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PubMed1993

The nucleolar transcription activator UBF relieves Ku antigen-mediated repression of mouse ribosomal gene transcription.

Previously we have shown that the RNA polymerase I (Pol I)-specific transcription factor UBF stimulates transcription by both facilitating transcription complex formation and by relieving repression exerted by a negative-acting factor which competes for binding of the murine factor TIF-IB to the ribosomal gene promoter (1). We have purified and functionally characterized this repressor protein from Ehrlich ascites cells. The final preparation contained two polypeptides with molecular masses of 75 and 90 kDa, respectively. Both polypeptides interact with the rDNA promoter as revealed by UV-crosslinking experiments. The specificity of binding to the ribosomal gene promoter was demonstrated in an electrophoretic mobility shift assay and by DNase footprinting. The biochemical properties of this negative-acting factor closely resemble those of the Ku antigen, a human nuclear DNA-binding heterodimer which is the target of autoantibodies in several autoimmune diseases. Anti-Ku antibodies precipitate the repressor activity and overcome transcription inhibition. The data demonstrate that regulation of Pol I gene transcription may involve an antirepression mechanism as already documented for Pol II genes and suggest that Ku protein may be causally involved in repressor-mediated down regulation of rRNA synthesis.

Animals↗
PubMed1992

Immunostaining of nucleolus organizers in mammalian cells by a human autoantibody against the polymerase I transcription factor UBF.

A mammalian autoantigen of the nucleolus organizing regions (NORs) was identified by a human autoantibody from the serum of a rheumatoid arthritis patient. The distribution and changes of NORs during the cell cycle of mammals were followed by using this autoantiserum in indirect-immunofluorescence microscopy. In interphase cells the staining pattern indicated that the autoantigen is restricted exclusively within the nucleolus. This fluorescence appeared punctuated rather than uniform, and it was reorganized during inhibition of transcription in cells treated with actinomycin D. During mitosis, the autoantigen was detected by light microscopy at the chromosomal nucleolus organizer regions, indicating that presumably the protein remains bound to the rRNA genes. Biochemical analysis by immunoblotting showed that the NOR autoantigen consists of two polypeptides with molecular masses apparent of 90-92 kDa in all of the mammalian cell lines tested. The identity of some epitopes, recognized by this autoantibody as the ribosomal transcription factor UBF, is discussed.

3T3 Cells↗
PubMed1994

Regulation of mouse UBF gene by multiple growth-related control elements.

Transcription of the mouse Upstream Binding Factor (mUBF) gene, that encodes one of the essential transcription factors for ribosomal DNA transcription, starts from several nucleotides. Neither typical TATA-box nor CCAAT-box is found upstream of the transcription initiation site. In the promoter region, there are eight GC-boxes, eight AP-2 binding consensus sequences, four cAMP response elements, and several serum response element equivalent sequences. These elements appear to play a positive role for the regulation of the mUBF gene as a whole. Among serum response elements, members located between -1182 and -343 are indeed responsive to serum, suggesting their important role in the high expression of mUBF under cell growth conditions.

Animals↗
PubMed1996

The species-specific RNA polymerase I transcription factor SL-1 binds to upstream binding factor.

Transcription of the 45S rRNA genes is carried out by RNA polymerase I and at least two trans-acting factors, upstream binding factor (UBF) and SL-1. We have examined the hypothesis that SL-1 and UBF interact. Coimmunoprecipitation studies using an antibody to UBF demonstrated that TATA-binding protein, a subunit of SL-1, associates with UBF in the absence of DNA. Inclusion of the detergents sodium dodecyl sulfate and deoxycholate disrupted this interaction. In addition, partially purified UBF from rat cell nuclear extracts and partially purified SL-1 from human cells coimmunoprecipitated with the anti-UBF antibody after mixing, indicating that the UBF-SL-1 complex can re-form. Treatment of UBF-depleted extracts with the anti-UBF antibody depleted the extracts of SL-1 activity only if UBF was added to the extract prior to the immunodepletion reaction. Furthermore, SL-1 activity could be recovered in the immunoprecipitate. Interestingly, these immunoprecipitates did not contain RNA polymerase I, as a monospecific antibody to the 194-kDa subunit of RNA polymerase I failed to detect that subunit in the immunoprecipitates. Treatment of N1S1 cell extracts with the anti-UBF antibody depleted the extracts of SL-1 activity but not TFIIIB activity, suggesting that the binding of UBF to SL-1 is specific and not solely mediated by an interaction between UBF and TATA-binding protein, which is also a component of TFIIIB. These data provide evidence that UBF and SL-1 interact.

