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Non-Rabl patterns of centromere and telomere distribution in the interphase nuclei of plant cells.

At the anaphase of cell divisions, the divided chromosomes move to the two poles, with the centromeres as heads and telomeres as tails. Such a polarized orientation of centromeres and telomeres is believed to be preserved in the interphase and is known as Rabl model. We analyzed the distributions of centromeres and telomeres in interphase nuclei from several plant species. Although Rabl polarity was observed in wheat, rye, barley and oats, non-Rabl patterns were discovered in sorghum, rice and maize. In the non-Rabl patterns, both centromeres and telomeres were dispersed throughout the interphase nucleus, except in the area occupied by the nucleolus. Both Rabl and non-Rabl distribution patterns of centromeres and telomeres were consistent in interphase nuclei derived from meristematic root tip cells, microspore mother cells and differentiated leaf cells. Our study demonstrated that there is a diversity of interphase chromatin organization and that the classical Rabl model is not universal in plant species.

Avena↗

Modification of wheat rDNA loci by rye B chromosomes: a chromatin organization model.

The rDNA loci, and their associated NORs, on chromosomes 1B and 6B of the hexaploid wheat cv. Lindström have been used as a chromatin marker to investigate the functional basis of the phenotype effects of introgressed supernumerary B chromosomes (Bs) of rye. The rye Bs themselves lack genes, other than those which determine their mitotic drive mechanism, and the way in which they can modulate characters determined by the A chromosome background has always been a puzzle. An isogenic line of Lindström plants carrying different numbers of Bs was used as the experimental system to see how different doses of Bs (from 0 to 6) affected the activity of the wheat NORs and the organization of their rDNA loci at interphase. Silver staining on metaphase chromosomes was used to evaluate the previous activity of the NORs, and to reveal variations in their size; and the pTa71 FISH probe from wheat was used to visualize structural modifications to the interphase rDNA loci. A single B had no measurable effect, but, as the B-number increased, there were significant changes in the physical dimensions of the metaphase NORs, reflecting reduced levels of their activity earlier in the cell cycle, and also in the condensation patterns of the interphase rDNA loci. In addition, the higher B-numbers caused a size heteromorphism between the homologous NORs. A model is discussed which interprets the phenotypic effects of Bs generically, in nucleotypic terms, based of their being 'genetically inert' but 'chromosomally active'.

Chromatin↗

Rye chromosome variability in wheat-rye addition and substitution lines.

In a study of polymorphism and stability in rye chromosomes, three rye varieties and the sets of wheat-rye addition and substitution lines were compared using two non-homologous highly repetitive DNA families, pSc200 and pSc250. The rye varieties, Petkus, Imperial and Onohoiskaya, showed polymorphism for the presence and the size of the pSc200 in-situ hybridization signals on chromosome pairs, 2R, 4R and 7R, and the pSc250 signals on chromosomes, 5R, 6R and 7R. Chromosome 1R was heteromorphic within the Onohoiskaya variety. Differences in the distribution of chromosome polymorphisms imply that intervarietal changes to these highly repetitive DNA families occurred independently, despite their juxtaposition or even overlapping locations in subtelomeric heterochromatic regions. In the set of Saratovskaya 29 wheat/Onohoiskaya substitution lines, only chromosome 2R was altered relative to its counterpart in the parental rye variety due to amplification of the pSc250 signal on the long arm, although this did not exceed intervarietal polymorphism. In the set of Chinese Spring wheat/Imperial addition lines, only two Imperial chromosomes, 4R and 6R, were unchanged. We detected the loss of one or both rye homologous chromosomes, the loss of one arm, and the deletion of subtelomeric heterochromatin accompanied by the loss of the pSc200 signal. The results show that Saratovskaya 29/Onohoiskaya chromosome substitution lines possess increased chromosome stability compared with Chinese Spring/Imperial addition lines.

