Translational control of embryonic cell division by CPEB and maskin.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
R Mendez
Publications and source records attributed to R Mendez.
CPEB phosphorylation and cytoplasmic polyadenylation are catalyzed by the kinase IAK1/Eg2 in maturing mouse oocytes.
In both vertebrates and invertebrates, the expression of several maternal mRNAs is regulated by cytoplasmic polyadenylation. In Xenopus oocytes, where most of the biochemical details of this process have been examined, polyadenylation is controlled by CPEB, a sequence-specific RNA binding protein. The activity of CPEB, which is to recruit cleavage and polyadenylation specificity factor (CPSF) and poly(A) polymerase (PAP) into an active cytoplasmic polyadenylation complex, is controlled by Eg2-catalyzed phosphorylation. Soon after CPEB phosphorylation and resulting polyadenylation take place, the interaction between maskin, a CPEB-associated factor, and eIF4E, the cap-binding protein, is destroyed, which results in the recruitment of mRNA into polysomes. Polyadenylation also occurs in maturing mouse oocytes, although the biochemical events that govern the reaction in these cells are not known. In this study, we have examined the phosphorylation of CPEB and have assessed the necessity of this protein for polyadenylation in maturing mouse oocytes. Immunohistochemistry has revealed that all the factors that control polyadenylation and translation in Xenopus oocytes (CPEB, CPSF, PAP, maskin, and IAK1, the murine homologue of Eg2) are also present in the cytoplasm of mouse oocytes. After the induction of maturation, a kinase is activated that phosphorylates CPEB on a critical regulatory residue, an event that is essential for CPEB activity. A peptide that competitively inhibits the activity of IAK1/Eg2 blocks the progression of meiosis in injected oocytes. Finally, a CPEB protein that acts as a dominant negative mutation because it cannot be phosphorylated by IAK1/Eg2, prevents cytoplasmic polyadenylation. These data indicate that cytoplasmic polyadenylation in mouse oocytes is mediated by IAK1/Eg2-catalyzed phosphorylation of CPEB.
Amino Acid Sequence↗
CPEB, maskin, and cyclin B1 mRNA at the mitotic apparatus: implications for local translational control of cell division.
In Xenopus development, the expression of several maternal mRNAs is regulated by cytoplasmic polyadenylation. CPEB and maskin, two factors that control polyadenylation-induced translation are present on the mitotic apparatus of animal pole blastomeres in embryos. Cyclin B1 protein and mRNA, whose translation is regulated by polyadenylation, are colocalized with CPEB and maskin. CPEB interacts with microtubules and is involved in the localization of cyclin B1 mRNA to the mitotic apparatus. Agents that disrupt polyadenylation-induced translation inhibit cell division and promote spindle and centrosome defects in injected embryos. Two of these agents inhibit the synthesis of cyclin B1 protein and one, which has little effect on this process, disrupts the localization of cyclin B1 mRNA and protein. These data suggest that CPEB-regulated mRNA translation is important for the integrity of the mitotic apparatus and for cell division.
Animals↗
Phosphorylation of CPE binding factor by Eg2 regulates translation of c-mos mRNA.
Full-grown Xenopus oocytes arrest at the G2/M border of meiosis I. Progesterone breaks this arrest, leading to the resumption of the meiotic cell cycles and maturation of the oocyte into a fertilizable egg. In these oocytes, progesterone interacts with an unidentified surface-associated receptor, which induces a non-transcriptional signalling pathway that stimulates the translation of dormant c-mos messenger RNA. Mos, a mitogen-activated protein (MAP) kinase kinase kinase, indirectly activates MAP kinase, which in turn leads to oocyte maturation. The translational recruitment of c-mos and several other mRNAs is regulated by cytoplasmic polyadenylation, a process that requires two 3' untranslated regions, the cytoplasmic polyadenylation element (CPE) and the polyadenylation hexanucleotide AAUAAA. Although the signalling events that trigger c-mos mRNA polyadenylation and translation are unclear, they probably involve the activation of CPEB, the CPE binding factor. Here we show that an early site-specific phosphorylation of CPEB is essential for the polyadenylation of c-mos mRNA and its subsequent translation, and for oocyte maturation. In addition, we show that this selective, early phosphorylation of CPEB is catalysed by Eg2, a member of the Aurora family of serine/threonine protein kinases.