Animals↗
PubMed1994

The RNA polymerase I transactivator upstream binding factor requires its dimerization domain and high-mobility-group (HMG) box 1 to bend, wrap, and positively supercoil enhancer DNA.

Upstream binding factor (UBF) is an important transactivator of RNA polymerase I and is a member of a family of proteins that contain nucleic acid binding domains named high-mobility-group (HMG) boxes because of their similarity to HMG chromosomal proteins. UBF is a highly sequence-tolerant DNA-binding protein for which no binding consensus sequence has been identified. Therefore, it has been suggested that UBF may recognize preformed structural features of DNA, a hypothesis supported by UBF's ability to bind synthetic DNA cruciforms, four-way junctions, and even tRNA. We show here that full-length UBF can also bend linear DNA to mediate circularization of probes as small as 102 bp in the presence of DNA ligase. Longer probes in the presence of UBF become positively supercoiled when ligated, suggesting that UBF wraps the DNA in a right-handed direction, opposite the direction of DNA wrapping around a nucleosome. The dimerization domain and HMG box 1 are necessary and sufficient to circularize short probes and supercoil longer probes in the presence of DNA ligase. UBF's sequence tolerance coupled with its ability to bend and wrap DNA makes UBF an unusual eukaryotic transcription factor. However, UBF's ability to bend DNA might explain how upstream and downstream rRNA gene promoter domains interact. UBF-induced DNA wrapping could also be a mechanism by which UBF counteracts histone-mediated gene repression.

Animals↗
PubMed2002

Upstream binding factor up-regulated in hepatocellular carcinoma is related to the survival and cisplatin-sensitivity of cancer cells.

Upstream binding factor (UBF) is an RNA polymerase I-specific transcription factor. By representational difference analysis, Northern blot, and cDNA array analysis, up-regulation of UBF was detected in 12 of 17 clinical hepatocellular carcinoma samples comparing to the paired normal liver tissues. Introduction of UBF in human lung fibroblast cells that do not express UBF resulted in an accelerated rate of cell growth; on the other hand, antisense oligodeoxynucleotides (ODNs) treatment of UBF-expressing hepatoma cell lines reduced the level of UBF protein, suppressed the colony formation capacity of these cells on soft agarose, and finally caused cell death. Annexin V binding analysis suggested that anti-UBF ODN-caused cell death might involve weak apoptosis, however, DNA laddering and cleavage of poly (ADP-ribose) polymerase were not observed in these ODN-treated cells. Expression profiling of the anti-UBF ODN-treated cells using a human cDNA array revealed that the expression of 30 genes was altered in response to the inhibition of UBF expression. Notably, UBF expression could increase the cell sensitivity to the chemotherapeutic reagent cis-diaminedichloroplatinum (II). We proposed that UBF is fundamental to the survival of cells expressing the gene, and is potential as a target for screening anti-cancer drugs and an indicator in selecting chemotherapeutic reagents.

Antineoplastic Agents↗
PubMed1997

Nucleosome binding by the polymerase I transactivator upstream binding factor displaces linker histone H1.

