Assessing the impacts of citizen-led policies on emissions, air quality and health.

Air pollution is a global challenge, and especially urban areas are particularly affected by acute episodes. Traditional approaches used to mitigate air pollution primarily consider the technical aspects of the problem but not the role of citizen behaviour and day-to-day practices. ClairCity, a Horizon 2020 funded project, created an impact assessment framework considering the role of citizen behaviour to create future scenarios, aiming to improve urban environments and the wellbeing and health of its inhabitants. This framework was applied to six pilot cases: Bristol, Amsterdam, Ljubljana, Sosnowiec, Aveiro Region and Liguria Region, considering three-time horizons: 2025, 2035 and 2050. The scenarios approach includes the Business As Usual (BAU) scenario and a Final Unified Policy Scenarios (FUPS) established by citizens, decision-makers, local planners and stakeholders based on data collected through a citizen and stakeholder co-creation process. Therefore, this paper aims to present the ClairCity outcomes, analysing the quantified impacts of selected measures in terms of emissions, air quality, population exposure, and health. Each case study has established a particular set of measures with different levels of ambition, therefore different levels of success were achieved towards the control and mitigation of their specific air pollution problems. The transport sector was the most addressed by the measures showing substantial improvements for NO2, already with the BAU scenarios, and overall, even better results when applying the citizen-led FUPS scenarios. In some cases, due to a lack of ambition for the residential and commercial sector, the results were not sufficient to fulfil the WHO guidelines. Overall, it was found in all cities that the co-created scenarios would lead to environmental improvements in terms of air quality and citizens' health compared to the baseline year of 2015. However, in some cases, the health impacts were lower than air quality due to the implementation of the measures not affecting the most densely populated areas. Benefits from the FUPS comparing to the BAU scenario were found to be highest in Amsterdam and Bristol, with further NO2 and PM10 emission reductions around 10%-16% by 2025 and 19%-28% by 2050, compared to BAU.

Cancer incidence among Danish seafarers: a population based cohort study.

AIMS: Seafarers aboard oil and chemical tankers may be exposed to many chemicals, including substances like benzene that are known to be carcinogenic. Other seafarers are exposed to engine exhaust, different oil products, and chemicals used aboard and some years ago asbestos was also used extensively in ships. The aim of this study was to study cancer morbidity among Danish seafarers in relation to type of ship and job title. METHODS: A cohort of all Danish seafarers during 1986-1999 (33,340 men; 11,291 women) registered by the Danish Maritime Authority with an employment history was linked with the nationwide Danish Cancer Registry and followed up for cancer until the end of 2002. The number of person years at risk was 517,518. Standardised incidence ratios (SIR) were estimated by use of the corresponding national rates. RESULTS: The SIR of all cancers combined was higher than expected: 1.26 (95% CI 1.19 to 1.32) for men and 1.07 (95% CI 0.95 to 1.20) for women. This was mainly due to an excess of cancer of the larynx, lung, tongue, mouth, pharynx, oesophagus, pancreas, kidney, urinary bladder, colon, and bone as well as skin melanomas among men (the three latter borderline significantly increased), and an excess of cancer of the lung, rectum, and cervix uteri among women. The differences in risk pattern for lung cancer between the different job categories among men ranged in terms of SIR from 1.2 (95% CI 0.9 to 1.7) (engine officers) to 2.3 (1.6 to 3.3) (engine room crew), and 4.1 (2.1 to 7.4) among maintenance crew. Non-officers had a 1.5 times higher lung cancer risk than officers. No increased occurrence of all lymphatic and haematopoietic malignancies combined was found for employees on tankers, but the number of cases was limited to a total of 7. CONCLUSIONS: Danish seafarers, especially men, face an increased overall cancer risk, in particular a risk for lung cancer and other tobacco associated cancers.

HCf-6, a novel class II hydrophobin from Cladosporium fulvum.

C. fulvum, a fungal tomato pathogen, has previously been shown to express a complex family of hydrophobin genes including four class I hydrophobins and one class II hydrophobin. Here we describe a gene for HCf-6, a sixth member of the hydrophobin family and the second class II gene. The protein is predicted to consist of a signal sequence, an N-terminus rich in glycine and asparagine and a C-terminal hydrophobic domain which bears the hall-marks of hydrophobins. In contrast to the previously described class II hydrophobin HCf-5, HCf-6 is expressed in mycelium growing in pure culture and mRNA levels do not increase during sporulation. It is down-regulated by carbon starvation but not by depletion of nitrogen in the growth medium.

