies in water use by plant canopies, but the underlying assumptions are now being seriously questioned.

Analysis of patients with inherited hypokalaemic alkalosis resulting from salt-wasting has proved fertile ground for identification of essential elements of renal salt homeostasis and blood-pressure regulation. We now demonstrate linkage of this phenotype to a segment of chromosome 1 containing the gene encoding a renal chloride channel, CLCNKB. Examination of this gene reveals loss-of-function mutations that impair renal chloride reabsorption in the thick ascending limb of Henle's loop. Mutations in seventeen kindreds have been identified, and they include large deletions and nonsense and missense mutations. Some of the deletions are shown to have arisen by unequal crossing over between CLCNKB and the nearby related gene, CLCNKA. Patients who harbour CLCNKB mutations are characterized by hypokalaemic alkalosis with salt-wasting, low blood pressure, normal magnesium and hyper- or normocalciuria; they define a distinct subset of patients with Bartter's syndrome in whom nephrocalcinosis is absent. These findings demonstrate the critical role of CLCNKB in renal salt reabsorption and blood-pressure homeostasis, and demonstrate the potential role of specific CLCNKB antagonists as diuretic antihypertensive agents.

Monoliths of two contrasting vegetation types, a species-rich grassland on a brown earth soil over limestone and species-poor community on a peaty gley, were transferred to solardomes and grown under ambient (350 μ 1 -1 ) and elevated (600 μ11 -1 ) CO 2 for 2 yr. Shoot biomass was unaltered but root biomass increased by 40-50% under elevated CO 2 . Root production was increased by elevated CO 2 in the peat soil, measured both as instantaneous and cumulative rates, but only the latter measure was increased in the limestone soil. Root growth was stimulated more at 6 cm depth than at 10 cm in the limestone soil. Turnover was faster under elevated CO 2 in the peat soil, but there was only a small effect on turnover in the limestone soil. Elevated CO 2 reduced nitrogen concentration in roots and might have increased mycorrhizal colonization. Respiration rate was correlated with N concentration, and was therefore lower in roots grown at elevated CO 2 . Estimates of the C budget of the two communities, based upon root production and on net C uptake, suggest that C sequestration in the peat soil increases by c. 0.2 kg C m -2 yr -1 (= 2 t ha yr -1 ) under elevated CO 2 .

Essential hypertension is a common multifactorial trait. The molecular basis of a number of rare diseases that after blood pressure in humans has been established, identifying pathways that may be involved in more common forms of hypertension. Pseudohypoaldosteronism type II (PHAII, also known as familial hyperkalaemia and hypertension or Gordon's syndrome; OMIM #145260), is characterized by hyperkalaemia despite normal renal glomerular filtration, hypertension and correction of physiologic abnormalities by thiazide diuretics. Mild hyperchloremia, metabolic acidosis and suppressed plasma renin activity are variable associated findings. The pathogenesis of PHAII is unknown, although clinical studies indicate an abnormality in renal ion transport. As thiazide diuretics are among the most efficacious agents in the treatment of essential hypertension, understanding the pathogenesis of PHAII may be of relevance to more common forms of hypertension. Analysis of linkage in eight PHAII families showing autosomal dominant transmission demonstrates locus heterogeneity of this trait, with a multilocus lod score of 8.1 for linkage of PHAII to chromosomes 1q31-q42 and 17p11-q21. Interestingly, the chromosome-17 locus overlaps a syntenic interval in rat that contains a blood pressure quantitative trait locus (QTL). Our findings provide a first step toward identification of the molecular basis of PHAII.