Upstream binding factor (UBF) is a vertebrate RNA polymerase I transcription factor that can bend and wrap DNA. To investigate UBF's likely role as an architectural protein of rRNA genes organized in chromatin, we tested UBF's ability to bind rRNA gene enhancers assembled into nucleosome cores (DNA plus core histones) and nucleosomes (DNA plus core histones plus histone H1). UBF bound with low affinity to nucleosome cores formed with enhancer DNA probes of 162 bp. However, on nucleosome cores which contained approximately 60 bp of additional linker DNA, UBF bound with high affinity similar to its binding to naked DNA, forming a ternary DNA-core histone-UBF complex. UBF could be stripped from ternary complexes with competitor DNA to liberate nucleosome cores, rather than free DNA, suggesting that UBF binding to nucleosome cores does not displace the core histones H2A, H2B, H3, and H4. DNase I, micrococcal nuclease, and exonuclease III footprinting suggests that UBF and histone H1 interact with DNA on both sides flanking the histone octamer. Footprinting shows that UBF outcompetes histone H1 for binding to a nucleosome core and will displace, if not dissociate, H1 from its binding site on a preassembled nucleosome. These data suggest that UBF may act to prevent or reverse the assembly of transcriptionally inactive chromatin structures catalyzed by linker histone binding.

Animals↗
PubMed2004

Development and use of an ovarian synchronization model to study the effects of endogenous estrogen and nitric oxide on uterine blood flow during ovarian cycles in sheep.

The objective of the current study was to develop an ovine animal model for consistent study of uterine blood flow (UBF) changes during synchronized ovarian cycles regardless of season. Sheep were surgically bilaterally instrumented with uterine artery blood flow transducers and 5-7 days later implanted with a vaginal progesterone (P(4))-controlled internal drug-releasing device (CIDR; 0.3 g) for 7 days. On Day 6 of P(4), sheep were given two prostaglandin F(2 alpha) injections (7.5 mg i.m. 4 h apart). At CIDR removal, Experimental Day 0, zero (n = 9), 500 IU (n = 8), or 1000 IU (n = 7) eCG was injected i.m.; UBF was monitored continuously for 55-75 h. Jugular blood was sampled every 8 h to evaluate levels of P(4), estradiol-17 beta (E(2)beta) and luteinizing hormone (LH). The inhibitor of nitric oxide synthase, L-nitro-arginine methyl ester (L-NAME) was infused in a stepwise fashion unilaterally into one uterine artery at 48-50 h after 500 IU eCG and the effects on UBF were examined (n = 7). The zero-eCG group gradually increased UBF from a baseline of 17.4 +/- 3.9 to 80.5 +/- 1.1 ml/min. The 500-IU-eCG group increased UBF between 10 and 15 h from a baseline of 11 +/- 3.3 to 83.3 +/- 1.0 ml/min, whereas UBF for the 1000-IU-eCG group was higher (100.1 +/- 1.7 ml/min) than that seen in either of the other groups. Plasma P(4) fell to baseline within 8 h of CIDR removal, while E(2)beta rose gradually in association with elevations in UBF. LH surges occurred between 32 and 56 h after CIDR removal and the LH surge occurred earlier in the 1000-IU-eCG group than the other two groups (P < 0.01). L-NAME infusion dose dependently reduced maximum levels of UBF ipsilaterally by 54.6% +/- 6.2%, but contralaterally only by 27.4% +/- 8.5%. Regardless of season, either dose of eCG will result in analogous UBF responses. During the follicular phase, elevations in UBF are in part locally controlled by the de novo production of nitric oxide.

Animals↗
PubMed1992

Cooperative binding of the Xenopus RNA polymerase I transcription factor xUBF to repetitive ribosomal gene enhancers.

Upstream binding factor (UBF) is a DNA-binding transcription factor implicated in ribosomal gene promoter and enhancer function in vertebrates. UBF is unusual in that it has multiple DNA-binding domains with homology to high-mobility-group (HMG) nonhistone chromosomal proteins 1 and 2. However, a recognizable DNA consensus sequence for UBF binding is lacking. In this study, we have used gel retardation and DNase I footprinting to examine Xenopus UBF (xUBF) binding to Xenopus laevis ribosomal gene enhancers. We show that UBF has a minimum requirement for about 60 bp of DNA, the size of the short enhancer variant in X. laevis. Stronger UBF binding occurs on the longer enhancer variant (81 bp) and on multiple enhancers linked head to tail. In vivo, Xenopus ribosomal gene enhancers exist in blocks of 10 alternating 60- and 81-bp repeats within the intergenic spacer. In vitro, UBF binds cooperatively to probes with 10 enhancers, with five intermediate complexes observed in titration experiments. This suggests that, on average, one UBF dimer binds every two enhancers. A single UBF dimer can produce a DNase I footprint ranging in size from approximately 30 to about 115 bp on enhancer probes of different lengths. This observation is consistent with the hypothesis that multiple DNA-binding domains or subdomains within UBF bind independently, forming more-stable interactions on longer probes.