Transcriptional regulation of the Saccharomyces cerevisiae amino acid permease gene BAP2.

Uptake of branched-chain amino acids by Saccharomyces cerevisiae from media containing a preferred nitrogen source is mediated by the permeases encoded by BAP2, BAP3, and VAP1/TAT1. The transcriptional activity of the BAP2 promoter is affected by a number of genes, including SSY1, which encodes an amino acid permease homologue that is necessary for transcription of BAP2. Other genes that control BAP2 encode known (Leu3p, Tup1p) and putative (Stp1p, Stp2p) transcription factors. We present evidence that the zinc-finger proteins Stp1p and Stp2p bind directly to the BAP2 promoter. Binding of Stplp to the BAP2 promoter in vivo and in vitro indicates that the STP gene family indeed encodes transcription factors. The presence of a Leu3p binding site in the BAP2 promoter is required for full promoter activity on synthetic complete medium. The capacity of Leu3p to activate BAP2 transcription correlates with conditions that affect the level of alpha-isopropyl malate. The effect of a tup1 deletion on BAP2 transcription depends on SSY1. In an ssy1 strain, the phenotype of tup1 conforms to the well-established role of Tup1p as part of a repressor complex, but in the SSY1 strain deletion of TUP1 causes a decrease in transcription, indicating that Tup1p may also have an activating role at the BAP2 promoter. Our results thus suggest a complex interplay between several transcription factors in the expression of BAP2.

Developmental time of Blattisocius tarsalis (Acari: Ascidae) at different temperatures.

The developmental time of the predatory mite Blattisocius tarsalis (Berlese) (Acari: Ascidae) was investigated at temperatures of 15, 21 and 25 degrees C and 75% r.h. Eggs 1-3 days old of Ephestia kuehniella Zeller (Lep.: Pyralidae), killed by freezing, were used as food. Mean developmental times were found to be 22.4, 8.5 and 7.0 days, respectively. Within the investigated thermal limits the developmental rate showed a linear relationship with temperature and the corresponding thermal threshold for development was calculated to 10.2 degrees C.

Stp1p, Stp2p and Abf1p are involved in regulation of expression of the amino acid transporter gene BAP3 of Saccharomyces cerevisiae.

Expression of the BAP3 gene of Saccharomyces cerevisiae, encoding a branched chain amino acid permease, is induced in response to the availability of several naturally occurring amino acids in the medium. This induction is mediated via an upstream activating sequence (called UAS(aa)) in the BAP3 promoter, and dependent on Stp1p, a nuclear protein with zinc finger domains, suggesting that Stp1p is a transcription factor involved in BAP3 expression. In this paper, we show that Stp2p, a protein with considerable similarity to Stp1p, is also involved in the induction of BAP3 expression. To gain more insight into the roles of STP1 and STP2, we have overexpressed both Stp1p and Stp2p in yeast cells. Gel shift assays with the UAS(aa)as a probe show that the UAS(aa)can form two major complexes. One complex is dependent on Stp2p overexpression and the other is formed independently of STP1 or STP2, suggesting that the UAS(aa)is also bound by another factor. Here we show that the other factor is Abf1p, which binds specifically to the UAS(aa)of BAP3.

Assessment of IgE allergen specificity among latex-allergic health care workers: review of IgE-binding components of various latex extracts.

