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Biomedical subjects

B Wells

Publications and source records attributed to B Wells.

At least 19 recordsLinked to original sources
PubMed2002

Coherence incoherence and dimensional crossover in layered strongly correlated metals.

The properties of an interacting electron system depend on the electron correlations and the effective dimensionality. For example, Coulomb repulsion between electrons may inhibit, or completely block, conduction by intersite electron hopping, thereby determining whether a material is a metal or an insulator. Furthermore, correlation effects increase as the number of effective dimensions decreases; in three-dimensional systems, the low-energy electronic states behave as quasiparticles, whereas in one-dimensional systems, even weak interactions break the quasiparticles into collective excitations. Dimensionality is particularly important for exotic low-dimensional materials where one- or two-dimensional building blocks are loosely connected into a three-dimensional whole. Here we examine two such layered metallic systems with angle-resolved photoemission spectroscopy and electronic transport measurements, and we find a crossover in the number of effective dimensions from two to three with decreasing temperature. This is apparent from the observation that, in the direction perpendicular to the layers, the materials have an insulating character at high temperatures but become metal-like at low temperatures, whereas transport within the layers remains metallic over the whole temperature range. We propose that this change in effective dimensionality correlates with the presence of coherent quasiparticles within the layers.

Journal Article↗
PubMed2001

Single ribosomal transcription units are linear, compacted Christmas trees in plant nucleoli.

The rDNA transcription units are enormous macromolecular structures located in the nucleolus and containing 50-100 RNA polymerases together with the nascent pre-rRNA attached to the rDNA. It has not previously been possible to visualize nucleolar transcription units directly in intact nucleoli, although highly spread preparations in the electron microscope have been imaged as "Christmas trees" 2-3 microm long. Here we determine the relative conformation of individual transcription units in Pisum sativum plant nucleoli using a novel labelling technique. Nascent transcripts were detected by a highly sensitive silver-enhanced 1 nm gold procedure, followed by 3D electron microscopy of entire nucleoli. Individual transcription units are seen as conical, elongated clusters approximately 300 nm in length and 130 nm in width at the thickest end. We further show that there were approximately 300 active ribosomal genes in the nucleoli examined. The underlying chromatin structure of the transcribing rDNA was directly visualized by applying a novel limited extraction procedure to fixed specimens in order to wash out the proteins and RNA, thus specifically revealing DNA strands after uranyl acetate staining. Using this technique, followed by post-embedding in situ hybridization, we observed that the nucleolar rDNA fibres are not extended but show a coiled, thread-like appearance. Our results show for the first time that native rDNA transcription units are linear, compacted Christmas trees.

Cell Nucleolus↗
PubMed2001

Cell wall architecture of the elongating maize coleoptile.

The primary walls of grasses are composed of cellulose microfibrils, glucuronoarabinoxylans (GAXs), and mixed-linkage beta-glucans, together with smaller amounts of xyloglucans, glucomannans, pectins, and a network of polyphenolic substances. Chemical imaging by Fourier transform infrared microspectroscopy revealed large differences in the distributions of many chemical species between different tissues of the maize (Zea mays) coleoptile. This was confirmed by chemical analyses of isolated outer epidermal tissues compared with mesophyll-enriched preparations. Glucomannans and esterified uronic acids were more abundant in the epidermis, whereas beta-glucans were more abundant in the mesophyll cells. The localization of beta-glucan was confirmed by immunocytochemistry in the electron microscope and quantitative biochemical assays. We used field emission scanning electron microscopy, infrared microspectroscopy, and biochemical characterization of sequentially extracted polymers to further characterize the cell wall architecture of the epidermis. Oxidation of the phenolic network followed by dilute NaOH extraction widened the pores of the wall substantially and permitted observation by scanning electron microscopy of up to six distinct microfibrillar lamellae. Sequential chemical extraction of specific polysaccharides together with enzymic digestion of beta-glucans allowed us to distinguish two distinct domains in the grass primary wall. First, a beta-glucan-enriched domain, coextensive with GAXs of low degrees of arabinosyl substitution and glucomannans, is tightly associated around microfibrils. Second, a GAX that is more highly substituted with arabinosyl residues and additional glucomannan provides an interstitial domain that interconnects the beta-glucan-coated microfibrils. Implications for current models that attempt to explain the biochemical and biophysical mechanism of wall loosening during cell growth are discussed.

