Expression analysis of five tobacco EIN3 family members in relation to tissue-specific ethylene responses.

Ethylene induces different sets of genes in different tissues and at different stages of development. To investigate whether these differential responses are caused by differential expression of members of the EIN3 family transcription factors, five tobacco family members were isolated. They can be divided into three subgroups, which is probably due to the amphidiploid nature of tobacco. In phylogenetic analysis, each of the subgroups clustered with one of the three tomato EIL proteins and all NtEILs proved to be most homologous to Arabidopsis EIN3 and EIL1. Although organ-specific ethylene responses have been observed before, northern blot analysis showed that all NtEILs were expressed in all organs. To study differential NtEIL expression at the cellular level, in situ hybridization was used on the tobacco ovary. It was found that different ovary tissues displayed variable ethylene-induced expression of two ethylene-responsive marker genes. By contrast, no differences were found in expression level or tissue-specificity for any of the NtEILs in the ovary, before or after ethylene treatment. This indicates that the organ and tissue-specific ethylene responses are not caused by differential expression of NtEIL family members. These results support a model in which the developmental signals that regulate the tissue-specific responses are integrated with the ethylene signal downstream of a common primary ethylene-signalling pathway.

Ethylene regulates the timing of anther dehiscence in tobacco.

We investigated the involvement of ethylene signaling in the development of the reproductive structures in tobacco ( Nicotiana tabacum L.) by studying flowers that were insensitive to ethylene. Ethylene-insensitivity was generated either by expression of the mutant etr1-1 ethylene-receptor allele from Arabidopsis thaliana or by treatment with the ethylene-perception inhibitor 1-methylcyclopropene (MCP). Development of ovaries and ovules was unaffected by ethylene-insensitivity. Anther development was also unaffected, but the final event of dehiscence was delayed and was no longer synchronous with flower opening. We showed that in these anthers degeneration of the stomium cells and dehydration were delayed. In addition, we found that MCP-treatment of detached flowers and isolated, almost mature anthers delayed dehiscence whereas ethylene-treatment accelerated dehiscence. This indicated that ethylene has a direct effect on a process that takes place in the anthers just before dehiscence. Because a similar function has been described for jasmonic acid in Arabidopsis, we suggest that ethylene acts similarly to or perhaps even in concurrence with jasmonic acid as a signaling molecule controlling the processes that lead to anther dehiscence in tobacco.

Regional localization of suspensor mRNAs during early embryo development.

We investigated gene activity within the giant embryos of the scarlet runner bean (Phaseolus coccineus) to gain understanding of the processes by which the apical and basal cells become specified to follow different developmental pathways after division of the zygote. We identified two mRNAs, designated G564 and C541, that accumulate specifically within the suspensor of globular-stage embryos. G564 mRNA accumulates uniformly throughout the suspensor, whereas C541 mRNA accumulates to a higher level within the large basal cells of the suspensor that anchor the embryo to the surrounding seed tissue. Both G564 and C541 mRNAs begin to accumulate shortly after fertilization and are present within the two basal cells of embryos at the four-cell stage. In contrast, at the same stage, these mRNAs are not detectable within the two descendants of the apical cell. Nor are they detectable within cells of the embryo sac before fertilization, including the egg cell. We used a G564/beta-glucuronidase reporter gene to show that the G564 promoter is activated specifically within the basal region and suspensor of preglobular tobacco embryos. Analysis of the G564 promoter identified a sequence domain required for transcription within the suspensor that contains several copies of a conserved motif. These results show that derivatives of the apical and basal cells transcribe different genes as early as the four-cell stage of embryo development and suggest that the apical and basal cells are specified at the molecular level after division of the zygote.

Tea and cancer prevention: an evaluation of the epidemiologic literature.

Animal and in vitro studies provide evidence of an anticarcinogenic potential of active ingredients in teas. This review encompasses epidemiologic studies of stomach, colon, and lung cancer as well as the evidence of a relationship between tea drinking and cancer at large in humans. Cohort studies do not suggest a protective role for tea drinking in the total risk of cancer. Site-specific studies reveal a more complex picture. The epidemiologic studies on tea drinking and stomach cancer do not justify claims of a cancer-protective effect. A protective effect of green tea on the development of colon cancer is suggested. The evidence regarding black tea is less clear, with some indication of a risk of colon or rectal cancer associated with regular use of black tea. The studies on tea and lung cancer also suggest an increased risk with increased tea consumption. The range and crude categorization of tea consumption, choice of control groups, and inadequate control for confounding might have obscured possible relationships. From the limited studies that suggest a favorable effect from tea, it is likely that benefits are restricted to high intakes in high-risk populations.

Functional dissection of the promoter of the pollen-specific gene NTP303 reveals a novel pollen-specific, and conserved cis-regulatory element.

Regulatory elements within the promoter of the pollen-specific NTP303 gene from tobacco were analysed by transient and stable expression analyses. Analysis of precisely targeted mutations showed that the NTP303 promoter is not regulated by any of the previously described pollen-specific cis-regulatory elements. However, two adjacent regions from -103 to -86 bp and from -86 to -59 bp were shown to contain sequences which positively regulated the NTP303 promoter. Both of these regions were capable of driving pollen-specific expression from a heterologous promoter, independent of orientation and in an additive manner. The boundaries of the minimal, functional NTP303 promoter were determined to lie within the region -86 to -51 bp. The sequence AAATGA localized from -94 to -89 bp was identified as a novel cis-acting element, of which the TGA triplet was shown to comprise an active part. This element was shown to be completely conserved in the similarly regulated promoter of the Bp 10 gene from Brassica napus encoding a homologue of the NTP303 gene.

