WEEE management in Europe and China - A comparison.

Over the last years Europe and China have developed specific regulations to address the challenge of managing Waste Electrical and Electronic Equipment (WEEE). Households in today's urban China are similarly equipped with electrical and electronic appliances as households in European metropolitan areas, which in turn will lead to similar per capita generation rates in WEEE. While the challenge is a similar one, the systems, technologies and legislation in place in Europe and China are partly different, partly aligned to each other. In Europe WEEE collection is based on existing municipal structures. Additionally, retail and other take-back channels are in place. In China the informal sector dominates WEEE collection, being more competitive and flexible and offering pecuniary reimbursement to consumers. In Europe manual dismantling as a first treatment step has been gradually replaced by mechanical break up of appliances, followed by sorting out of hazardous and valuable components. In the subsequent second treatment level, cathode ray tubes are separated, whereby compound materials like motors and coils are mechanically treated, printed circuit boards go to special smelters, and plastics are separated and partly recycled. In China large formal dismantling capacities have been set up in recent years. There dismantling practices follow similar principles as in European plants; however, further processing is only partly implemented in Chinese recycling facilities. Specifically metallurgical treatment of printed circuit boards is still not existent in China. Companies selling electrical and electronic products within the EU are obliged to organise collection and treatment. This has led to a larger number of producer responsibility organisations. Financed and controlled by producers and importers, these systems aim to fulfil legal requirements at optimised costs subject to compliance with environmental standards and monitoring requirements. The Chinese system is built on a state controlled fund which subsidies formal recyclers. For these recyclers this financial support is essential to compete with informal recyclers, who operate at lower costs and do not necessarily comply with environmental standards.

The telomere profile distinguishes two classes of genetically distinct cutaneous squamous cell carcinomas.

The incidence of skin cancer is increasing worldwide and cutaneous squamous cell carcinomas (SCCs) are associated with considerable morbidity and mortality, particularly in immunosuppressed individuals ('carcinomatous catastrophy'). Yet, molecular mechanisms are still insufficiently understood. Besides ultraviolet (UV)-indicative mutations, chromosomal aberrations are prominent. As telomeres are essential in preserving chromosome integrity, and telomere erosion as well as aberrant spatial telomere distribution contribute to genomic instability, we first established telomere length profiles across the whole tissue and identified normal skin (10/30) harboring discrete epidermal sites (stem cell territories) of evenly short telomeres. Precancerous actinic keratoses (AKs) (17) and SCCs (27) expressed two telomere phenotypes: (i) tissue-wide evenly short to intermediate and (ii) longer and tissue-wide heterogeneous telomere lengths, suggesting two modes of initiation, with one likely to originate in the epidermal stem cells. Although tumor histotype, location, patient gender or age failed to distinguish the two SCC telomere phenotypes, as did telomerase activity, we found a trend for a higher degree of aberrant p53 and cyclin D1 expression with long/heterogeneous telomeres. In addition, we established an association for the short/homogeneous telomeres with a simpler and the heterogeneous telomeres with a more complex karyotype correlating also with distinct chromosomal changes. SCCs (13) from renal transplant recipients displayed the same telomere dichotomy, suggesting that both telomere subtypes contribute to 'carcinomatous catastrophy' under immunosuppression by selecting for a common set (3, 9p and 17q) and subtype-specific aberrations (e.g., 6p gain, 13q loss). As a second mechanism of telomere-dependent genomic instability, we investigated changes in telomere distribution with its most severe form of telomeric aggregates (TAs). We identified a telomere length-independent but progression-dependent increase in cells with small telomere associations in AKs (17/17) and additional TAs in SCCs (24/32), basal cell carcinomas (30/31) and malignant melanomas (15/15), and provide evidence for a reactive oxygen species-dependent mechanism in this UV-induced telomere organization-dependent genomic instability.

The human papillomavirus type 16 E5-protein modulates ligand-dependent activation of the EGF receptor family in the human epithelial cell line HaCaT.

The E5 open reading frame of the human papillomavirus type 16 encodes a transmembrane protein associated with the Golgi, ER, and plasma membranes. We have analyzed the effect of E5 expression on the activation of the EGF receptor family. We find that expression of the E5-protein strongly enhances EGFR activation in a ligand-dependent manner. This activation takes place immediately after addition of ligand, demonstrating that increased tyrosine phosphorylation cannot solely be due to an impaired downregulation of the receptors. Furthermore, this activation is not a result of impaired activity of EGFR-specific phosphatase through the E5-protein, as demonstrated by using inhibitors specifically blocking EGFR activation. In addition, treatment with EGF results in an enhanced activation of the ErbB2 receptor in E5-expressing cells. This superactivation must be a result of heterodimer formation between EGFR and ErbB2, since EGF is not a ligand for ErbB2. Finally, treatment of E5-expressing cells with HB-EGF shows no increased phosphorylation of the ErbB4 receptor, suggesting a specific effect of E5 on the activation of the different members of the EGFR family.

