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PubMed2004

Nucleolar localization of the human telomeric repeat binding factor 2 (TRF2).

The telomeric repeat binding factor 2 (TRF2) specifically recognizes TTAGGG tandem repeats at chromosomal ends. Unexpectedly immunofluorescence studies revealed a prominent nucleolar localization of TRF2 in human cells, which appeared as discrete dots with sizes similar to those present in the nucleoplasm. The TRF2 dots did not overlap with dots stemming from the upstream binding factor (UBF) or the B23 protein. After treatment with a low concentration of actinomycin D (0.05 microg/ml), TRF2 remained in the nucleolus, although this condition selectively inhibited RNA polymerase I and led to a relocalization of UBF and B23. TRF2 was prominent in the nucleolus at G0 and S but seemed to diffuse out of the nucleolus in G2 phase. During mitosis TRF2 dispersed from the condensed chromosomes and returned to the nucleolus at cytokinesis. Treatment with low doses of actinomycin D delayed the release of TRF2 from the nucleolus as cells progressed from G2 phase into mitosis. With actinomycin D present TRF2 was detected in discrete foci adjacent to UBF in prophase, while in metaphase a complete overlap between TRF2 and UBF was observed. TRF2 was present in DNase-insensitive complexes of nucleolar extracts, whereas DNA degradation disrupted the protein-DNA complexes consisting of Ku antigen and B23. Following treatment with actinomycin D some of the mitotic cells displayed chromosome end-to-end fusions. This could be correlated to the actinomycin D-suppressed relocalization of TRF2 from the nucleolus to the telomeres during mitosis. These results support the view that the nucleolus may sequester TRF2 and thereby influences its telomeric functions.

Animals↗
PubMed1996

The effects of xylazine on intrauterine pressure, uterine blood flow, maternal and fetal cardiovascular and pulmonary function in pregnant goats.

To investigate the effects of xylazine on pregnant goats, xylazine induced changes in heart rate (HR), arterial blood pressure (ABP) and arterial blood pH and gases in the mother and fetus, as well as changes in intrauterine pressure (IUP) and uterine blood flow (UBF), were studied using a chronic preparation model of pregnant goats. Intramuscular administration of xylazine (0.2 mg/kg b.w.) caused a marked decrease in HR and ABP of the mother, which remained significantly reduced up to 120 min. In the fetus, a significant decrease in HR and a significant increase in ABP were recorded after only 5 min. Significant decreases of maternal arterial blood pH and oxygen partial pressure (Po2) were observed up to 120 and 30 min after the injection, respectively. Fetal arterial blood pH and Po2 also decreased, but these changes were milder and less persistent than those of the mother. Within 2 to 5 min after the administration of xylazine, IUP began to increase and remained high for about 15 min. Thereafter, there was a frequent periodic increase in IUP. After 5 min, UBF decreased significantly and remained low up to 120 min. A fall in UBF was associated with a rise in IUP. These findings suggested that administration of xylazine to pregnant goats results in a decrease in UBF arising from the induction of uterine contractions, a decrease in circulating blood volume, hypoxemia and acidosis in the mother.

Animals↗
PubMed1997

The effects of medetomidine on maternal and fetal cardiovascular and pulmonary function, intrauterine pressure and uterine blood flow in pregnant goats.