We have investigated the cellular basis for the effects of oxidative stress on stomatal behavior using stomatal bioassay and ratio photometric techniques. Two oxidative treatments were employed in this study: (a) methyl viologen, which generates superoxide radicals, and (b) H2O2. Both methyl viologen and H2O2 inhibited stomatal opening and promoted stomatal closure. At concentrations [less than or equal to]10-5 M, the effects of methyl viologen and H2O2 on stomatal behavior were reversible and were abolished by 2 mM EGTA or 10 [mu]M verapamil. In addition, at 10-5 M, i.e. the maximum concentration at which the effects of the treatments were prevented by EGTA or verapamil, methyl viologen and H2O2 caused an increase in guard cell cytosolic free Ca2+ ([Ca2+]i), which was abolished in the presence of EGTA. Therefore, at low concentrations of methyl viologen and H2O2, removal of extracellular Ca2+ prevented both the oxidative stress-induced changes in stomatal aperture and the associated increases in [Ca2+]i. This suggests that in this concentration range the effects of the treatments are Ca2+-dependent and are mediated by changes in [Ca2+]i. In contrast, at concentrations of methyl viologan and H2O2 > 10-5 M, EGTA and verapamil had no effect. However, in this concentration range the effects of the treatments were irreversible and correlated with a marked reduction in membrane integrity and guard cell viability. This suggests that at high concentrations the effects of methyl viologen and H2O2 may be due to changes in membrane integrity. The implications of oxidative stress-induced increases in [Ca2+]i and the possible disruption of guard-cell Ca2+ homeostasis are discussed in relation to the processes of Ca2+-based signal transduction in stomatal guard cells and the control of stomatal aperture.

The DCC (deleted in colorectal cancer) candidate tumor suppressor gene spans greater than 1350 kilobases at chromosomes 18q21.1 and encodes a transmembrane protein of unknown function. Although DCC is expressed in a number of adult tissues, its expression is highest in the brain and we have, therefore, undertaken studies to determine if DCC inactivation might contribute to tumors arising there. Decreased or absent DCC protein expression was noted in more than 50% of the thirty brain tumors studied. Although specific mutations in the DCC gene were not identified, a variety of mechanisms appeared to contribute to the altered DCC expression, including allelic loss, aberrant splicing of transcripts and allele-specific loss of transcripts. In total, the data suggest that DCC inactivation may be important in brain tumor pathogenesis.

Responses to elevated CO 2 have been studied using Agrostis capillaris L., an upland grass which is abundant on nutrient-poor soils. Plants were grown in sand culture with a wide range of nitrogen, phosphorus and potassium concentrations, and the impact of CO 2 on the demand for nutrients was determined using isotopic root bioassays. Plants grown with the smallest concentrations of N and P showed typical foliar symptoms associated with deficiency of these elements. However, even when supplies of N and P were limiting to growth, additional CO 2 (250 ppm above ambient) influenced neither total N nor total P in above-ground tissues, nor nutrient demands as indicated by the bioassay. The estimates of the demand of the plants for K from the 86 Rb bioassay indicated an appreciable increase when plants were raised in elevated CO 2 . For plants of the same size with the same nutrient supply, those grown in elevated CO 2 consistently displayed an increased internal demand for K. Uptake of K was not however, enhanced by elevated CO 2 even in non-limiting conditions and it might therefore be limited by a factor other than K supply. The overall conclusion from the experiments is that when A. capillaris is grown in elevated CO 2 , uptake of N, P and K fails to increase proportionally with dry mass. This was true even when nutrient supplies were adequate, and it appears that nutrient-use-efficiency might increase to enable the plants to maintain growth in elevated CO 2 .

Bud break and growth of beech trees were measured during the growing season of 1992. The trees had previously been exposed, through the growing season of 1991, to one of four treatments: (1) episodic ozone for 128 d, total dose = 46-3 ppm h; (2) air purified by filtration through Purified and charcoal; (3) ozone together with a period of water shortage: (4) filtered air with a period of water shortage. Despite uniform growth conditions during 1992. the previous water stress treatment caused bud break to begin slightly earlier, but the rates of shoot growth and the length of the first flush branches were reduced (40%). In well-watered plants ozone caused a reduced Tate of shoot growth during the first week after bud break. The total amount of growth during 1992 was shown to be reduced (36%) by the previous water stress treatment, In the case of the well-watered trees, exposure to ozone in the previous year reduced the amount of new growth by 17%, which was the result of reduced internal expansion. When ozone was combined with water stress, there was no further reduction in the amount of new growth, but there was a fall in the number of internodes relative to water-stressed plants grown in filtered air. Ozone and water stress applied singly reduced growth. A combination of the two stresses, however, caused no additional reduction in growth, but reduced the number of internodes. The possible implications of the aftereffects of these stresses are discussed in relation to reduced productivity and canopy structure.