Animals↗
PubMed1985

Effect of graded reductions in uteroplacental blood flow on the fetal lamb.

Graded reductions in uteroplacental blood flow (UBF), produced by an externally adjustable vascular occluder, were studied in unanesthetized pregnant sheep. This method was designed to simulate the physiological occurrence of reduced UBF to study its effects on fetal status. Sheep were divided into four groups in which average percent changes (+/- SE) in UBF, as measured by electromagnetic flow probes, were 7 +/- 2, -24 +/- 1, -49 +/- 1, and -63 +/- 2% for 1 h. Maternal and fetal cardiovascular (heart rate and blood pressure) and blood gas measurements of PO2 and PCO2, pH, and O2 content were determined before, during, and after recovery from UBF reduction. UBF reduction had no significant effect on maternal measurements. Changes in fetal PO2 and O2 content were directly related to the extent of vascular occlusion (P less than or equal to 0.05). Fetal pH markedly decreased after 1 h of 49% UBF reduction (P less than or equal to 0.05) and within 10 min after 63% UBF reduction (P less than or equal to 0.05). Fetal bradycardia (P less than or equal to 0.05) resulted from 63% reduction in UBF. Fetal heart rates at recovery were greater than base-line heart rate values (P less than or equal to 0.05) in the three groups whose UBF was restricted. The results of the present study suggest that the uterus of the pregnant ovine is perfused in sufficient excess to protect the fetus during short-term reductions of less than 50% UBF.

Animals↗
PubMed1996

Regulation of rDNA transcription during endothelin-1-induced hypertrophy of neonatal cardiomyocytes. Hyperphosphorylation of upstream binding factor, an rDNA transcription factor.

Treatment of cultured neonatal cardiomyocytes with endothelin-1 and phorbol 12-myristate 13-acetate (PMA) results in cardiomyocyte hypertrophy. However, the signal transduction pathways involved in this process are poorly understood. Because increased ribosome biogenesis is a requisite for hypertrophy, we sought to (1) confirm the hypothesis that these two hypertrophic agents did indeed induce rRNA synthesis and (2) examine the mechanism through which this induction was accomplished. In this study, hypertrophy of contraction-arrested neonatal cardiomyocytes induced by treatment with either endothelin-1 or PMA was associated with increased rDNA transcription. Western blots demonstrated that the enhanced rates of rDNA transcription were not mediated by increased amounts of either RNA polymerase I or upstream binding factor (UBF), an rDNA transcription factor. However, immunoprecipitation of [32P] orthophosphate-labeled UBF from hypertrophying neonatal cardiomyocytes suggested that the increased rate of rDNA transcription may be due to the hyperphosphorylation of UBF, which would increase the activity of UBF. The increase in UBF phosphorylation occurred within 3 to 6 hours after exposure to either agent, was maximal at 12 hours, and was sustained for at least the first 24 hours of exposure. Phosphoamino acid analysis of UBF immunoprecipitated from control and treated cardiomyocytes demonstrated that UBF was phosphorylated exclusively on serine residues. Our previous studies have shown that the cellular UBF content increased in adrenergic- and contraction-induced models of cardiac hypertrophy. This study with endothelin-1 and PMA demonstrates that the modulation of UBF phosphorylation is an additional pathway by which ribosome biogenesis may be regulated in neonatal cardiomyocytes. These results support the hypothesis that UBF is an important regulatory factor during the initiation and maintenance of the accelerated rate of rDNA transcription observed during neonatal cardiomyocyte hypertrophy mediated by both phorbol esters and endothelin-1.