Chimera↗

Age and sex influence on oxidative damage and functional status in human skeletal muscle.

A reduction in muscle mass, with consequent decrease in strength and resistance, is commonly observed with advancing age. In this study we measured markers of oxidative damage to DNA, lipids and proteins, some antioxidant enzyme activities as well Ca2+ transport in sarcoplasmic reticulum membranes in muscle biopsies from vastus lateralis of young and elderly healthy subjects of both sexes in order to evaluate the presence of age- and sex-related differences. We found a significant increase in oxidation of DNA and lipids in the elderly group, more evident in males, and a reduction in catalase and glutathione transferase activities. The experiments on Ca2+ transport showed an abnormal functional response of aged muscle after exposure to caffeine, which increases the opening of Ca2+ channels, as well a reduced activity of the Ca2+ pump in elderly males. From these results we conclude that oxidative stress play an important role in muscle aging and that oxidative damage is much more evident in elderly males, suggesting a gender difference maybe related to hormonal factors.

Adolescent↗

Structural and functional organization of the '1S0.8 gene-rich region' in the Triticeae.

Wheat genes are present in physically small, gene-rich regions, interspersed by gene-poor blocks of retrotransposon-like repetitive sequences. One of the largest gene-rich regions is present around fraction length (FL) 0.8 of the short arm of wheat homoeologous group 1 chromosomes and is called '1S0.8 region'. The objective of this study was to reveal the structural and functional organization of the '1S0.8 region' in various Triticeae and other Poaceae species. Consensus genetic linkage maps of the '1S0.8 region' were constructed for wheat, barley, and rye by combining mapping information from 16, 11, and 12 genetic linkage maps, respectively. The consensus genetic linkage maps were compared with each other and with a consensus physical map of wheat homoeologous group 1. Comparative analyses localized 75 agronomically important genes to the '1S0.8 region'. This high-resolution comparison revealed exceptions to the rule of conserved gene synteny, established using low-resolution marker comparisons. Small rearrangements such as duplications, deletions, and inversions were observed among species. Proportion ofchromosomal recombination occurring in the '1S0.8 region' was very similar among species. Within the gene-rich region, the extent of recombination was highly variable but the pattern was similar among species. Relative recombination among markers was similar except for a few loci where drastic differences were observed among species. Chromosomal rearrangements did not always change the extent of recombination for the region. Differences in gene order and relative recombination were the least between wheat and barley, and were the highest between wheat and oat.

Chromosome Mapping↗

Randomized trial of a combination of natural products (cernitin, saw palmetto, B-sitosterol, vitamin E) on symptoms of benign prostatic hyperplasia (BPH).

Because benign prostatic hyperplasia (BPH) is relatively common, it is important to discover safe and effective means to treat this often debilitating perturbation. Accordingly, we examined the effectiveness of a combination of natural products (cernitin, saw palmetto, B-sitosterol, vitamin E) in treating symptoms of BPH. We undertook a randomized, placebo-controlled, double-blind study. Patients were enrolled from 3 urological practices in the USA. 144 subjects were randomized for study. 17 subjects eventually withdrew, leaving 70 patients in the test group and 57 in the placebo group to complete the study. Inclusion criteria consisted of a diagnosis of BPH, no evidence of cancer, and a maximal urinary flow rate between 5 and 15 ml/second. Patients received either placebo or the combined natural products for 3 months. Evaluations were performed via the American Urological Association (AUA) Symptom Index score, urinary flow rate, PSA measurement, and residual bladder volume. Nocturia showed a markedly significant decrease in severity in patients receiving the combined natural products compared to those taking placebo (p < 0.001). Daytime frequency was also lessened significantly (p < 0.04). When the average individual total AUA Symptom Index score in the test group was compared to that in the placebo group at the end of the study, the difference proved highly significant (p < 0.014). PSA measurements, maximal and average urinary flow rates, and residual volumes showed no statistically significant differences. When taken for 3 months, a combination of natural products (cernitin, saw palmetto, B-sitosterol, vitamin E) compared to placebo can significantly lessen nocturia and frequency and diminish overall symptomatology of BPH as indicated by an improvement in the total AUA Symptom Index score. The combination of natural products caused no significant adverse side effects.