To investigate the effects of medetomidine on late pregnant goats, medetomidine induced changes in maternal or fetal circulation and acid-base balance, as well as changes in intrauterine pressure (IUP) and uterine blood flow (UBF), were studied. Intramuscular administration of medetomidine (40 micrograms/kg b.w.) decreased the heart rate (HR) and arterial blood pressure (ABP) of the mother, and the change in HR was significant statistically (p < 0.05). In the fetus, HR and ABP showed a transient decrease and increase (p < 0.05), respectively. A decrease in maternal arterial blood pH and oxygen partial pressure (PO2) and an increase in carbon dioxide partial pressure (PCO2) were recorded after the injection, but none was significant. In the fetus, arterial blood PO2 decreased significantly (p < 0.05) after 5 min of administration, and a significant metabolic acidemia supported by a decrease in base excess was observed. Within 1 to 4 min after the administration of medetomidine, IUP began to rise and remained high for 10 to 14 min. Thereafter, the rise in IUP was frequent and periodical. After the injection, UBF significantly (p < 0.05) decreased, and the fall in UBF was associated with a rise in IUP. The maternal and fetal serum medetomidine concentration increased remarkably after the injection of medetomidine into the mother. These observations in late pregnant goats suggested that medetomidine induced a decrease in maternal cardiac output, a decrease in UBF arising from the induction of uterine contractions, and transplacental medetomidine can have a suppressive effect on the fetus.

Adrenergic alpha-Agonists↗
PubMed1991

Transcription complex formation at the mouse rDNA promoter involves the stepwise association of four transcription factors and RNA polymerase I.

We have used purified transcription factors and RNA polymerase I (pol I) to analyze the individual steps involved in the formation of transcription initiation complexes at the mouse ribosomal gene promoter in vitro. Complete assembly of transcription complexes requires pol I and at least four auxiliary factors, termed TIF-IA, TIF-IB, TIF-IC, and UBF. Preincubation and template commitment, as well as order of addition protocols, were used to discriminate between various intermediate complexes generated during assembly of the initiation complex. As a first step, TIF-IB binds to the core promoter, a process that is facilitated by the upstream control element and the upstream binding factor (UBF). Binding of TIF-IB to the rDNA promoter results in the formation of a functional preinitiation complex (complex 1), which is stable for many rounds of transcription. UBF, which on its own does not stably associate with the rDNA promoter, triggers a 5-10-fold increase in the overall amount of this primary complex. Following binding of TIF-IB and UBF to the template DNA, pol I and TIF-IC successively bind, yielding complexes 2 and 3, respectively. Transcription-competent initiation complexes are built up by the final association of the growth-regulated factor TIF-IA. The various complexes can be distinguished by their different sensitivity to Sarkosyl. Only the complete complex consisting of all four factors and pol I shows resistance to intermediate concentrations of Sarkosyl (0.045%) and is competent to catalyze the formation of the first phosphodiester bond. The initiated complex is, on the other hand, resistant to high concentrations of Sarkosyl (0.3%). The hierarchical nature of the different complexes formed suggests a model for transcription initiation and predicts functions for the individual factors.

Animals↗
PubMed1989

The sufficiency of blood flow in human masseter muscle during endurance of biting in the intercuspal position and on a force transducer.

Blood flow changes caused by endurance of voluntary isometric muscle contraction and the sufficiency of flow during contraction were assessed by measuring 133Xe clearance. Aiming at 50% maximum voluntary contraction (MVC), 9 healthy persons sustained biting in the intercuspal position (ICP group) at an actual medium EMG activity level of 55% MVC, while receiving a visual feedback of the average-integrated EMG activity. Eleven persons sustained unilateral biting (UBF group) on a force transducer at 40% MVC, receiving feedback of the force output. The significantly lower % MVC for the UBF group was due in part to a decrease in the EMG activity during endurance, while force was constant. Blood flow changes over time were significant; however, the changes differed significantly between groups during endurance: the ICP group had a median reduction in blood flow to 0.4 of the initial resting value, and the UBF group had a 0.2-fold median increase. Following endurance, flow changes integrated over 3 minutes were about 43-fold the initial resting values of both groups. This similarity was probably a result of their equivalent effort. The endurance flow accounted for 1% of the total change for the ICP group and 5% for the UBF group. Overall, the proportion of the total flow that took place during endurance decreased logarithmically with greater level of contraction and masseter effort. Therefore, sufficiency of blood flow to maintain muscle fibre homeostasis is less when the rate of metabolic turnover is greater, thus contributing to an earlier onset of masseter fatigue, pain and exhaustion at high contraction levels.