Animals↗
PubMed2005

Objective and subjective upper body function six months following diagnosis of breast cancer.

Whether based on self-reported disability or direct physical measurements, studies of upper body function (UBF) among women with breast cancer consistently reveal high prevalence of limitation. Unfortunately, limited research and conflicting results undermine the ability to identify those factors that contribute to these UBF problems. This study describes the influence of selected personal and treatment characteristics on UBF, six-months following treatment for unilateral breast cancer, in a population-based sample of women residing in Southeast Queensland, Australia (n = 214). UBF was assessed by three objective measures (upper body strength and endurance, flexibility and hand grip strength) and two subjective measures (Disability of the Arm, Shoulder and Hand questionnaire and arm morbidity scale of the Functional Assessment of Cancer Treatment, Breast questionnaire). Modest correlations between the various UBF measures suggest that each captures a somewhat different dimension of physical function and disability. Advancing age, being treated on the non-dominant side, more childcare responsibilities, lower socioeconomic status, more extensive lymph node removal, having lymphoedema, and UBF scores of the untreated side emerged as significant correlates depending on whether UBF was assessed objectively or subjectively. There also was evidence that the relationships between the UBF measure of flexibility and (i) radiation treatment and (ii) side of treatment were modified depending on dominance. These results highlight the necessity of taking dominance into account when assessing functional impairment among breast cancer survivors. The findings also highlight the importance of considering whether subjective or objective UBF measures are more relevant in the design of future studies.

Activities of Daily Living↗
PubMed2005

Large-conductance Ca2+-dependent K+ channels regulate basal uteroplacental blood flow in ovine pregnancy.

OBJECTIVES: The mechanisms regulating basal uteroplacental blood flow (UBF) and the greater than 30-fold increase observed in normal pregnancy remain unclear. Although vascular growth contributes in early gestation, vasodilation accounts for the exponential rise seen in the last third of pregnancy. Large conductance potassium channels (BK(Ca)) are expressed in uterine vascular smooth muscle (VSM), but the extent of their role in regulating UBF in pregnancy is unclear. Therefore, we determined if BK(Ca) regulate basal UBF during ovine pregnancy. METHODS: Studies were performed at 113 to 127 days and 135 to 150 days of gestation in eight pregnant ewes instrumented with uterine artery flow probes and uterine arterial and venous catheters. Tetraethylammonium chloride (TEA), a BK(Ca)-specific inhibitor at less than 1.0 mM, was infused intra-arterially into the pregnant uterine horn over 60 minutes to achieve levels of 0.001-0.35 mM while continuously monitoring UBF, arterial pressure (MAP), and heart rate (HR). Uterine arterial and venous blood was collected simultaneously to measure uterine cyclic guanosine monophosphate (cGMP) synthesis. RESULTS: Intra-arterial TEA dose-dependently decreased basal UBF in the early (R = 0.81, n = 36, P <.001) and late (R = 0.72, n = 31, P <.001) study periods without altering contralateral UBF, MAP, and HR. The IC(50) was 0.2 mM and basal UBF decreased >or=80% at 0.35 mM in both periods. Although UBF fell greater than 40% at estimated plasma TEA levels of 0.3 mM, uterine arterial cGMP was unchanged, uterine venous cGMP rose, and uterine cGMP synthesis was unchanged; therefore, upstream events associated with BK(Ca) activation were unaffected by blockade. CONCLUSIONS: These are the first data demonstrating that BK(Ca) are essential in the maintenance of basal UBF in the last third of ovine pregnancy.

Animals↗
PubMed1997

Involvement of in situ conformation of ribosomal genes and selective distribution of upstream binding factor in rRNA transcription.