Cell Division↗
PubMed2000

How many women lose bone mineral density while taking hormone replacement therapy? Results from the Postmenopausal Estrogen/Progestin Interventions Trial.

BACKGROUND: The frequency of bone loss among women using postmenopausal hormone therapy is unknown. METHODS: We used data from the Postmenopausal Estrogen/Progestin Interventions Trial to address the frequency of bone loss among women using postmenopausal hormone replacement therapy. Of 701 women randomized to active treatment (conjugated equine estrogens alone or in combination with 1 of 3 progestins), 538 (76.7%) were adherent and had replicate bone mineral density (BMD) measures at baseline, 12 months, and 36 months. Of 174 placebo-assigned women, 132 (75.9%) were similarly eligible. Replicate BMD measures were used to calculate within-person measurement errors, which were then used to delineate cut points that defined bone losers with 97.5%, 95.0%, 90.0%, or 75.0% confidence. RESULTS: At the lumbar spine, during the first 12 months, 1.5% of hormone users lost BMD with 97.5% confidence, corresponding to a decline of -3% per year; during months 12 to 36, only 0.6% of treated women lost spinal BMD to this degree. An annual loss of -1% or more was the criterion for spinal bone loss at the 75.0% confidence level; 5.1% and 8.0% of hormone users met this criterion in the first year and in months 12 to 36, respectively. For the total hip, during the first 12 months, 2.3% of hormone-adherent women lost -3.0% per year or more, the 97.5% confidence definition of loss; 0.4% were so classified during months 12 to 36. To be 75.0% confident of hip BMD loss, a -1.0% per year decline in BMD was required; using this criterion, 14.5% and 11.8% of hormone users lost total hip BMD between 0 to 12 and 12 to 36 months, respectively. Among hormone-adherent women, at the spine and hip, there was virtually no overlap between women classified as bone losers in the first 12 months and those classified as such in the last 24 months. With 95.0% certainty, corresponding to an approximate loss of -2.5% at the spine and hip, 31.3% and 11.7% of placebo-adherent women lost spinal BMD in the first 12 and last 24 months, respectively. Parallel figures for the hip were 32.3% and 7.9%, respectively. CONCLUSION: Bone loss while taking postmenopausal hormones is rare, and bone loss among untreated women is far from universal.

Bone Density↗
PubMed2000

Altering the inclusion properties of CTV through crystal engineering: CTV, carborane, and DMF supramolecular assemblies

The complexes [Na(CTV)2(OH)(H2O)](H2O)(DMF)2(o-carborane) (3; CTV = cyclotriveratrylene), [K(OH)(CTV)(DMF)]2(o-carborane) (4), [(DMF)(CTV)]2(H2O)4(o-carborane) (5), and (o-carborane)(CTV)(DMF)2 (6) all form as crystalline inclusion complexes from N,N'-dimethylformamide (DMF) solution. Complexes 3 and 4 are the first reported examples of CTV acting as a chelating ligand, with two CTV molecules coordinating cis to the six-coordinate M+ centers (M=Na, K). The extended structures of complexes 3-5 are similar, forming extended coordinate and/or hydrogen-bonding interactions and all feature intracavity complexation of DMF by CTV, while the complex 6 forms an assembly of (o-carborane) intersection of two sets (CTV) ball-and-socket supermolecules with DMF as a channel-type included guest.

Journal Article↗
PubMed2000

ATP-dependent regulation of nuclear Ca(2+) levels in plant cells.