Adult↗
PubMed1983

[Effects of indomethacin on renal and uterine blood flow and pressor responsiveness to angiotensin II or norepinephrine in pregnant dogs].

Pressor responsiveness to angiotensin II (A II) or norepinephrine (NE) in anesthetized nonpregnant and pregnant dogs, and the changes in renal blood flow (RBF) and uterine blood flow (UBF) elicited by the pressure rise in pregnant dogs were studied. Intervention of prostaglandins (PGs) in these hemodynamic responses was also examined, comparing the responses before and after the administration of indomethacin (IDM). The pressor responsiveness to the pressor substances was diminished in near term pregnant dogs in common with normal human pregnancy, and the refractoriness disappeared after the administration of IDM. RBF decreased following the infusion of A II or NE, and IDM enhanced the responses. A characteristic increase in UBF was observed following the infusion of A II exhibiting the maximum response at a dose of 0.05 microgram/kg, while it decreased following the infusion of NE, and these responses remained almost unchanged after IDM administration, although there was a 24.2% decrease in the base line value of UBF. It seems appropriate to conclude that PGs play a pivotal role in the development of the relative vascular refractoriness to infused pressor substances characterizing normal pregnancy, and also in preservation of RBF and UBF during pregnancy. Moreover, PGs attenuate the response of the renal vascular bed as well as systemic vascular bed to the pressor substances. However, the refractoriness of uterine vascular bed to A II remained almost unchanged regardless of PGs synthesis, suggesting the existence of another competitive factor involved in the effect of A II.

Angiotensin II↗
PubMed2002

Nucleolar protein allocation and ultrastructure in bovine embryos produced by nuclear transfer from granulosa cells.

In the present study immunofluorescence confocal laser scanning microscopy, autoradiography following (3)H-uridine incubation and transmission electron microscopy were used to evaluate the nucleolar protein localization, transcriptional activity, and nucleolar ultrastructure during genomic re-programming in bovine embryos reconstructed by nuclear transfer from granulosa cells into non-activated cytoplasts followed by activation. During the 1st cell cycle (1-cell embryos), no autoradiographic labelling was detected. Ultrastructurally, nucleoli devoid of a granular component were observed. During the 2nd cell cycle (2-cell embryos) autoradiographic labelling was also lacking and the embryos displayed varying degrees of nucleolar inactivation. During both the 3rd (4-cell embryos) and 4th (tentative 8-cell embryos), cell cycles autoradiographic labelling was lacking in some embryos, while others displayed labelling and associated formation of fibrillo-granular nucleoli. During the 5th cell cycle (tentative 16-cell embryos), all embryos displayed autoradiographic labelling and fibrillo-granular nucleoli. In some blastomeres, however, deviant nucleolar ultrastructure was observed. During the first cell cycle labelling of RNA polymerase I, fibrillarin, upstream binding factor (UBF) and nucleolin (C23) was localized to nuclear entities. During the 2nd cell cycle, only labelling of RNA polymerase I and fibrillarin persisted. During the 3rd and 4th cell cycle labelling of fibrillarin persisted, labelling of nucleophosmin (B23) appeared and that of nucleolin re-appeared. During the 5th cell cycle almost all embryos showed complete labelling of all proteins except for UBF, which lacked in more than half of the embryos. In conclusion, bovine granulosa cell nuclear transfer embryos showed re-modelling of the nucleoli to an inactive form followed by re-formation of fibrillo-granular nucleoli. The re-formation of fibrillo-granular nucleoli was initiated already during the 3rd cell cycle, which is one cell cycle earlier than in in vivo- and in vitro-derived bovine embryos. Moreover, in more than half of the embryos, UBF could not be immunocytochemically localized to the nucleolar compartment during the 5th cell cycle indicating lack of developmental potentials.

Animals↗
PubMed1997

Regulation of ribosomal RNA gene transcription during retinoic acid-induced differentiation of mouse teratocarcinoma cells.