Amino Acid Sequence↗
Safety and efficacy of tacrolimus in combination with mycophenolate mofetil (MMF) in cadaveric renal transplant recipients. FK506/MMF Dose-Ranging Kidney Transplant Study Group.
BACKGROUND: Tacrolimus (FK506) is a safe and effective treatment for the prevention of rejection of renal allografts. Mycophenolate mofetil (MMF) has been used as adjunct immunosuppressive therapy with cyclosporine and corticosteroids for the same purpose. The objective of this study was to investigate the safety and efficacy of FK506 and MMF in renal transplant recipients. METHODS: After cadaveric renal transplant, patients were randomized to receive tacrolimus in combination with either azathioprine (AZA, n=59), MMF 1 g/day (n=59), or MMF 2 g/day group (n=58). Patients were followed for 1 yr posttransplant for the incidence of biopsy-confirmed acute rejection, patient and graft survival, and adverse events. RESULTS: Tacrolimus doses and trough concentrations were similar between treatment groups at all time points; 80% of patients were maintained within a range of 5.0-13.9 ng/ml at 12 months posttransplant. The mean dose of MMF decreased in the 2 g/day group to 1.5 g/day by 6 months posttransplant, primarily due to gastrointestinal GI-related disorders. The incidence of biopsy-confirmed acute rejection at 1 year was 32.2%, 32.2%, and 8.6% in the AZA, MMF 1 g/day, and MMF 2 g/day groups, respectively (P<0.01). The use of antilymphocyte antibodies for the treatment of rejection was comparable across treatment groups. The incidence of most adverse events was similar across treatment groups and comparable with previous reports. The overall incidence of posttransplant diabetes mellitus was 11.9%, with the lowest rate observed in the MMF 2 g/day group (4.7%), and was reversible in 40% of patients. The incidence of malignancies and opportunistic infections was low and not different across treatment groups. CONCLUSION: Tacrolimus in combination with an initial dose of MMF 2 g/day is a very effective and safe regimen in cadaveric kidney transplant recipients.
Acute Disease↗
A mutation determining the loss of HLA-A2 antigen expression in a cervical carcinoma reveals novel splicing of human MHC class I classical transcripts in both tumoral and normal cells.
Explore the source record for details and available documents.
Alternative Splicing↗
Phosphorylation of CPEB by Eg2 mediates the recruitment of CPSF into an active cytoplasmic polyadenylation complex.
The release of Xenopus oocytes from prophase I arrest is largely driven by the cytoplasmic polyadenylation-induced translation of dormant maternal mRNAs. Two cis elements, the CPE and the hexanucleotide AAUAAA, and their respective binding factors, CPEB and a cytoplasmic form of CPSF, control polyadenylation. The most proximal stimulus for polyadenylation is Eg2-catalyzed phosphorylation of CPEB serine 174. Here, we show that this phosphorylation event stimulates an interaction between CPEB and CPSF. This interaction is direct, does not require RNA tethering, and occurs through the 160 kDa subunit of CPSF. Eg2-stimulated and CPE-dependent polyadenylation is reconstituted in vitro using purified components. These results demonstrate that the molecular function of Eg2-phosphorylated CPEB is to recruit CPSF into an active cytoplasmic polyadenylation complex.
Animals↗
Kinetic study of addition of volatile organic compounds to a nitrifying sludge.