Localised alterations in cytoplasmic Ca(2+) levels are an integral part of the response of eukaryotic cells to a plethora of external stimuli. Due to the large size of nuclear pores, it has generally been assumed that intranuclear Ca(2+) levels reflect the prevailing cytoplasmic Ca(2+) levels. Using nuclei prepared from carrot (Daucus carota L.) cells, we now show that Ca(2+) can be transported across nuclear membranes in an ATP-dependent manner and that over 95% of Ca(2+) is accumulated into a pool releasable by the Ca(2+) ionophore A.23187. ATP-dependent nuclear Ca(2+) uptake did not occur in the presence of ADP or ADPgammaS and was abolished by orthovanadate. Confocal microscopy of nuclei loaded with dextran-linked Indo-1 showed that the initial ATP-induced rise in [Ca(2+)] occurs in the nuclear periphery. The occurrence of ATP-dependent Ca(2+) uptake in plant nuclei suggests that alterations of intranuclear Ca(2+) levels may occur independently of cytoplasmic [Ca(2+)] changes.

Adenosine Triphosphate↗
PubMed2000

Nucleoside analogs plus ritonavir in stable antiretroviral therapy-experienced HIV-infected children: a randomized controlled trial. Pediatric AIDS Clinical Trials Group 338 Study Team.

CONTEXT: Although protease inhibitors are used routinely in adults with human immunodeficiency virus (HIV) infection, the role of these drugs in the treatment of clinically stable HIV-infected children is not clear. OBJECTIVE: To evaluate the safety, tolerance, and virologic response produced by a change in antiretroviral therapy in HIV-infected children who were clinically and immunologically stable while receiving previous therapy. DESIGN: The Pediatric AIDS Clinical Trials Group 338, a multicenter, phase 2, randomized, open-label controlled trial conducted from February 6 to April 30, 1997 (patient entry period); patients were followed up for 48 weeks. SETTING: Pediatric HIV research clinics in the United States and Puerto Rico. PATIENTS: Two hundred ninety-seven antiretroviral-experienced, protease inhibitor-naive, clinically stable HIV-infected children aged 2 to 17 years. INTERVENTIONS: Children were randomized to receive zidovudine, 160 mg/m2 3 times per day, plus lamivudine, 4 mg/kg 2 times per day (n = 100); the same regimen plus ritonavir, 350 mg/m2 2 times per day (n = 100); or ritonavir, 350 mg/m2 2 times per day, and stavudine, 4 mg/kg 2 times per day (n = 97). MAIN OUTCOME MEASURE: Plasma HIV-1 RNA levels at study weeks 12 and 48, compared among the 3 treatment groups. RESULTS: At study week 12, 12% of patients in the zidovudine-lamivudine group had undetectable plasma HIV RNA levels (<400 copies/mL) compared with 52% and 54% of patients in the 2- and 3-drug ritonavir-containing groups, respectively (P<.001). Through study week 48, 70% of children continued receiving their ritonavir-containing regimen. At study week 48, 42% of children receiving ritonavir plus 2 nucleosides compared with 27% of those receiving ritonavir and a single nucleoside had undetectable HIV RNA levels (P = .04); however, similar proportions in each group continuing initial therapy had HIV RNA levels of less than 10000 copies/mL (58% vs 48%, respectively; P = .19). CONCLUSIONS: In our study, change in antiretroviral therapy to a ritonavir-containing regimen was associated with superior virologic response at study week 12 compared with change to a dual nucleoside analog regimen. More children receiving ritonavir in combination with 2 compared with 1 nucleoside analog had undetectable HIV RNA levels at study week 48.

Adolescent↗
PubMed2000

A glucoamylase::GFP gene fusion to study protein secretion by individual hyphae of Aspergillus niger.