Characterization of a pollen-specific cDNA clone from Nicotiana tabacum expressed during microgametogenesis and germination.

This report describes the isolation and characterization of a cDNA clone representing a gene specifically expressed in pollen. A cDNA library was constructed against mRNA from mature pollen of Nicotiana tabacum. It was screened differentially against cDNA from mRNA of leaf and of pollen. One clone, NTPc303, was further characterized. On northern blot this clone hybridizes to a transcript 2100 nucleotides in length. NTPc303 is abundant in pollen. Expression of the corresponding gene is restricted to pollen, because no other generative or vegetative tissue contains transcripts hybridizing to NTPc303. Expression of NTP303 is evolutionarily conserved: homologous transcripts are present in pollen from various plant species. The first NTP303 transcripts are detectable on northern blot at the early bi-nucleate stage and accumulate until the pollen has reached maturity. During germination and pollen tube growth in vitro new NTP303 transcripts appear. This transcription has been proved by northern blots as well as by pulse labelling experiments. Nucleotide sequence analysis revealed that NTPc303 has an open reading frame coding for a predicted protein of 62 kDa. This protein shares homology to ascorbate oxidase and other members of the blue copper oxidase family. A possible function for this clone during pollen germination is discussed.

Calcium intake and 28-year cardiovascular and coronary heart disease mortality in Dutch civil servants.

Data obtained from a general health examination in 1953-1954 of 2605 middle-aged Dutch civil servants were analysed to investigate the relation between dietary calcium and cardiovascular (CVD) and coronary heart disease (CHD) mortality. Calcium intake was assessed at baseline by a 1-week food frequency recall. Multivariate adjusted odds ratios (OR) were calculated using the highest quintile of calcium intake as the reference. No statistically significant associations were observed for low calcium intake in 15 and 28 years of follow-up in both men and women. For men, multivariate adjusted OR for the lowest quintile of calcium intake were 1.3 (95% confidence interval (CI): 0.8-1.9) and 0.9 (95% CI: 0.6-1.6) for 28-year CVD and CHD mortality, respectively. For women, corresponding OR were 1.1 (95% CI: 0.6-2.0) and 1.1 (95% CI: 0.5-2.5). Although an inverse association between calcium intake and CVD and CHD mortality, possibly mediated by blood pressure, might be hypothesized, no clear association was observed. Because dietary patterns in the 1950s were quite stable, and major calcium sources were addressed, misclassification of calcium intake may not be fully responsible for this finding.

Stage-related expression of mRNAs during pollen development in lily and tobacco.

Homogeneous populations of developing microspores and pollen from anthers of lily (Lilium longiflorum Thumb.) and tobacco (Nicotiana tabacum L.) show a continuous production of biomass, reaching a maximum in young pollen. The rate of RNA synthesis was 460 fg · h(-1) in young binucleate cells, 138 fg · h(-1) in late binucleate cells and 56 fg · h(-1) in microspores. The mRNA population in developing pollen can be separated into three groups. In the first group, certain types of mRNAs are present at a constant level during all stages of development. A second group is characteristic of young pollen and increases quantitatively until anthesis. A third group is seen transiently; to this belong mRNAs present only before mitosis or at a distinct cell stage after mitosis. Some of the translation products of this latter group of mRNAs showed similarities between lily and tobacco on two-dimensional gels in respect of molecular weight and isolectric point, indicating that those mRNAs and proteins play a role in the regulation of pollen development.

Physiologically-induced changes in proopiomelanocortin mRNA levels in the pituitary gland of the amphibian Xenopus laevis.

In the pars intermedia of the pituitary gland of the amphibian Xenopus laevis the level of mRNA encoding proopiomelanocortin (POMC), the precursor protein for alpha-melanophore-stimulating hormone (alpha-MSH), is shown to be dependent on physiological parameters. POMC mRNA levels in the pars intermedia of black-background-adapted Xenopus are much higher than those of white-adapted animals. These physiological changes in POMC mRNA levels are tissue-specific because they were not found in the pars distalis of the pituitary gland. Background transfer experiments revealed that modulation of POMC gene activity is much slower than changes in the secretion of alpha-MSH.

Unexpected 226Ra build-up in wet-process phosphoric-acid plants.

During an investigation of the distribution of radium and uranium over the different process streams in phosphoric-acid production and in the nitrophosphate route, a higher than average radiation level was detected in the vicinity of some piping and vessels near the gypsum filtres. This higher radiation level is caused by radium, which, as a decay product in the 238U series, is present in very low concentrations (1-50 pCi 226R/g) in the phosphates used for fertilizer production. Specific research later on established that there were detectable radiation levels around vessels and piping in other phosphoric-acid works as well. It turned out that in all factories the filter part, and especially the washing-acid section showed the highest radiation-level. In the piping of the gypsum filter, through which the washing acid is discharged, a scaling of bariumsulphate and calciumsulphate with a higher radiumconcentration (+/- 0.1 Ci/g) is formed. The radium ions appeared to be incorporated in the bariumsulphate lattice, hence the solubility of the radium precipitate is very low. The danger of internal pollution, for example in cleaning operations by radio-active intake or inhallation of radon is very small in the cases described. The risk of external radiation can be prevented by periodically cleaning (every 1-3 yr) the equipment in which the radio-active scaling is formed.