Human papillomavirus type 16 E5 protein (review).

The association of certain human papillomavirus (HPV) types with malignancies of the anogenital tract is well established. The virus type most frequently associated with cellular transformation is HPV 16, as has been shown in epidemiological studies. Its transforming capacity has also been demonstrated in many in vitro cell transformation experiments. The most potent oncogenes of HPV 16 are the E6 and E7 proteins, but the E5 protein, whose homologue is the main oncogene of bovine papillomavirus, has recently been identified as an oncogene also for HPV. On the basis of epidemiological and clinical data from tumor material as well as from in vitro data it has been suggested, that the HPV 16 E5 protein would have a function at the early stages of cervical carcinogenesis. The E5 protein enhances growth factor-mediated signal transduction to the nucleus and consequently augments cellular proliferation. Expression of the E5 protein enables the infected cell to escape growth control provided by surrounding cells by inhibiting gap junctional intercellular communication in epithelial cells. This viral oncogene seems to interfere with the control mechanisms of cellular growth and proliferation and thus facilitate the function of the E6 and E7 proteins and further steps towards epithelial cell transformation.

Promoter of the gene encoding the bovine catalytic subunit of cAMP-dependent protein kinase isoform C beta 2.

Genomic sequences flanking the 5' end of the cDNA encoding isoform C beta 2 of the catalytic subunit of bovine cAMP-dependent protein kinase were cloned, sequenced and analyzed for promoter activity and transcription initiation sites. A region of 913 bp upstream the translation initiator ATG was amplified from genomic DNA by vectorette polymerase chain reaction. In primer extension reactions and RNase protection assays, residues C (at position -91), T (-71) and G (-70) were found to serve as transcription initiation sites of the gene. Amplification products and sub-fragments thereof were ligated upstream of the reporter gene chloramphenicol acetyltransferase to test for promoter activity. Constructs were transiently transfected into a Chinese hamster ovary cell line which was shown to express endogenous C beta 2 mRNA. The genomic sequence upstream the C beta 2 cDNA does have promoter activity. The region from position -51 to -292 proved sufficient to drive efficient transcription of the reporter gene. The promoter is AT rich (68%), does not contain a TATA box within 50 bp upstream of the first initiation site and possesses putative binding sites for several transcription factors such as PEA-3 and a glucocorticoid receptor.

Specific expression in adult mice and post-implantation embryos of a transgene carrying the histone H1(0) regulatory region.

Histone H1(0), a variant of the H1 group, has been found associated with the repressed state of chromatin and its content is increased in terminally differentiated cells. We have cloned a mouse H1(0) histone gene and introduced the promoter region, ligated to the beta-galactosidase reporter gene, into transgenic mice. By histochemistry we demonstrated a strong expression of the transgene in adult kidney, testis and brain. Intestine, uterus and ovarium were also positive. This expression followed the same pattern as that of the endogenous H1(0) gene, as demonstrated by in situ hybridization with a non-coding fragment of the mRNA, by Northern analysis, and by immunofluorescence with specific antibodies. In post-implantation embryos, the expression was very low up to day ten p.c. At this time, most of the X-Gal staining was found in the brain, retina and some of the large blood vessels. Hence, expression of the transgene as well as of the endogenous H1(0) gene is not exclusively linked to a differentiated phenotype or to a reduced cell proliferation capacity.

Basal level transcription of the histone H1(0) gene is mediated by a 80 bp promoter fragment.

The replacement histone H1(0) of the H1 group, known to interact with general transcription factors, has been found associated with transcriptionally repressed chromatin. Transcription of the gene in F9 stem cells is low but can be stimulated by treating the cells with retinoic acid. Using mutant deletions, we now demonstrate that basal level transcription in F9 cells is mediated by an 80 bp DNA fragment, located 430 bp upstream of the TATA box, which does not include the retinoic acid responsive element (RARE) known to bind retinoic acid receptors and stimulate transcription from an heterologous promoter after retinoic acid treatment. By footprinting, DMS interference, site-directed mutagenesis and UV-cross linking techniques we demonstrate that at least two nuclear factors, with MW of 90,000 and 30,000, bind to the 80 bp fragment and that this binding is necessary for transcription. Furthermore, positioning of this fragment upstream of the HSV-tk gene promoter stimulates transcription 2-3 times over control values, far less than the activity observed for this fragment in the homologous promoter, indicating that full activity of this fragment requires sequences located in the proximal part of the promoter.

Genomic rearrangements of retroviral vectors carrying two genes in F9 EC cells.