The response of photosynthetic gas exchange to oxides of nitrogen (NOx) was studied in leaves of lettuce (Lactuca sativa L.) at different temperatures. Exposure to > high concentrations (e.g. 13 μmol NO x mol -1 ), similar to those often found in commercial glasshouses, caused a rapid inhibition of the net assimilation of CO 2 . This appeared to be by a direct effect on photosynthesis rather than by a change in the stomatal conductance. In ambient CO 2 , (345 μmol mol -1 ), the percentage inhibition at 10 and 5°C was approximately 3× and 5×, respectively, that measured at 20°C. This effect of temperature also occurred when measured in CO 2 , enriched air (1050 μmol mol -1 ), which would normally accompany NO x in a glasshouse. The extent of photosynthetic inhibition caused by NO x was, however, always less in high than in low CO 2 . The results suggest that when burning fuel to raise the CO 2 , concentration and heat the glasshouse air, growers should avoid generating high concentrations of NO x , in conditions of low temperature.

The effects of a mixture of SO(2) and NO(2) at a concentration of 0.08 to 0.10 ppm (in microg m(-3): 164-205 for NO(2); 229-286 for SO(2)) were tested for four weeks on two old varieties of winter wheat, Little Joss and Holdfast, introduced in 1908 and 1938, respectively, grown in simulated autumn conditions. After two weeks, root dry weights of both varieties were significantly decreased but, although there was some leaf necrosis, shoot dry weights were unaltered. After a further two weeks, the dry weights of the shoots as well as the roots were significantly decreased. These effects, amounting to a combined 40% decrease in total dry weight, were greater than those in identical experiments carried out with the present day variety Avalon, in which the decrease was 20%. The results suggest that the modern cultivar Avalon is more tolerant of SO(2) and NO(2) than Little Joss and Holdfast, which were grown many years ago.

Polyamines (PA) are known to be involved in the areas of plant physiology and biochemistry which are related to the response of a plant to air pollution. This study examines the role of arginine decarboxylase (ADC), an important rate-limiting enzyme in polyamine synthesis, in barley plants exposed to ozone (O(3)). The activity of ADC increased significantly in O(3)-treated leaves when visible injury was hardly apparent. The increase in ADC activity may be a mechanism to increase the PA levels in O(3)-treated leaves and so minimize the damaging effects of O(3). Supporting this, foliar applications of DL-alpha-difluoromethylarginine (DFMA), a specific inhibitor of ADC, prevented the rise in ADC activity and visible injury was considerable on exposure to O(3). This damage was not due to the foliar sprays, as little visible injury was seen in leaves in the O(3)-free controls. The results are discussed in terms of the roles of PA in conferring O(3) resistance in plants.

A new large-scale closed chamber fumigation system with cooling facilities is described for studying effects of low concentrations of SO(2), NO(2) and O(3) and low temperatures on woody species and herbaceous plants. The system is based on modified hemispherical greenhouses with a forced air ventilation system. This provides a chamber environment with low spatial variability of pollutant gas concentrations and rapid air circulation which allows exposure of plants at near ambient temperatures and relative humidity. Large capacity cooling units come into operation when ambient temperatures fall below 0 degrees C, and these allow chamber temperatures to be lowered by an additional 4 to 8 degrees C in experiments designed to test whether exposure to pollutants enhances the frost sensitivity of plants.