We have examined the mechanism of regulation of rRNA synthesis in mouse F9 teratocarcinoma cells that were induced to differentiate by retinoic acid and dibutyryl cAMP. Ribosomal RNA (rRNA) synthesis was significantly reduced during differentiation of F9 cells into parietal endoderm cells. Nuclear run-on assay revealed that the rRNA gene transcription rates were reduced in differentiated cells, and this phenomenon could be mimicked by in vitro transcription assay using nuclear extracts prepared from F9 stem and F9 parietal endoderm cells. Analysis of the DNA-binding activities of two RNA polymerase I (pol I) transcription factors E1BF/Ku and UBF revealed decreased affinity for their cognate recognition sequences. Immunoblot analysis showed a marked reduction in the amounts of E1BF/Ku and UBF in the differentiated cells. Analysis of the steady-state RNA levels for the smaller subunit of E1BF/Ku and for UBF in differentiating F9 cells revealed decreased mRNA synthesis and increase in message level for the differentiation-specific marker laminin B1 with progression of the differentiated status of the cells. This study has demonstrated that differentiation of mouse F9 teratocarcinoma cells into parietal endoderm cells leads to diminished rRNA synthesis, which may be mediated by reduced DNA-binding activities and amounts of at least two pol I transcription factors.

Animals↗
PubMed1999

Simultaneous localization of transcription and early processing markers allows dissection of functional domains in the plant cell nucleolus.

Nucleolar transcription in isolated onion cell nuclei was visualized, after Br-UTP incorporation, under the conventional fluorescence microscope, the confocal microscope, and the transmission electron microscope. The confocal microscopy study of transcription was combined with immunodetection of fibrillarin, a component of the RNP complex involved in the early processing of pre-rRNA. Superposition of transcription and fibrillarin images from the same optical section showed some small "black holes" in the nucleolus, around which a lateral and radial differentiation of labeling was observed: laterally, zones corresponding to transcription labeling alternated with zones of fibrillarin labeling; radially, areas of transcription gradually became areas of colocalization of transcription and fibrillarin, and, further outward, of fibrillarin alone, which occupied the major part of the labeled nucleolar area. Three-dimensional reconstruction of the nucleolar transcription labeling, from confocal optical sections, showed clusters of foci arranged around an area of low or no labeling. Thin labeled extensions, connecting single foci, were observed. Visualization of transcription at the ultrastructural level identified the black holes as fibrillar centers, in view of their size and the absence of labeling in them. In fact, most of the labeling was observed in discrete areas of the dense fibrillar component, near fibrillar centers, including the transition area between these two components. This observation was supported by a quantitative study. Otherwise, the outline of fibrillar centers did not appear entirely surrounded by particles, and a minor proportion of particles was detected dispersed throughout the dense fibrillar component. As a complementary study, the transcription factor upstream binding factor (UBF) and the protein NopA64, a plant nucleolin homologue, were immunolocalized. Small foci of UBF localization alone and other foci in which the two protein markers overlapped were observed. The outer areas of the nucleolus showed the exclusive presence of NopA64. Under the electron microscope, UBF labeling, quantitatively assessed, appeared as clusters of particles, most of them surrounding fibrillar centers. A graphic model is presented to give a molecular interpretation of these data.

Bromodeoxyuridine↗
PubMed1995

Assessment of the fetal PO2 changes by cerebral and umbilical Doppler on lamb fetuses during acute hypoxia.