Androgen Antagonists↗

Interphase arrangement of rye B chromosomes in rye and wheat.

Probes for B chromosome-specific sequences in the distal region of the long arm of the rye B have been used to investigate the interphase arrangement of the Bs in rye and in hexaploid wheat. The Lindström strain of wheat carries the rye Bs as additions. The number of in situ signals in nuclei with two, three and four Bs is often less than the maximum B number, and it seems that the Bs may be grouped together in various ways rather than being randomly dispersed throughout the nucleus. The degree of physical association is greater in rye than in the allen wheat background. The results are discussed in relation to the pairing and recombination preferences of the Bs in rye and in Lindström wheat.

Chromosomes↗

Development-dependent inheritance of 5-azacytidine-induced epimutations in triticale: analysis of rDNA expression patterns.

Genomic imprinting of rye origin rDNA sequences in triticale is modulated by DNA methylation responsible for ontogenic expression patterns of those sequences. Considering the dynamic nature of these phenomena, we evaluated the influence of plant development on the inheritance of modified rye rDNA expression patterns. DNA hypomethylation was induced in triticale by 5-azacytidine (5AC) treatments at distinct developmental stages of M1 plants, and expression patterns were analysed in M2. The activity of rye origin rRNA genes in progeny of untreated and 5AC-treated plants was evaluated by silver staining in meristematic root tip cells and in meiocytes at diplotene. In the progeny of 5AC-treated plants, a significant increase in rye rDNA expression was observed, contrasting with the residual activity in untreated plants. Significant differential effects of 5AC treatments were observed in M2 plants and correlated with the M1 plant developmental stage in which DNA hypomethylation was induced. Hypotheses to explain the origin of those differences are discussed here.

Antimetabolites, Antineoplastic↗

Nucleolar dominance in triticales: control by unlinked genes.

Hybrid plants and animals often show suppression of activity of ribosomal genes (rDNA) originating from one of the parental or ancestral species. In the wheat x rye amphiploid triticale, containing 28 chromosomes of wheat origin and 14 from rye, rDNA of rye origin (on chromosome 1R) is not normally expressed, while the 1B- and 6B-origin rDNA from wheat shows strong expression. Expression of rDNA can be accurately assessed by the silver staining method, which stains both interphase nucleoli and metaphase rDNA sites that were actively expressed at the previous interphase. We show here that substitution of another rye chromosome, 2R, by a chromosome from hexaploid wheat, 2D (triticale-2D(2R)), prevents suppression of the rye-origin rDNA, and leads to activity of all six major rDNA loci. These results were found in two different triticales and supported by rDNA behaviour in wheat-rye chromosomal addition lines. Models for chromosomal interactions leading to control of rDNA expression are presented.

Chimera↗

Alterations in the distribution of histone H3 phosphorylation in mitotic plant chromosomes in response to cold treatment and the protein phosphatase inhibitor cantharidin.

The function of the phosphorylation of histone H3 at Ser 10 in plant cell division is uncertain. The timing correlates with chromosome condensation, and studies in plant meiosis suggest that it is involved in sister chromatid cohesion. In mitosis, plant chromosomes are highly phosphorylated in the pericentromeric region only. In order to modulate H3 phosphorylation, root meristems of different plant species were treated with the protein phosphatase inhibitor cantharidin or with ice-water. Immunostaining using an antibody specific to phosphorylated H3 at Ser 10 revealed a high level of H3 phosphorylation along the whole mitotic chromosome after cantharidin treatment, which resembles the distribution seen exclusively in first meiotic division. In chromosomes that were isolated from meristems treated with ice-water, the heterochromatic regions and nucleolar organizer regions, in addition to the pericentromeric region, were highly phosphorylated at H3. Cantharidin and ice-water also affected spindle assembly and chromosome length, but these effects did not seem to be directly linked to changes in H3 phosphorylation.