BACKGROUND: Allergic reactions to natural rubber latex have increased during the past 10 years, especially in many health care workers who have high exposure to latex allergens both by direct skin contact and by inhalation of latex particles from powdered gloves. Development of satisfactory diagnostic methods to verify the presence of latex allergy in health care workers requires characterization of the immunoreactive proteins in latex products and identification of specific IgE antibodies in sensitized patients. A number of different latex preparations are now available for in vitro evaluations. OBJECTIVES: Utilizing different in vitro methods, this study examines IgE sensitization to components of latex in a selected population of hospital employees, employing a raw natural latex glove extract and various commercial latex extracts. METHODS: Two hundred hospital employees exposed to latex were evaluated using an allergy history questionnaire. To further identify sensitized patients, two different specific IgE tests and leukocyte histamine release tests were performed using a panel of latex extracts obtained from different manufacturers. Sodium dodecylsulfate polyacrylamide electrophoresis (SDS-PAGE) profiles were obtained. Sera from 34 subjects suspected to be latex-sensitized were IgE immunoblotted to assess the presence of IgE antibodies directed toward specific latex proteins. RESULTS: Thirty-four participants (17%) were considered sensitized to latex by a positive clinical history in conjunction with positive specific IgE tests (18 individuals) and/or positive histamine release tests (26 individuals). Significant extract differences in both the histamine release response profile and the frequency of positive test results were noted in the histamine release test. Significant individual differences in patients' latex epitope-specificity were found by IgE immunoblotting, substantiated by sodium dodecylsulfate polyacrylamide profiles revealing differences in protein band patterns among the various extracts. The IgE immunoblots indicated that the majority of patients reacted to proteins with molecular weights of 14, 21, 30 to 35, and 42 kD; the 30 to 35 kD protein being predominant. Seven subjects (22%) of the 34 considered to be latex-sensitized did not reveal binding of specific IgE in immunoblots. One latex extract (Stallergene) with the widest IgE-reacting protein repertoire identified the majority of subjects (63%) as latex sensitive by leukocyte histamine release and also provided the best quantitative histamine release test results. CONCLUSION: Only by testing with a combination of latex extracts were all sensitized individuals identified. This study demonstrates that currently several in vitro methods may be necessary to detect IgE sensitization to latex. Latex extracts to be employed in future skin tests must contain a wide epitope repertoire of IgE-binding proteins to identify all latex-sensitized individuals.

Methionine synthase, a gene required for methionine synthesis, is expressed in planta by Cladosporium fulvum.

Abstract The nutritional requirements of phytopathogenic fungi growing in planta has to date been largely ignored. We have begun to address this problem by investigating the methionine requirement for the biotrophic pathogen of tomato Cladosporium fulvum during infection. The Met6 gene from Cladosporium fulvum encoding a cobalamin-independent 5-methyltetrahydropteroyltriglutamate-homocysteinemethyltransferase, was cloned by functional yeast complementation. The open reading frame was found to be 2304 bp, containing no introns and encoding a protein of 87 kDa. In vitro Northern analysis demonstrated high levels of Met6 expression in the absence of externally supplied methionine. However in the presence of methionine or in the absence of carbon, expression of Met6 decreased significantly. Analysis of Met6 expression in planta revealed a strong increase during infection suggesting the requirement for methionine synthesis in planta by Cladosporium fulvum. This study demonstrates that Cladosporium fulvum is starving for methionine during infection and thus implies the essentiality of primary biosynthetic pathways to the infecting fungus.

Chromosomal aberrations in humans induced by urban air pollution: influence of DNA repair and polymorphisms of glutathione S-transferase M1 and N-acetyltransferase 2.

We have studied the influence of individual susceptibility factors on the genotoxic effects of urban air pollution in 106 nonsmoking bus drivers and 101 postal workers in the Copenhagen metropolitan area. We used the frequency of chromosomal aberrations in peripheral blood lymphocytes as a biomarker of genotoxic damage and dimethylsulfate-induced unscheduled DNA synthesis in mononuclear WBCs, the glutathione S-transferase M1 (GSTM1) genotype, and the N-acetyltransferase 2 (NAT2) genotype as biomarkers of susceptibility. The bus drivers, who had previously been observed to have elevated levels of aromatic DNA adducts in their peripheral mononuclear cells, showed a significantly higher frequency of cells with chromosomal aberrations as compared with the postal workers. In the bus drivers, unscheduled DNA synthesis correlated negatively with the number of cells with gaps, indicating a protective effect of DNA repair toward chromosome damage. Bus drivers with the GSTM1 null and slow acetylator NAT2 genotype had an increased frequency of cells with chromosomal aberrations. NAT2 slow acetylators also showed elevated chromosomal aberration counts among the postal workers. Our results suggest that long-term exposure to urban air pollution (with traffic as the main contributor) induces chromosome damage in human somatic cells. Low DNA repair capacity and GSTM1 and NAT2 variants associated with reduced detoxification ability increase susceptibility to such damage. The effect of the GSTM1 genotype, which was observed only in the bus drivers, appears to be associated with air pollution, whereas the NAT2 genotype effect, which affected all subjects, may influence the individual response to some other common exposure or the baseline level of chromosomal aberrations.