The distribution of the ribosomal genes (rDNA) and the upstream binding factor (UBF), correlatively with their RNA transcripts, was investigated in G1, S-phase, and G2. rDNA was distributed in nucleoli, with alternate sites of clustered and dispersed genes. UBF was found associated with some but not all clustered genes and proportionally more with dispersed genes. It was distributed in several foci that were more numerous and heterogeneous in size during G2 than G1. We suggest that UBF associated with rDNA during S-phase because its nucleolar amount increased during that time and remained stable in G2. 5,6-Dichloro-1-beta-D-ribofuranosylbenzimidazole treatment indicated a similar amount of UBF per transcription unit, and consequently heterogeneous size of the UBF foci can represent a variable number of transcription units per foci. Direct visualization of the transcripts demonstrated that only part of UBF is associated with active transcription and that rDNA distribution varied with transcription. We propose that in the same rDNA locus three types of configuration coexist that are correlated with gene activity: 1) clustered genes without UBF; 2) clustered genes with UBF, of which some are associated with transcription; and 3) dispersed genes with UBF and transcription. These results support the hypothesis that rDNA transcription involved several steps of regulation acting successively and locally in the same locus to promote the repressed clustered genes to become actively transcribed dispersed genes.

Animals↗
PubMed2005

Uterine blood flow responses to ICI 182 780 in ovariectomized oestradiol-17beta-treated, intact follicular and pregnant sheep.

Oestrogen dramatically increases uterine blood flow (UBF) in ovariectomized (Ovx) ewes. Both the follicular phase and pregnancy are normal physiological states with elevated levels of circulating oestrogen. ICI 182 780 is a pure steroidal oestrogen receptor (ER) antagonist that blocks oestrogenic actions in oestrogen-responsive tissue. We hypothesized that an ER-mediated mechanism is responsible for in vivo rises in UBF in physiological states of high oestrogen. The purpose of the study was to examine the effect of an ER antagonist on exogenous and endogenous oestradiol-17beta (E2beta)-mediated elevations in UBF. Sheep were surgically instrumented with bilateral uterine artery blood flow transducers, and uterine and femoral artery catheters. Ovx animals (n = 8) were infused with vehicle (35% ethanol) or ICI 182 780 (0.1-3.0 microg min(-1)) into one uterine artery for 10 min before and 50 min after E2beta was given (1 microg kg(-1) I.V. bolus) and UBF was recorded for an additional hour. Intact, cycling sheep were synchronized to the follicular phase using progesterone, prostaglandin F2alpha(PGF2alpha) and pregnant mare serum gonadotrophin (PMSG). When peri-ovulatory rises in UBF reached near peak levels, ICI 182 780 (1 or 2 microg (ml uterine blood flow)-1) was infused unilaterally (n = 4 sheep). Ewes in the last stages of pregnancy (late pregnant ewes) were also given ICI 182 780 (0.23-2.0 microg (ml uterine blood flow)-1; 60 min infusion) into one uterine artery (n = 8 sheep). In Ovx sheep, local infusion of ICI 182 780 did not alter systemic cardiovascular parameters, such as mean arterial blood pressure or heart rate; however, it maximally decreased ipsilateral, but not contralateral, UBF vasodilatory responses to exogenous E2beta by approximately 55-60% (P < 0.01). In two models of elevated endogenous E2beta, local ICI 182 780 infusion inhibited the elevated UBF seen in follicular phase and late pregnant ewes in a time-dependent manner by approximately 60% and 37%, respectively; ipsilateral >> contralateral effects (P < 0.01). In late pregnant sheep ICI 182 780 also mildly and acutely (for 5-30 min) elevated mean arterial pressure and heart rate (P < 0.05). We conclude that exogenous E2beta-induced increases in UBF in the Ovx animal and endogenous E2beta-mediated elevations of UBF during the follicular phase and late pregnancy are partially mediated by ER-dependent mechanisms.

Animals↗
PubMed1996

Nitric oxide contributes to estrogen-induced vasodilation of the ovine uterine circulation.