The objective of the present study was to validate one or a combination of fetal Doppler parameters in order to assess acute fetal hypoxia in an ovine model. Acute hypoxia was induced by reducing umbilical, or maternal aortic flow (approx. 70%). A CW Doppler probe was fixed on the fetal cervical skin, facing the internal carotid artery and the fetal abdominal skin adjacent to the umbilical arteries. (The angle between Doppler beam and flow vector remained constant.) A "Doptek 3000" spectrum analyser was used to measure the maximal and mean Doppler frequencies. Heart rate (HR), umbilical blood flow (UBF), carotid blood flow (CBF), umbilical RI (URI), cerebral RI (CRI) and cerebroplacental ratio (CPR = CRI/URI) were calculated in real time. A catheter was inserted into the fetal femoral artery, for blood gas (PO2, PCO2 pH) and blood pressure (BP) measurements. After 1 min of aorta compression (70% aortic flow reduction), the URI increased by 10% (P < 0.05), and the UBF decreased by 10% (P < 0.05), but the CRI decreased by 20% (P < 0.02), and the CBF did not change significantly. Fetal PO2 and CPR fell down after 1 min (59% and 38%, respectively; P < 0.001), although strong fetal heart rate decelerations were observed. The blood pressure, PCO2 and pH did not change significantly during this test. Throughout the 12 min of cord compression (70% umbilical flow reduction) the URI increased (70% to 80% P < 0.001), and the UBF decreased (approx. 60%; P < 0.001), but the CRI decreased (approx. 25%; P < 0.01), and the CBF remained constant (+/- 5%; ns). Fetal PO2 and CPR all decreased during the compression (30% to 44% and 40% to 60%, respectively; P < 0.001). HR, pH and PCO2 did not change significantly. During cord compression the blood pressure did not change significantly. In both cases, the CPR decreased significantly (P < 0.001) with the PO2 in the same direction and with a comparable amplitude (-30% to -50%). Nevertheless, the drop in CPR was greater during cord compression than during aorta compression, probably because the compression of the cord induced a central hypovolemia in addition to the hypoxia. The CPR was found to be the hemodynamic parameter that followed most closely the PO2 acute changes. The amplitude of the variations of this parameter (-30% to -50%) were quite similar to those of the PO2 during the period of acute hypoxia.(ABSTRACT TRUNCATED AT 400 WORDS)

Acute Disease↗
PubMedDate not supplied

Ethanol increases uterine blood flow and fetal arterial blood oxygen tension in the near-term pregnant ewe.

Fetal hypoxia is a hypothesized mechanism of ethanol teratogenesis. The objective of this study was to test this hypothesis by determining the effects of maternal ethanol infusion on uterine blood flow (UBF) and fetal oxygen status. UBF was measured with an electromagnetic flow probe placed around the left maternal uterine artery of the surgically recovered instrumented near-term pregnant sheep at 124 +/- 3 days of gestation (term = 147 days). Experimental treatment involved maternal infusion of 2 g (n = 3) or 4 g (n = 5) ethanol/kg maternal body weight, or 0.9% saline (n = 4) over a 5-h period. Arterial blood samples were collected at regular intervals to monitor maternal ethanol concentration and fetal PO2. Maternal ethanol infusion produced a dose-dependent increase (p = 0.0009) in UBF. Ethanol infusion also increased (p = 0.03) fetal arterial PO2. Overall, these findings indicate that fetal hypoxia is not a mechanism of ethanol teratogenesis.

Abnormalities, Drug-Induced↗
PubMed2005

Fetal hypertension and abnormal Doppler velocimetry in an ovine model of intrauterine growth restriction.

OBJECTIVE: Our objective was to test the hypothesis that systemic blood pressure (BP) is increased above normal in intrauterine growth restricted (IUGR) fetal lambs with elevated umbilical artery (UmA) Doppler indices. STUDY DESIGN: Five pregnant ewes were exposed to hyperthermic conditions for 80 days beginning at 40 days' gestation (dGA) to induce IUGR. They were then placed in ambient conditions with 6 additional ewes that served as controls. Doppler indices were calculated from UmA Doppler flow velocity waveforms. At 128 dGA, fetal catheters were placed for measurement of umbilical blood flow (UBF) by an ethyl alcohol steady-state diffusion technique and for aortic BP measurements. At 132 dGA, fetal mean systemic BP and blood flow were determined. At necropsy the placental and fetal weights were recorded. UBF was normalized for fetal weight. Linear regression, F tests and t tests were performed as appropriate. P < .05 was considered significant. RESULTS: Compared with control pregnancies, the IUGR pregnancies showed: (1) reduced fetal and placental weights, (2) elevated systemic BP, (3) reduced UBF, (4) elevated UmA and aortic Doppler velocimetry indices, (5) increased resistance per 100 g placenta, and (6) decreased UmA oxygenation and increased lactic academia. The UmA Doppler index of resistance (systolic/diastolic ratio) correlated strongly with calculated resistance (R2 = 0.7). Doppler indices also correlated with systemic BP (R2 = 0.5). CONCLUSION: Ovine IUGR fetuses with high UmA Doppler indices have elevated systemic BPs. UmA Doppler indices of resistance correlate well with (1) fetal systemic BPs and (2) resistance as calculated by pressure/flow. This whole animal study shows that IUGR fetuses are hypertensive and that increased UmA Doppler resistance indices are consistent with a fetal-placental hypertensive state.