[Microdialysis. A method for measurement of local tissue metabolism]. / Mikrodialyse. En metode til måling af lokal vaevsmetabolisme.

Studies of pharmacological, immunological, and biochemical reactions in intact tissues have been hampered by the lack of appropriate techniques. The purpose of the review is to give a short introduction to the microdialysis technique which permits measurement of low molecular weight compounds in the extracellular water compartment in different tissue. Originally developed for use in animal brain, microdialysis is now being applied in humans for both clinical and experimental studies in different tissues, e.g. subcutaneous adipose tissue, brain, skin, and skeletal muscle. This review summarizes theoretical and practical aspects of microdialysis, and describes its main applications in humans.

Biomarkers for exposure to ambient air pollution--comparison of carcinogen-DNA adduct levels with other exposure markers and markers for oxidative stress.

Human exposure to genotoxic compounds present in ambient air has been studied using selected biomarkers in nonsmoking Danish bus drivers and postal workers. A large interindividual variation in biomarker levels was observed. Significantly higher levels of bulky carcinogen-DNA adducts (75.42 adducts/10(8) nucleotides) and of 2-amino-apidic semialdehyde (AAS) in plasma proteins (56.7 pmol/mg protein) were observed in bus drivers working in the central part of Copenhagen, Denmark. In contrast, significantly higher levels of AAS in hemoglobin (55.8 pmol/mg protein), malondialdehyde in plasma (0. 96 nmol/ml plasma), and polycyclic aromatic hydrocarbon (PAH)-albumin adduct (3.38 fmol/ microg albumin) were observed in the suburban group. The biomarker levels in postal workers were similar to the levels in suburban bus drivers. In the combined group of bus drivers and postal workers, negative correlations were observed between bulky carcinogen-DNA adduct and PAH-albumin levels (p = 0.005), and between DNA adduct and [gamma]-glutamyl semialdehyde (GGS) in hemoglobin (p = 0.11). Highly significant correlations were found between PAH-albumin adducts and AAS in plasma (p = 0.001) and GGS in hemoglobin (p = 0.001). Significant correlations were also observed between urinary 8-oxo-7, 8-dihydro-2'-deoxyguanosine and AAS in plasma (p = 0.001) and PAH-albumin adducts (p = 0.002). The influence of the glutatione S-transferase (GST) M1 deletion on the correlation between the biomarkers was studied in the combined group. A significant negative correlation was only observed between bulky carcinogen-DNA adducts and PAH-albumin adducts (p = 0.02) and between DNA adduct and urinary mutagenic activity (p = 0.02) in the GSTM1 null group, but not in the workers who were homozygotes or heterozygotes for GSTM1. Our results indicate that some of the selected biomarkers can be used to distinguish between high and low exposure to environmental genotoxins.

Barley elongation factor 1 alpha: genomic organization, DNA sequence, and phylogenetic implications.

A full length cDNA clone encoding the 447 amino acid long barley (Hordeum vulgare cv. Bomi) endosperm elongation factor 1 alpha (eF-1 alpha) was isolated by a differential screening procedure. RFLP mapping of eF-1 alpha showed that the barley genome contains a small eF-1 alpha gene family of 4 copies, with 1 copy of the gene being located on each of chromosomes 2, 4, 6, and 7. Analysis of barley endosperm total proteins by Western blot with antibodies directed towards wheat eF-1 alpha and the sea urchin 51 kDa proteins gave a single band of the expected molecular weight. Amino acid sequence comparison with other plant eF-1 alpha sequences showed that the isolated barley endosperm eF-1 alpha is more similar to the published wheat eF-1 alpha sequence than to eF-1 alpha sequences previously published for the barley cultivars Igri and Dicktoo. The phylogenetic analysis suggests that the barley eF-1 alpha gene family can be divided into two subfamilies and that two ancestral genes existed before the divergence of monocotyledonous and dicotyledonous plants.

Biomonitoring of diesel exhaust-exposed workers. DNA and hemoglobin adducts and urinary 1-hydroxypyrene as markers of exposure.