Sorghum plants that had been subjected to different degrees of water stress were examined for the occurrence of endogenous compounds capable of inducing stomatal closure, i.e. "antitranspirants". Acidic extracts contained increased amounts of abscisic acid (ABA) as the amount of stress increased, but another highly active compound easily distinguished from ABA also accumulated. This compound, also found in neutral extracts, was probably all trans-farnesol, an isoprenoid alcohol which, like ABA, is a sesquiterpenoid. Highly dilute solutions of "commercial" farnesol induced stomatal closure when applied to isolated epidermis of Commelina.

Experiments are described which attempted to clarify the reasons for differences in stomatal opening on adaxial and abaxial epidemics. Previous studies had suggested that endogenous auxin levels might be responsible. It was found that adaxial stomata were more sensitive than abaxial stomata to externally supplied abscisic acid (ABA). It is suggested that differing endogenous concentrations of calcium ions in the upper and lower epidermes may contribute to the observed differences in sensitivity to ABA. It is further suggested that studies of stomata on isolated epidermis may represent a useful model system for examining the role of cell or tissue sensitivity in determining the responses to ABA and other hormones. Data are presented which lend support to the suggestion that calcium ions play an important part. Treatment with lanthanum ions, which block calcium channels in cell membranes, had little effect on abaxial stomata, but caused the apertures of adaxial stomata to increase considerably. The chelating agent EGTA eliminated the differential behaviour of adaxial and abaxial stomata, which clearly suggests that calcium ions have a dominant role. Experiments in which EGTA and fusicoccin were presented separately and in combination indicated that their effects were independent and additive. It is suggested that they operate at different locations, or on different processes in the guard cells.

The effects of SO 2 and NO 2 on the control of water loss by birch trees were investigated using clonal populations of both Betula pendula Roth, (silver birch) and Betula pubescens Khr. (downy birch). Plants were grown in controlled environments and were fumigated in a 2 × 2 factorial experiment with 65 nl l -1 SO 2 and/or NO 2 , or with equal concentrations of both SO 2 and NO 2 in four treatments, namely zero, 20, 40 and 60 nl l -1 . Excised leaves showed in increased rate of water loss approximately corresponding to the concentration of pollutant to which they had been previously exposed. When petroleum jelly was applied to the abaxial epidermal surface of polluted leaves the increased rate of water loss was not found, strongly implying that the damage had occurred only at this site, and not on the adaxial epidermis, which is virtually devoid of stomata. Measurements of gas exchange were made on B. pubescens exposed to the four doses of the mixture of the two gases. The rate of transpiration was approximately doubled as a result of the two higher pollution treatments, judging from measurements made later in clean air. When transpiration was measured in high and low CO 2 concentrations, polluted plants had a slightly reduced and more variable response to the change in CO 2 concentration. Both clones were also exposed first to the four doses of the gas mixture, and then to drought. There was a more rapid onset of water stress, accompanied by an earlier decrease in effective leaf area in the polluted plants, when compared with the unpolluted plants. The abaxial surface of frozen hydrated leaves of the Betula pubescens clone were examined at low temperature with a scanning electron microscope. Damaged epidermal cells were clearly visible on the polluted leaves and thought to be responsible for areas of wide open stomata. It is concluded that leaves of these species are less efficient in the utilization of water after a period of exposure to pollution and are less able to restrict water loss in a time of shortage.

Pea aphids feeding from birth to maturity on pea plants (Pisum sativum) exposed to SO(2) concentrations of 50 nl litre(-1) or 80 nl litre(-1) showed a significant 19% increase in the rate of nymph production during the reproductive period, compared to control aphids feeding on plants in charcoal-filtered air. The higher nymph production resulted in a mean 4.6% increase in the intrinsic rate of population increase (rm). In longer term glasshouse fumigation experiments pea aphid populations were, on average, 1.8 times greater on pea plants in ambient air plus 45 nl litre(-1) SO(2) than in ambient air alone. Aphid infestation in ambient air caused a 42% reduction in pea yield and affected most plant parameters adversely. Ambient air plus SO(2) had no direct effect on yield, but, in combination with aphid infestation, a further 10% reduction in yield was recorded.