We have used two classes of double-expression retroviral vectors for the expression of foreign genetic information in embryonal carcinoma cell lines. The splice-vector pM5neo takes advantage of mutated sequences that mediate an LTR-driven expression in F9 EC cells. The second vector (pXT1 type) uses an internal HSV-tk promoter as the control element for the transcription of the second gene. Genomic analysis of DNA from infected F9 cell lines revealed that most of the proviruses have rearranged upon integration into the host genome. This reorganization always included the nonselected gene and is sequence independent, but depends on the selective pressure applied. No retroviral genomic rearrangements were observed in F9 cells infected with pM5 proviruses carrying only the neo resistance gene. On the contrary, gross rearrangements were found in cells infected with parental pXT1 retroviruses. In both vectors the transcriptional activity was very low. A direct correlation between selective pressure, proviral reorganization, and transcription was observed.

Multiple cis-acting elements of the proximal promoter region are required for basal level transcription of the H1(0) histone gene.

Basal level transcription of the mouse histone H1(0) gene is mediated by 531 base pairs of the promoter region. Deletion of the most distal upstream 80 bp of this fragment reduces transcription to very low values. By in vitro footprinting we demonstrate now that multiple factors bind to the DNA fragment localized between the 80 bp and the cap nucleotide. In addition to the presence of motifs for the binding of SP1, H1-box, H4TF-2 and TATA-box-factors, other not yet described protein-binding elements were identified. Internal deletions in the wild type promoter enclosing these motifs strongly restrict transcription. Furthermore, when one of these motifs was modified by site-directed mutagenesis a strong impairment of transcription followed. Thus for basal level transcription, in addition to the 80 bp distal fragment, cis-acting elements localized in the 450 bp proximal promoter region are required.

Transcription of BPV-1 genes in transfected F9 cells.

In F9 cells transformed with bovine papillomavirus type 1 (BPV-1) sequences two different phenotypes can be recognized. One cell type shows the characteristics of the parental stem cell line, whereas the other comprises cells with spindle-like morphology that do not adhere to each other, similar to retinoic acid-treated F9 embryonal carcinoma cells. The phenotypically altered cells plate more efficiently than the stem cells, grow well in soft agar and show an extended lifespan in the differentiated stage. Both types of cells contain BPV-1 DNA sequences as episomes, but only the non-stem-like cells have RNA transcripts for the unspliced E5 reading frame as well as for the spliced E6/E4.

The F9-EC cell line as a model for the analysis of differentiation.

The teratocarcinoma stem cell line F9 has been widely used as a model for the analysis of molecular mechanisms associated with differentiation. This cell line has been considered to be nullipotent and able to differentiate into endodermal-like derivatives upon treatment with retinoic acid. Nevertheless, under definite culture conditions, F9 cells are able to differentiate into derivatives of all three germ layers. The F9 cells express characteristics of early mouse embryonal cells and possess all repression factors known to be present in cells of the early mouse embryogenesis. Induction of differentiation can be achieved not only by adding chemical agents to the culture medium but also by transfection of several oncogenic sequences. In somatic cell genetic experiments, immortalized, differentiated F9-like cells have been shown to express dominantly genes responsible for the appearance of the differentiated phenotype.

Differentiation of EC cells in vitro by the fluorescent dye Hoechst 33342.

The fluorescent dye Hoechst 33342 is able to differentiate F9 EC cells at low concentrations. This differentiation is accompanied by synthesis of large amounts of laminin, production of a well-developed cytoskeleton, disappearance of the SSEA-1 antigen, and synthesis of large amounts of fibronectin, all characteristics of the primitive endoderm. The dye immediately blocks the cells at the S/G2 phase of the cell cycle and produces a complete arrest in proliferation. This effect is not specific for the nullipotent F9 cell line, as multipotent EC cell lines like PCC3, P19, and PCC4 can also be easily differentiated into the same pathway by treatment with the Hoechst dye. In contrast, the dye has no remarkable effects on terminal differentiated, immortalized cells like NIH 3T3 or the parietal endoderm-like cell PYS-2.

The efficiency of water use in water stressed plants is increased due to ABA induced stomatal closure.

Gas exchange and abscisic acid content of Digitalis lanata EHRH. have been examined at different levels of plant water stress. Net photosynthesis, transpiration and conductance of attached leaves declined rapidly at first, then more slowly following the withholding of irrigation. The intercellular partial pressure of CO2 decreased slightly. The concentration of 2-cis(S)ABA increased about eight-fold in the leaves of non-irrigated plants as compared with well-watered controls. A close linear correlation was found between the ABA content of the leaves and their conductance on a leaf area basis. In contrast, the plot of net assimilation versus ABA concentration was curvilinear, leading to an increased efficiency of water use during stress. After rewatering, photosynthesis reached control values earlier than transpiration, leaf conductance and ABA content. From these data it is concluded that transpiration through the stomata is directly controlled by the ABA content, whereas net photosynthesis is influenced additionally by other factors.Possible reasons for the responses of photosynthesis and water use efficiency to different stress and ABA levels are discussed.