Animals↗
PubMed1998

The first high-mobility-group box of upstream binding factor assembles across-over DNA junction by basic residues.

Upstream binding factor (UBF) is a eukaryotic RNA polymerase I-specific transcription factor. Its predominant DNA-binding motif, ubfHMG box 1, preserves DNA assembling activity that can bind two or more DNA duplexes simultaneously to form a crossover DNA junction. Here we investigate the basis of crossover DNA-assembling activity of ubfHMG box 1 by extensive mutagenesis analyses and mobility shift assay. Although the ubfHMG box 1 preserves a high mobility group (HMG) core structure, changing a number of the consensus hydrophobic and aromatic residues to alanine did not inhibit its crossover-assembling activity. This indicates that these residues do not directly participate in protein-DNA interaction. However, altering a series of basic residues in the helices 1 and 2 regions or the N-terminal extended strand of the ubfHMG box 1 motif had severe effects on DNA-assembling activity; however, certain non-specific DNA binding activity still remained. This suggests that the ubfHMG box 1 motif might extensively contact the backbone of a crossover junction through its multiple basic residues. Mutating a hydrophobic residue in the terminal dimerization domain inhibited the association of truncated Xenopus UBF, but had little effect on its crossover-assembling activity. This indicates that the UBF-crossover DNA complex is not established by the association of individual DNA-bound peptides.

Amino Acid Sequence↗
PubMed2005

Nucleolar biogenesis: the first small steps.

The nucleolus is the site of rRNA transcription, pre-rRNA processing and ribosome subunit assembly. The nucleolus assembles around clusters of ribosomal gene repeats during late telophase, persists throughout interphase and then disassembles as cells enter mitosis. The initial step in nucleolar formation is ribosomal gene transcription, which is mediated by Pol I (RNA polymerase I) and its associated transcription factors: UBF (upstream-binding factor), SL1 (selectivity factor) and TIF-IA (transcription initiation factor IA)/Rrn3. Ribosomal gene clusters, termed NORs (nucleolar organizer regions), are found on each of the five human acrocentric chromosomes. Though transcription is repressed during metaphase, NORs that were active in the previous interphase form prominent cytogenetic features, namely secondary constrictions. The main defining characteristic of these constrictions is under-condensation in comparison with the rest of the chromosome. Extensive binding of UBF over the ribosomal gene repeat is responsible for the formation of this chromosomal feature. During interphase, the majority of the Pol I transcription machinery, though present in nucleoli, is not actively engaged in transcription. Interaction with UBF bound across the gene repeat provides an explanation for how this non-engaged Pol I machinery is sequestered by nucleoli.

Cell Nucleolus↗
PubMed2004

The Treacher Collins syndrome (TCOF1) gene product is involved in ribosomal DNA gene transcription by interacting with upstream binding factor.