Although Aspergillus niger is used as a host for heterologous protein production, yields are generally lower than those obtained for homologous proteins. Mechanisms of protein secretion and the secretory pathway in filamentous fungi are poorly characterised, although there is evidence to suggest that secretion occurs by a mechanism similar to that in other eukaryotes, but with proteins destined for secretion being directed to the hyphal tip. We report on a method using a glucoamylase: GFP gene fusion which allows us for the first time to monitor, in vivo, protein secretion in A. niger at the single hyphal level. A synthetic green fluorescent protein (sGFP(S65T)) was fused to truncated A. niger glucoamylase (GLA:499). Southern blot analysis of transformants confirmed that the gene fusion had successfully integrated into the A. niger genome. Confocal and fluorescence microscopy revealed that the GLA::GFP fusion protein is fluorescent in A. niger and appears to be directed to the hyphal tip. In young mycelia, hyphal cell wall fluorescence is apparent and immunogold labelling of GFP confirmed that GFP was partially localised within the hyphal cell wall. Using Western blotting, extracellular GLA::GFP was detected only in culture filtrates of young mycelia grown in a soya milk medium. The actin inhibitor latrunculin B was used to disrupt the secretion process, and its effects on the distribution of GLA::GFP were monitored.

Actins↗
PubMed2000

The nucleus: a highly organized but dynamic structure.

The nucleus in plants and animals is a highly structured organelle containing several well-defined subregions or suborganelles. These include the nucleolus, interphase chromosome territories and coiled bodies. We have visualized transcription sites in plants at both light- and electron-microscopy level by the incorporation of BrUTP. In the nucleolus many dispersed foci are revealed within the dense fibrillar component, each of which probably corresponds to a single gene copy. In the nucleoplasm there are also many dispersed foci of transcription, but not enough to correspond to one site per transcribed gene. We have shown that in wheat, and probably many other plant species, interphase chromosome territories are organized in a very regular way, with all the chromosomes in the Rabl configuration, all the centromeres clustered at the nuclear membrane and all the telomeres located at the nuclear membrane on the opposite side of the nucleus. However, despite this regular, polarized structure, there is no sign of polarization of transcription sites, or of any preferred location for them with respect to chromosome territorial boundaries. The nucleus is also highly dynamic. As an example, we have shown by the use of a green fluorescent protein fusion to the spliceosomal protein U2B" that coiled bodies move and coalesce within the nucleus, and may act as transport structures within the nucleus and nucleolus.

Autoantigens↗
PubMed2000

Personal and situational variables that describe coping with acute stress in competitive sport.

The authors examined the degree to which competitive basketball players in Australia were consistent in their cognitive appraisals and coping strategies in response to 4 types of stressful situations that they had experienced during previous basketball games as functions of perceived stress intensity. The authors predicted that both approach and avoidance coping strategies would be dependent on the type of stressful event, in accord with the transactional model. The results supported that prediction: Approach strategies were more prevalent than avoidance strategies following 3 of the 4 events. Cognitive appraisals and perceived stress intensity also strongly influenced the participants' use of coping strategies, accounting for 34% of the variance.

Acute Disease↗
PubMed1999

The relationship of biochemical markers of bone turnover to bone density changes in postmenopausal women: results from the Postmenopausal Estrogen/Progestin Interventions (PEPI) trial.