The effects of different concentrations of several volatile organic compounds (VOC) such as ethanol, acetate, propionate, and butyrate ranging from 0 to 2000 mg/L as well as a mixture of volatile fatty acids (MVFA) at a 4:1:1 (acetate:propionate:butyrate) ratio on the nitrification rate in batch cultures were studied. The results showed that ethanol and acetate were consumed in a mixotrophic way by the nitrifying sludge. At a concentration of 500 mg/L, the nitrification rate of inhibition was different for each compound in which propionate and butyrate were the most inhibitory. At 2000 mg/L the inhibition was 80% with ethanol and 100% with acetate, propionate, and butyrate. With similar concentrations of MVFA, the inhibition was also similar to that in acetate. The effect of the addition of pulses of MVFA at a ratio of 4:1:1 during 14 h on the performance of the continuous nitrifying process with a hydraulic retention time of 3 d was also studied. No inhibition of the nitrification process was observed with pulses of 750-3000 mg of MVFA/L in the input of the reactor. The results in batch cultures suggest that the different degrees of inhibition of the nitrification process were related to the type of organic matter added. The noninhibitory effect of the organic matter in the continuous cultures on the nitrification efficiency of the nitrifying sludge might be related to the feeding pattern.
Acetates↗
Valacyclovir for the prevention of cytomegalovirus disease after renal transplantation. International Valacyclovir Cytomegalovirus Prophylaxis Transplantation Study Group.
BACKGROUND: Cytomegalovirus (CMV) disease is a major complication of organ transplantation. We hypothesized that prophylactic treatment with valacyclovir would reduce the risk of CMV disease. METHODS: A total of 208 CMV-negative recipients of a kidney from a seropositive donor and 408 CMV-positive recipients were randomly assigned to receive either 2 g of valacyclovir or placebo orally four times daily for 90 days after transplantation, with the dose adjusted according to renal function. The primary end point was laboratory-confirmed CMV disease in the first six months after transplantation. RESULTS: Treatment with valacyclovir reduced the incidence or delayed the onset of CMV disease in both the seronegative patients (P<0.001) and the seropositive patients (P=0.03). Among the seronegative patients, the incidence of CMV disease 90 days after transplantation was 45 percent among placebo recipients and 3 percent among valacyclovir recipients. Among the seropositive patients, the respective values were 6 percent and 0 percent. At six months, the incidence of CMV disease was 45 percent among seronegative recipients of placebo and 16 percent among seronegative recipients of valacyclovir; it was 6 percent among seropositive placebo recipients and 1 percent among seropositive valacyclovir recipients. At six months, the rate of biopsy-confirmed acute graft rejection in the seronegative group was 52 percent among placebo recipients and 26 percent among valacyclovir recipients (P=0.001). Treatment with valacyclovir also decreased the rates of CMV viremia and viruria, herpes simplex virus disease, and the use of inpatient medical resources. Hallucinations and confusion were more common with valacyclovir treatment, but these events were not severe or treatment-limiting. The rates of other adverse events were similar among the groups. CONCLUSIONS: Prophylactic treatment with valacyclovir is a safe and effective way to prevent CMV disease after renal transplantation.
Acyclovir↗
Reduced intrapatient variability of cyclosporine pharmacokinetics in renal transplant recipients switched from oral Sandimmune to Neoral.
Sixty-six renal transplant patients maintained on Sandimmune, the traditional formulation of cyclosporine, participated in an open-label, sequential trial to compare intrapatient variability in drug exposure before and after a switch to Neoral. Three 12-hour cyclosporine pharmacokinetic profiles were obtained over approximately 6 weeks while patients were receiving Sandimmune. Patients were then switched to Neoral, with the dose adjusted as necessary to maintain target trough blood cyclosporine concentrations. At approximately 4 and 6 weeks postconversion, 2 additional pharmacokinetic profiles were obtained. Key pharmacokinetic variables analyzed were area under the concentration-time curve (AUC), maximum concentration (Cmax), and predose trough concentration (C0). Intrapatient variability in drug exposure for dose-normalized mean AUC, Cmax, and C0 was significantly reduced with Neoral, with 50 (76%), 57 (86%), and 45 (68%) patients experiencing reduced variability in AUC, Cmax, and C0, respectively (P < 0.001). Additionally, the total exposure to cyclosporine was more predictable from the trough level of cyclosporine with Neoral; the relationship between AUC and C0 was 0.81 for Neoral at both pharmacokinetic profiling time points but ranged from 0.49 to 0.69 for the 3 Sandimmune time points. The large reductions in intrapatient variability of pharmacokinetic variables for cyclosporine provided by Neoral indicate an improvement in the consistency of drug exposure, which may translate into important clinical benefits.