Cantharidin↗

Chromosome arrangement and behaviour of two rye homologous telosomes at the onset of meiosis in disomic wheat-5RL addition lines with and without the Ph1 locus.

Fluorescence in-situ hybridization (FISH) of total genomic and repetitive DNA on microsporocytes of ditelocentric addition lines of rye 5RL in hexaploid wheat was performed to study the behaviour of the rye homologous chromosome arms in relation to centromere and telomere dynamics at premeiotic interphase and meiotic prophase I. By comparing isogenic lines with and without the Ph1 locus, we established the effect of the Ph1 gene on appearance and behaviour of the rye chromosomes. Ph1 and ph1b lines demonstrated similar premeiotic chromosome arrangement with the two rye homologues occupying separated domains despite the occurrence of centromere association. Our study confirmed that bouquet arrangement of telomeres follows the Rabl configuration. In cells displaying bouquet clustering of telomeres, centromeres of the 5RL telosomes are still at the opposite pole, suggesting anchoring of centromeres at the cytoskeleton. Once the telomeres complete clustering, the rye centromeres migrate to the telomere pole, and the rye chromosomes begin to loosen their structure. While the rye homologues in the wild-type keep separate territories in the nucleus, they become intermingled in the ph1b mutant, possibly because of their lower condensation. In a subsequent stage, the 5RL homologues appear intimately associated mainly at the distal region. Our study suggests that the lower rate of chromosome synapsis in the ph1b mutant results from abnormal chromatin decondensation and organization.

Cell Nucleus↗

Molecular cloning of alpha-amylases from cotton boll weevil, Anthonomus grandis and structural relations to plant inhibitors: an approach to insect resistance.

Anthonomus grandis, the cotton boll weevil, causes severe cotton crop losses in North and South America. Here we demonstrate the presence of starch in the cotton pollen grains and young ovules that are the main A. grandis food source. We further demonstrate the presence of alpha-amylase activity, an essential enzyme of carbohydrate metabolism for many crop pests, in A. grandis midgut. Two alpha-amylase cDNAs from A. grandis larvae were isolated using RT-PCR followed by 5' and 3' RACE techniques. These encode proteins with predicted molecular masses of 50.8 and 52.7kDa, respectively, which share 58% amino acid identity. Expression of both genes is induced upon feeding and concentrated in the midgut of adult insects. Several alpha-amylase inhibitors from plants were assayed against A. grandis alpha-amylases but, unexpectedly, only the BIII inhibitor from rye kernels proved highly effective, with inhibitors generally active against other insect amylases lacking effect. Structural modeling of Amylag1 and Amylag2 showed that different factors seem to be responsible for the lack of effect of 0.19 and alpha-AI1 inhibitors on A. grandis alpha-amylase activity. This work suggests that genetic engineering of cotton to express alpha-amylase inhibitors may offer a novel route to A. grandis resistance.

Amino Acid Sequence↗

Effect of green manure on the incidence of cyanogenic Pseudomonas strains in hop garden soils.

Incidence of cyanogenic Pseudomonas strains in hop garden soils in relation to the kind of fertilization was studied. Incidence differed with respect to the fertilization treatment and the age of the plantation. Amendment of soil with rye and with white mustard as green manures limited the number of cyanogenic Pseudomonas strains relative to farmyard manures and NPK fertilization. Among all fertilization treatments, cyanogenic Pseudomonas spp. strains had lowest populations in soils amended with white mustard.

Fertilizers↗

Comparison of Saw Palmetto (extract and whole berry) and Cernitin on prostate growth in rats.