We assessed the associations of eight bone turnover markers (BTMs) with baseline and 1-year percentage changes in lumbar spine and hip bone mineral density (BMD) of 293 postmenopausal women undergoing treatment with hormone replacement therapy (HRT) or placebo using squared correlation coefficients (R2). In 239 women assigned to treatment with estrogen alone or with with estrogen plus progestins (active treatment), mean percentage changes for all markers decreased significantly and remained below baseline values through 3 years of study, whereas mean percentage changes for 54 women assigned to the placebo group showed no significant change from baseline in any marker. At baseline, age and body mass index (BMI) together accounted for 16% and 25% of the variance in spine and hip BMD, respectively. The telopeptide resorption marker, cross-linked N-telopeptide of type I collagen (NTX), alone accounted for 12% and 8% of variance, respectively. Another telopeptide, carboxy-terminal telopeptide of type I collagen (Crosslaps), accounted for 8% and 7% of variance, respectively. A bone-specific alkaline phosphatase (BALP-2) accounted for 8% of variance at the spine and 5% at the hip. No other marker accounted for more than 5% of total variance at either site; adding either baseline NTX, Crosslaps, or BAP-2 to regressions containing age and BMI increased R2 values at the spine and hip to about 22% and 28%, respectively. In the placebo group, baseline spine BMD accounted for 4% of the variance in 1-year spine BMD percentage change, whereas baseline values for age and BMI accounted for 1% and 0% of the variance, respectively; none of the three accounted for more than 0% of hip BMD percentage change; Crosslaps and NTX contributed 5% and 4% to the variance in 1-year spine BMD percentage change, but other markers accounted for < 2% of variance at the spine. At the hip, another BALP (BALP-1) accounted for 4% of variance, but no other baseline marker except NTX accounted for more than 1% of variance. In the active treatment group, baseline values for age, BMI, and spine BMD together accounted for 13% of the percentage change in spine BMD and for 4% of the BMD change at the hip. No individual or pair of baseline markers significantly enhanced these R2 values, but addition of 1-year percentage changes in some individual markers did significantly increase it. The largest R2 value was obtained by adding the percentage change in BALP-2, which increased the R2 in spine BMD percentage change to 20% and that at the hip to 8%. Adding baseline and change variables for all eight markers to the regression increased R2 to 28% at the spine and 12% at the hip. Restricting the set of analyses to individuals who suppressed marker activity beyond the precision error for the measurement did not improve R2s for the regressions. When baseline marker values were stratified into quartiles, only NTX and osteocalcin showed significant relationships between quartile and change in spine BMD, and these did not reach significance at the hip. When the 1-year change in markers was stratified into quartiles, significant relationships with percentage change in spine BMD were observed only for BALP phosphatases. We conclude that BTMs are not a surrogate for BMD to identify women with low bone mass and that they offer little useful information for predicting BMD changes for individual untreated or HRT-treated postmenopausal women.

Alkaline Phosphatase↗
PubMed1998

The ROOT HAIRLESS 1 gene encodes a nuclear protein required for root hair initiation in Arabidopsis.

The epidermis of Arabidopsis wild-type primary roots, in which some cells grow hairs and others remain hairless in a position-dependent manner, has become an established model system to study cell differentiation. Here we present a molecular analysis of the RHL1 (ROOT HAIRLESS 1) gene that, if mutated, prevents the formation of hairs on primary roots and causes a seedling lethal phenotype. We have cloned the RHL1 gene by use of a T-DNA-tagged mutant and found that it encodes a protein that appears to be plant specific. The predicted RHL1 gene product is a small hydrophilic protein (38.9 kD) containing putative nuclear localization signals and shows no significant homology to any known amino acid sequence. We demonstrate that a 78-amino-acid sequence at its amino terminus is capable of directing an RHL1-GFP fusion protein to the nucleus. The RHL1 transcript is present throughout the wild-type plant and in suspension culture cells, but in very low amounts, suggesting a regulatory function for the RHL1 protein. Structural evidence suggests a role for the RHL1 gene product in the nucleolus. We have examined the genetic relationship between RHL1 and GL2, an inhibitor of root hair initiation in non-hair cells. Our molecular and genetic data with double mutants, together with the expression analysis of a GL2 promoter-GUS reporter gene construct, indicate that the RHL1 gene acts independently of GL2.

Amino Acid Sequence↗
PubMed1998

Feasibility of prescribing injectable heroin and methadone to opiate-dependent drug users: associated health gains and harm reductions.