Diesel exhaust-exposed workers have been shown to have an increased risk of lung cancer. A battery of biomarkers were evaluated for their ability to assess differences in exposure to genotoxic compounds in bus garage workers and mechanics and controls. Lymphocyte DNA adducts were analyzed using the 32P-postlabelling method with butanol and P1 enrichment procedures. Hydroxyethylvaline (HOEtVal) adducts in hemoglobin were measured by gas chromatography-mass spectrometry (GC-MS) and 1-hydroxypyrene (HPU) in urine determined using HPLC analysis. The exposed workers had significantly higher levels of all three biomarkers compared to the controls. Total DNA adduct levels were 0.84 fmol/micrograms DNA vs 0.26 in controls (butanol) and 0.65 fmol/micrograms DNA vs. 0.08 (P1 nuclease). Median HOEtVal adduct level in exposed workers was 33.3 pmol/g hemoglobin vs. 22.1 in controls. HOEtVal adducts correlated with HPU but not with DNA adducts. The levels of HPU in urine were 0.11 micromol/mol creatinine compared to 0.05 in controls. All three assays applied were sensitive enough to evaluate a low level of exposure to environmental pollutants, with postlabelling and GC-MS as the most sensitive assays. The study indicated that skin absorption of polycyclic aromatic hydrocarbons (PAH) might be an important factor to consider when studying PAH exposure from air pollution sources.

Comparative synchronous fluorescence spectrophotometry and 32P-postlabeling analysis of PAH-DNA adducts in human lung and the relationship to TP53 mutations.

Polycyclic aromatic hydrocarbon (PAH)-DNA adducts were studied in human lung from 39 lung cancer patients by synchronous fluorescence spectrophotometric (SFS) and 32P-postlabeling assays. Regression analysis of the samples failed to detect any correlation between benzo[a]pyrene-diolepoxide (BPDE)-DNA adducts detected by SFS and the BPDE co-migrating spot detected by 32P-postlabeling. We have also analyzed the relationship between adduct levels and TP53 mutations. By postlabeling diagonal radioactive zone (DRZ) adducts were detected in 37 of 39 (95%) lung tissues from lung cancer patients and the adduct level ranged from 6.81 to 108.50 adducts/10(8) nucleotide. Thirty-three of 39 (85%) had detectable levels of BPDE-DNA adducts (> 1 adduct/10(9) nucleotide). Current heavy smokers (> 20 cigarettes/day) have significantly higher DRZ adduct levels compared to individuals smoking less than 20 cigarettes/day. By SFS combined with immunoaffinity column (IAC), 11 of 39 (28%) samples had detectable adduct levels, and 6 of 11 (55%) were detectable by SFS following purification of benzo[a]pyrene (BP)-tetrols by high pressure liquid chromatography (HPLC). Six of 33 (18%) samples were positive for BPDE-DNA adducts by both postlabeling and HPLC/SFS. No correlation was observed between the SFS and 32P-postlabeling assays for the detection of BPDE-DNA adducts. However, there was a good correlation between adduct levels detected by IAC/SFS and HPLC/SFS. We found a weak association between total PAH-DNA adduct levels in lung tissue and TP53 mutations.

Environmental air pollution and DNA adducts in Copenhagen bus drivers--effect of GSTM1 and NAT2 genotypes on adduct levels.

The lymphocyte bulky PAH-DNA adduct levels have been studied in persons occupationally exposed to ambient air pollution. The exposure group consisted of 90 healthy, nonsmoking bus drivers from the Copenhagen area, divided into three exposure groups according to driving area, and 60 rural controls (smokers and non-smokers). PAH-DNA adducts were determined by 32P-postlabelling with the butanol enrichment procedure. The bus drivers answered a comprehensive questionnaire on passive smoking, residential area, diet and other potential confounding variables. A significantly higher adduct level was observed in bus drivers working in central Copenhagen (1.214 fmol/microg DNA, n = 49) compared with both those driving in the dormitory (median: 0.507 fmol/microg DNA, P = 0.046, n = 16) and suburban (median: 0.585 fmol/microg DNA, P = 0.041, n = 25) areas. All three groups had higher adduct levels than rural controls (0.074 fmol/microg DNA, n = 60, P < 0.001). No significant influence on adduct levels was demonstrated from potential confounders, including smoking and diet. The effect of the metabolizing enzymes, GSTM1 and NAT2, on adduct levels was investigated. No statistically significant effects were observed on adduct levels from GSTM1 or NAT2, either individually or combined, but a non-significant trend was seen for individuals with GSTM1*0/0 (null), since they had higher adduct levels in all exposure groups. This study demonstrated that lymphocyte PAH-DNA adduct levels were related to levels of exposure to urban air pollution and indicated that these adducts might be helpful as a means of classifying better different exposure groups for epidemiological studies. Furthermore, it demonstrated the ability of 32P-postlabelling to discern small differences in low exposure to ambient air pollution and suggested a possible effect of GSTM1*0/0 on DNA adduct levels.