Treacher Collins syndrome (TCS) is an autosomal dominant disorder characterized by an abnormality of craniofacial development that arises during early embryogenesis. TCS is caused by mutations in the gene TCOF1, which encodes the nucleolar phosphoprotein treacle. Even though the genetic alterations causing TCS have been uncovered, the mechanism underlying its pathogenesis and the function of treacle remain unknown. Here, we show that treacle is involved in ribosomal DNA gene transcription by interacting with upstream binding factor (UBF). Immunofluorescence labeling shows treacle and UBF colocalize to specific nucleolar organizer regions and cosegregate within nucleolar caps of actinomycin d-treated HeLa cells. Biochemical analysis shows the association of treacle and UBF with chromatin. Immunoprecipitation and the yeast two-hybrid system both suggest physical interaction of the two nucleolar phosphoproteins. Down-regulation of treacle expression using specific short interfering RNA results in inhibition of ribosomal DNA transcription and cell growth. A similar correlation is observed in Tcof(+/-) mouse embryos that exhibit craniofacial defects and growth retardation. Thus, treacle haploinsufficiency in TCS patients might result in abnormal development caused by inadequate ribosomal RNA production in the prefusion neural folds during the early stages of embryogenesis. The elucidation of a physiological function of treacle provides important information of relevance to the molecular dissection of the biochemical pathology of TCS.

Animals↗
PubMed1991

The RNA polymerase I transcription factor xUBF contains 5 tandemly repeated HMG homology boxes.

The RNA polymerase I transcription factor UBF has been identified in human, mouse, rat and Xenopus and the primary structure of the human protein has been determined. Human UBF was shown to contain four tandem homologies to the folding domains of the HMG1 and 2 proteins and hence to belong to a previously unrecognised family of 'HMG-box' transcription factors. Here, cDNA clones encoding the Xenopus laevis UBF (xUBF) have been isolated and sequenced. Northern and Southern blots revealed that in tissue culture cells, xUBF is coded on a single major mRNA size species by a small number of genes. The deduced primary structure of xUBF is highly homologous with the human protein except for a central deletion which removes most of one HMG-box. This explains the major size difference between the X. laevis and human proteins and may well explain their different transcriptional specificities. It is shown that xUBF contains 5 tandemly repeated HMG-boxes and that by analogy the human protein contains 6.

Amino Acid Sequence↗
PubMed1995

Acanthamoeba castellanii contains a ribosomal RNA enhancer binding protein which stimulates TIF-IB binding and transcription under stringent conditions.

The intergenic spacer (IGS) of Acanthamoeba castellanii rRNA genes contains repeated elements which are weak enhancers for transcription by RNA polymerase I. A protein, EBF, was identified and partially purified which binds to the enhancers and to several other sequences within the IGS, but not to other DNA fragments, including the rRNA core promoter. No consensus binding sequence could be discerned in these fragments and bound factor is in rapid equilibrium with unbound. EBF has functional characteristics similar to vertebrate upstream binding factors (UBF). Not only does it bind to the enhancer and other IGS elements, but it also stimulates binding of TIF-IB, the fundamental transcription initiation factor, to the core promoter and stimulates transcription from the promoter. Attempts to identify polypeptides with epitopes similar to rat or Xenopus laevis UBF suggest that structurally the protein from A.castellanii is not closely related to vertebrate UBF.

Acanthamoeba↗
PubMed2004

Putative involvement of the histone acetyltransferase Tip60 in ribosomal gene transcription.

Tip60 is a histone acetyltransferase (HAT) implicated in a wide range of cellular functions, including mRNA synthesis and DNA repair. In the present report we propose a model based on which Tip60 is actively involved in ribosomal gene transcription through acetylation of UBF, a ribosomal specific transcription factor, as well as through its direct recruitment to the human ribosomal gene promoter, as shown by chromatin immunoprecipitation experiments. Electron microscopy studies revealed that Tip60 resides in sites of active rDNA transcription within the nucleolus, while it co-localizes with UBF as shown by confocal microscopy. In addition, in vivo transcription assays demonstrated that the nucleolar fraction of Tip60 localizes to sites of newly synthesized rRNA. Finally, functional assays established that Tip60 complexes with, and targets UBF for acetylation. The present study underlines the importance of acetylation in rDNA transcription and directly implicates Tip60 in the process of ribosomal gene transcription.

Acetyltransferases↗