OBJECTIVE: To assess the feasibility of offering the choice of prescribing injectable heroin (diamorphine) or injectable methadone to opiate-dependent injecting drug users and to assess whether there are health and social gains associated with prescribing injectable opiates. DESIGN: A protocol-driven prospective observational study. Type of injectable opiate received was based on self-selection. SETTING: A large west London drug clinic. PATIENTS: Fifty-eight patients admitted to the clinic between 1 June 1995 and 31 December 1996, who were long term opiate-dependent injecting drug users, who had previously tried and failed oral methadone and who were apparently unable or unwilling to give up injecting. MAIN OUTCOME MEASURES: Retention in treatment, illicit drug use, HIV risk behaviour, criminal activity, social functioning, health and psychological status as measured by self-report, urinalysis and doctor's ratings. RESULTS: Thirty-seven patients (64%) chose heroin and 21 (36%) chose injectable methadone. Fifty (86%) were retained in treatment after three months, 40 (69%) after six months and 33 (57%) after 12 months. Among those in treatment at three months, there were significant reductions in illicit drug use, illicit drug-injecting risk behaviour, and criminal activity, and significant improvements in social functioning, health status and psychological adjustment. Generally, these gains were sustained between three, six and 12 months. Doctors' ratings of health and urinalysis results further supported these findings. CONCLUSIONS: Injectable heroin is not always the drug of choice. This intervention retained most patients in treatment with substantial benefits to both patients and the community. Prescribing injectable opiates to long term injecting drug users is a feasible treatment option.

Adult↗
PubMed1998

Children's comprehension of unfamiliar regional accents: a preliminary investigation.

The effect of regional accent on children's processing of speech is a theoretically and practically important aspect of phonological development that has been little researched. 48 children from London, aged four and seven years old, were tested on their ability to repeat and define single words presented in their own accent and in a Glaswegian accent. Results showed that word comprehension was significantly reduced in the Glaswegian condition and that four-year-olds performed less successfully than seven-year-olds. Both groups made similar numbers of lexical misidentifications, but the younger children were more likely to fail to access any word at all. On the repetition task, the younger children showed a different pattern of errors to the older children, their productions being apparently more influenced by the phonetics of the Glaswegian stimuli. It is suggested that such phonetic responses are related to the younger children's failure to map the unfamiliar accent onto their own phonological representations. It is proposed that the lexical misidentifications, common to both age groups, are more likely to be induced by lack of context. The paper concludes with discussion of implications of these findings for our understanding of how children develop the ability to process unfamiliar regional accents.

Child↗
PubMed1998

Ribavirin and interferon alfa-2b in chronic hepatitis C: assessment of possible pharmacokinetic and pharmacodynamic interactions.