Exposure to urban and rural air pollution: DNA and protein adducts and effect of glutathione-S-transferase genotype on adduct levels.

Ambient air in urban areas is polluted by agents suspected of causing cancer in humans. A number of epidemiological studies have revealed an increased cancer risk in urban communities, especially in lung cancer. The relative risk have been estimated to be in the order of 1.5. The objective of this study was to evaluate differences in genotoxic exposure through air pollution in urban and rural areas using DNA and protein adducts as biomarkers. Another objective was to investigate whether the GSTM1 genotype has any effect on adduct level. The analyses included 32P postlabelling of DNA adducts in lymphocytes, enzyme-linked immunosorbent assay for measuring benzo[a]pyrene protein adducts and polymerase chain reaction amplification of the GSTM1 genotype. The study was a cross-sectional study of non-smoking, healthy males from rural and urban Danish areas and from Athens, Greece. All individuals in the study were healthy, non-smoking males. The Danish urban group included 74 university students, the rural group 29 students from agricultural colleges and the Greek group 17 individuals. Adduct levels differed significantly in the three groups with median levels of 0.152 fmol/micrograms DNA (rural), 0.205 fmol/micrograms (urban) and 0.285 fmol/micrograms (Athens). The adduct patterns showed some identical spots, but also specific adducts. Here we report increasing DNA adduct levels comparing residents in rural, small urban and large urban residential areas; we found no influence of GSTM1 genotype on DNA or protein adduct levels in non-smokers exposed to low levels of air pollution.

The promoter of the barley aleurone-specific gene encoding a putative 7 kDa lipid transfer protein confers aleurone cell-specific expression in transgenic rice.

This paper describes the aleurone-specific gene Ltp2 from barley, which encodes a putative 7 kDa non-specific lipid transfer protein. As shown by Northern and in situ hybridization analyses, the Ltp2 transcript is present in barley aleurone cells shortly after the initiation of aleurone cell differentiation. The expression of Ltp2 increases until grain mid-maturity, but the mRNA is absent from mature grains. The Ltp2 transcript is undetectable in the embryo and vegetative tissues, confirming the aleurone specificity of the Ltp2 gene. The ability of the isolated 801 bp Ltp2 promoter to direct aleurone-specific expression in immature barley grains is demonstrated by particle bombardment experiments. In these experiments, the activity of the Ltp2 promoter is 5% of the activity of the strong constitutive Actin1 promoter from rice, as quantified by GUS activity measurements. In stably transformed rice plants containing the Ltp2 promoter-Gus construct, the specificity of the Ltp2 promoter is confirmed in vivo by the presence of GUS activity exclusively in the aleurone layer. This study demonstrates the conserved nature of the regulatory signals involved in aleurone-specific gene transcription in cereal grains.

In vitro binding of nuclear proteins to the barley plastocyanin gene promoter region.

Plastocyanin is a nuclear-encoded chloroplast protein participating in electron transport during photosynthesis. The plastocyanin gene is expressed in photosynthetic tissue in a developmentally regulated manner and the expression is stimulated by light. A genomic clone encoding the plastocyanin precursor was isolated from a barley (Hordeum vulgare) lambda library using a barley cDNA clone as a probe and the sequence of a 1.9-kb DNA fragment containing the plastocyanin gene was determined. TATA and CCAAT boxes are located 34-bp and 68-bp, respectively, upstream of the transcription start site, the 5'-untranslated leader is 78 nucleotides long, and the intronless gene has at least two different polyadenylation sites. DNA sites in the plastocyanin gene that mediate binding of barley nuclear proteins were mapped by mobility-shift assays with fragments of the promoter/upstream region. Two of the three specific binding sites characterised in more detail were found to form complexes with the same factor in cross-competition experiments. One of these sites, narrowed down to a 17-bp sequence at position -512, contains the consensus binding site for Myb-like transcription factors. The third specific binding site, located at position -622, contains the sequence CACGTG which is a high-affinity-binding site for transcription factors of the basic-region leucine-zipper family.