Pharmaceuticals such as finasteride and alpha blockers are used to treat symptoms of benign prostatic hyperplasia (BPH) and are known to cause severe adverse reactions. Accordingly, a search for safer, natural products has been undertaken. Two natural agents (nutraceuticals) have come under recent scrutiny; because natural products, in general, often have evidence of long-term safety. The present study compares the in vivo effects on androgen-induced prostatic enlargement in rats of two nutraceuticals--the widely recognized Saw Palmetto (Serenoa repens) and the less well-known Cernitin (defined pollen extract). Non-castrated rats, had a mean prostate weight of 124 mg +/- 8.8 (S.E.M.) compared to the 24.5 mg +/- 1.9 (S.E.M.) of the castrated rat followed under the same regimen (p < 0.01). When castrated rats were given testosterone, the mass increased significantly to 250.0 mg +/- 31.7 (S.E.M.) (p < 0.01). In the five remaining groups, castrated rats receiving testosterone were given finasteride, an extract of Saw Palmetto, crushed whole berry derived from Saw Palmetto fruit, a water soluble and fat soluble extract of Cernitin or a combination of the Saw Palmetto extract and Cernitin. All treatments decreased the size of the prostate to roughly the same size as in the non-castrated rats, a size that was significantly smaller than castrated rats treated with testosterone in the same manner (p < 0.01). A second study examining non-castrated rats treated with very high doses of testosterone showed similar results. In both studies, the nutraceuticals generally decreased body weight. In conclusion, these studies show the ability of Saw Palmetto (whole berry and extract) and Cernitin to influence prostatic hyperplasia via effects on androgen metabolism.

Androgens↗

Meiosis in primary trisomics of rye: considerations for models of chromosome pairing.

Meiotic chromosome pairing of primary trisomics of rye was analysed by electron microscopy in surface-spread prophase I nuclei and compared with light-microscopic observations of metaphase I cells. Despite the large-sized chromosomes of rye, prophase I trivalent frequencies were close to the two thirds expected on a simple model with two terminal independent pairing initiation sites per trisome (set of three homologous chromosomes). Direct observations mostly reveal one pairing partner switch (PPS) in prophase I trivalents, which confirms this supposition. There were no significant differences between the number of trivalent and bivalent plus univalent configurations observed at prophase and metaphase I; therefore, synapsed segments form chiasmata. In all of the trisomics, the three homologues showed variations not only in the number of telomeric C-bands but also in the amount of heterochromatin of these bands, which allowed identification of chromosomes or chromosome arms associated in most metaphase I configurations. In trisomics for chromosomes 2, 3 and 5, some metaphase I chromosome configuration frequencies did not fit those expected under the assumption of random chromosome association among all partners, suggesting the existence of preferences for pairing between two given chromosome arms of the trisome. No preferential associations either at metaphase I or pachytene were observed in the trisomics for chromosome 6. The fit between theoretical pairing models and the experimental data is also discussed.

Cell Nucleus↗

Earlier onset of DNA fragmentation in leaves of wheat compared to barley and rye.

We have found extensive nucleosomal fragmentation of native DNA extracted from leaves of healthy cereal plants, as indicated by ladder patterns on agarose gels and TUNEL staining. The time of first appearance of fragmentation differed among cereals. Native DNA from the first leaf of 10-day-old plants formed a clear ladder pattern of multiples of 180 bp fragments in wheat and triticale but not in barley and oats. In one cultivar of rye a weak ladder pattern occurred but not in another. Freezing and thawing of samples before DNA extraction resulted in much more extensive DNA fragmentation in wheat but not in rye and barley, indicating that DNA-degrading enzymes are present in the cytoplasm of wheat, but not in barley and rye, at this stage. In barley, nucleosomal fragmentation was first detected in 25-day-old plants. These results indicate that programmed cell death takes place in developing leaves of young cereal plants, but that the time of onset differs among cereal species.

DNA Fragmentation↗