AIMS: The primary objective of this study was to determine whether pharmacokinetic interactions occurred between interferon alpha-2b (IFN) and ribavirin in patients with chronic hepatitis C infections. Additionally this study assessed the single and multiple-dose pharmacokinetics of ribavirin and IFN, and compared the safety, tolerability and antiviral pharmacodynamics of IFN plus ribavirin compared with either drug alone. METHODS: In this open label parallel group study, patients with chronic hepatitis C were randomized to receive IFN 3 million IU thrice weekly s.c. alone, ribavirin 600 mg twice daily p.o. alone or both drugs in combination over 6 weeks. Single and multiple dose pharmacokinetics and indices of antiviral pharmacodynamics were assessed during weeks 1 and 6, along with safety assessments during the study. RESULTS: The range of mean ribavirin terminal phase half-lives after single doses was 44-49 h. Comparison of week 1 and week 6 AUC(0,12h) values showed accumulation in plasma of approximately 6-fold. The range of mean washout half-lives after week 6 was 274-298 h, reflecting release of ribavirin from deep compartment stores. The range of single and multiple dose IFN terminal phase half-lives was 5-7 h. IFN demonstrated an increase in bioavailability (approximately 2-fold) upon multiple dose administration. Ribavirin and IFN pharmacokinetic parameters for combined ribavirin and IFN were similar to those during monotherapy with either compound, although the power of this study to detect differences was low. Serum HCV-RNA titers and ALT concentrations were reduced by IFN alone, ribavirin alone reduced ALT concentrations only, and combined IFN plus ribavirin produced numerically greater falls in both measurements than either treatment alone. Serum concentrations of neopterin and activity of 2',5'-oligoadenylate synthetase (2'5'-OAS) were increased by IFN alone and in combination with ribavirin, whereas serum 2'5'-OAS activity was decreased and neopterin concentrations unaltered by ribavirin monotherapy. IFN and ribavirin monotherapy produced characteristic changes in safety laboratory tests (IFN--reductions in white cells, neutrophils and platelets; ribavirin--reduced haemoglobin) and characteristic adverse event profiles (IFN--headache, flu-like symptoms, fatigue, anorexia, nausea, myalgia, and insomnia; ribavirin--headache, fatigue, myalgia, and pruritus). There was no additive effect of combination therapy on safety laboratory tests or reported adverse events. All changes were fully reversible upon treatment cessation. CONCLUSIONS: There was no evidence of pharmacokinetic interactions between IFN and ribavirin in this study. There were numerical trends indicating that the combination of IFN and ribavirin reduced titers of HCV-RNA to a greater extent than did either treatment alone, and the safety profile of combination therapy was similar to those of both monotherapy treatments.

Adolescent↗
PubMed1998

Plant-adapted green fluorescent protein is a versatile vital reporter for gene expression, protein localization and mitosis in the filamentous fungus, Aspergillus nidulans.

Green fluorescent protein (GFP) is a useful reporter to follow the in vivo behaviour of proteins, but the wild-type gfp gene does not function in many organisms, including many plants and filamentous fungi. We show that codon-modified forms of gfp, produced for use in plants, function effectively in Aspergillus nidulans both as gene expression reporters and as vital reporters for protein location. To demonstrate the use of these modified gfps as reporter genes we have used fluorescence to follow ethanol-induced GFP expression from the alcA promoter. Translational fusions with the modified gfp were used to follow protein location in living cells; plant ER-retention signals targeted GFP to the endoplasmic reticulum, whereas fusion to the GAL4 DNA-binding domain targeted it to the nucleus. Nuclear-targeted GFP allowed real-time observation of nuclear movement and division. These modified gfp genes should provide useful markers to follow gene expression, organelle behaviour and protein trafficking in real time.

Aspergillus nidulans↗
PubMed1997

Sites of rDNA transcription are widely dispersed through the nucleolus in Pisum sativum and can comprise single genes.

Incorporation by RNA polymerases of BrUTP into both plant root tissue and isolated plant nuclei as a method for localization of the sites of transcription has been used. In this paper pea root tissue was used, and under the conditions employed, nearly all the incorporation occurs in the nucleolus, and thus must be catalysed by RNA polymerase I. Immunofluorescence and confocal microscopy shows that incorporation occurs in a pattern consisting of many small foci distributed widely through the dense fibrillar component of the nucleoli. Immunogold labelling using silver-enhanced Nanogold probe at the electron microscopic level confirms the sites of transcription as small foci approximately 200 nm in diameter. Simultaneous fluorescence in situ hybridization with a probe to the external transcribed spacer (ETS) region of the pre-rRNA shows that the structures revealed by this probe and the BrUTP immunofluorescence labelling are very similar. A probe to the transcribed portion of the rDNA (18S) also shows a good correlation to the sites of BrUTP incorporation within the nucleolus. On the other hand a probe to the non-transcribed intergenic spacer region (NTS) shows very little coincidence with the sites of BrUTP incorporation, and double fluorescence in situ labelling with both 18S and NTS probes confirms this difference in localization. These results suggest that most BrUTP foci correspond to single transcribed genes.

Cell Nucleolus↗