Estradiol-17beta (E2beta), a potent vasodilator, has its greatest effects on the uterine vasculature, blood flow (UBF) increasing > or = 10-fold. The mechanism(s) responsible for E2beta-induced vasodilation is unclear. We determined if nitric oxide (NO)-induced increases in cGMP modulate estrogen-induced increases in UBF, and if cyclooxygenase inhibition modifies E2beta responses. Nonpregnant (n = 15) and pregnant (n = 8) ewes had flow probes implanted on main uterine arteries and catheters in branches of the uterine vein and artery bilaterally for blood sampling and infusion of the NO synthase inhibitor L-nitro-arginine methyl ester (L-NAME), respectively. In nonpregnant ewes E2beta (1 microg/kg) caused parallel increases (P < 0.001) in UBF (15+/-3 to 130+/-16 ml/min) and uterine cGMP secretion (23+/-10 to 291+/-38 pmol/min); uterine venous cGMP also rose (4.98+/-1.4 to 9.43+/-3.2 pmol/ml; P < 0.001). Intra-arterial L-NAME partially inhibited increases in UBF dose-dependently (r = 0.66, n = 18, P < 0.003) while completely inhibiting cGMP secretion (P = 0.025). Indomethacin, 2 mg/kg intravenously, did not alter E2beta-induced responses. After E2beta-induced increases in UBF, intraarterial L-NAME partially decreased UBF dose dependently (r = 0.73, n = 46, P < 0.001) while inhibiting cGMP secretion (178+/-48 to 50+/-24 pmol/min; n = 5, P = 0.006); both were reversed by L-arginine. In pregnant ewes, E2beta increased UBF and venous cGMP (9.1+/-0.96 to 13.2+/-0.96 pmol/ml, P < 0.01); however, intraarterial L-NAME decreased basal cGMP secretion 66% (P = 0.02), but not UBF. Acute estrogen-induced increases in UBF are associated with NO-dependent increases in cGMP synthesis, but other mechanisms may also be involved. However, vasodilating prostanoids do not appear to be important. In ovine pregnancy NO is not essential for maintaining uteroplacental vasodilation.

Animals↗
PubMed2002

[Activation of the ribosomal gene during blast transformation of human lymphocytes].

BACKGROUND: Natural lectin, phytohemagglutinin, initiates the transformation of normally quiescent T lymphocytes into proliferating lymphoblast-like cells. Recently we have shown that the transformation is accompanied by strong promotion of ribosomal RNA synthesis and by phosphorylation of its activator, initiating factor UBF, both culminating in a synthetic phase of the first cell division cycle. In contrast we have revealed that the UBF gene was activated and its transcription culminated in the early G1 phase. We examined three possible delaying mechanisms: the kinetics of unwinding of rDNA chromatin, the kinetics of transcription of genes coding for the second initiating factor, SL1 complex, and the kinetics of the translation of UBF protein product. METHODS AND RESULTS: Up to 48 hrs following the addition of phytohemagglutinin to the growth medium, we monitored structural changes in the rDNA chromatin using indirect antiUBF immunofluorescence. The data indicated an increased number of separated transcriptional units during the G1 phase of the first cycle. In a time interval of up to 70 hrs we measured the mRNA levels of four constituents of SL1 complex: TAF110, TAF63, TAF48 and TBP using the RT-PCR method. We found a close correlation between the kinetics of the transcription of UBF and SL1 genes and the maximal rate in the early G1 phase. Using metabolic labelling with 35S methionine/cysteine we monitored the translation of UBF protein in PHA stimulated lymphocytes. The data suggested that UBF translation, starting in the S phase, paralleled chromosomal DNA replication. CONCLUSIONS: During the transformation of normal T lymphocytes into proliferating blast-like cells, the multicopy rDNA gene unwinds in the G1 phase of the first cycle forming individual transcriptional units. Genes coding for factors which initiate synthesis of ribosomal precursors are activated in the early G1 phase. The G1 synthesis of ribosomal RNA is accelerated by phosphorylation of the hypophosphorylated UBF pool. As blastic transformation develops UBF translation is triggered in the S phase and neosynthesized UBF, activated by phosphorylation, pushes the synthesis of ribosomal precursors to maximal efficiency. The process of blastic transformation interferes throughout the entire prolonged G1 phase of the first cell division cycle.

DNA, Ribosomal↗