Adult↗
Maskin is a CPEB-associated factor that transiently interacts with elF-4E.
In Xenopus, the CPE is a bifunctional 3' UTR sequence that maintains maternal mRNA in a dormant state in oocytes and activates polyadenylation-induced translation during oocyte maturation. Here, we report that CPEB, which binds the CPE and stimulates polyadenylation, interacts with a new factor we term maskin. Maskin contains a peptide sequence that is conserved among elF-4E-binding proteins. Affinity chromatography demonstrates that CPEB, maskin, and elF-4E reside in a complex in oocytes, and yeast two-hybrid analyses indicate that CPEB and maskin bind directly, as do maskin and elF-4E. While CPEB and maskin remain together during oocyte maturation, the maskin-elF-4E interaction is substantially reduced. The dissolution of this complex may result in the binding of elF-4E to elF-4G and the translational activation of CPE-containing mRNAs.
3' Untranslated Regions↗
In vitro susceptibilities of clinical yeast isolates to the new antifungal eberconazole compared with their susceptibilities to clotrimazole and ketoconazole.
The antifungal activity of eberconazole, a new imidazole derivative, against 124 clinical isolates of Candida comprising eight different species and to 34 isolates of Cryptococcus neoformans was compared to those of clotrimazole and ketoconazole. MICs of eberconazole, determined by the National Committee for Clinical Laboratory Standards standardized microbroth method, were equal to or lower than those of other azoles, especially for Candida krusei and Candida glabrata, which are usually resistant to triazoles.
Antifungal Agents↗
Using gadolinium-enhanced three-dimensional MR angiography to assess arterial inflow stenosis after kidney transplantation.
OBJECTIVE: Our objective was to evaluate use of gadolinium-enhanced three-dimensional (3D) MR angiography in the assessment of suspected arterial inflow stenosis after kidney transplantation. SUBJECTS AND METHODS: Twenty-eight consecutive patients receiving kidney transplants (26 single-kidney transplants and two en block transplants) with suspected arterial inflow stenosis were examined with two MR angiography sequences: gadolinium-enhanced 3D fast spoiled gradient-recalled (SPGR) imaging and 3D phase-contrast imaging. Twenty-four of these patients then were examined using the gold standards: either digital subtraction angiography (DSA) (n = 23) or surgery (n = 1). MR angiography and DSA studies were independently and prospectively analyzed for the presence of arterial stenoses (mild [<50%], severe [50-90%], or critical [>90%]) in the iliac, anastomotic, and renal artery segments. Two independent observers retrospectively evaluated the MR angiography sequences for ability to detect or exclude significant (> or = 50%) arterial stenoses. RESULTS: In 22 single-kidney transplants, DSA showed eight significant stenoses in 66 arterial segments. MR angiograms adequately showed 66 of 66 segments (prospective observers) and 64 of 66 segments (each retrospective observer), which were subsequently evaluated. The sensitivity and specificity of MR angiography in revealing significant stenoses were 100% and 98% (prospective analysis), 88% and 98% (retrospective observer 1), and 86% and 100% (retrospective observer 2). Concordance between observers showed kappa values exceeding .85 for all comparisons except the analysis of phase-contrast series (kappa = .62). In one en block transplant, DSA showed that stenosis was greater than 90%, although it had been graded at less than 50% with MR angiography. CONCLUSION: Gadolinium-enhanced 3D MR angiography accurately evaluated arterial inflow in single-kidney transplants.
Angiography, Digital Subtraction↗
The influence of donor age on kidney graft survival in the 1990s.
Based on analyses of the UNOS Registry data for cadaver kidney transplants performed between 1991-1999 we showed that: 1. 15-40 year old donor kidneys provided the best one-year graft survival rates. When donors were analyzed with recipients, younger (0-10) and older (70-90) donors and recipients (Table 2) had the lowest one-year graft success rates. 2. Chronic loss rate, the constant rate of graft loss between one and 5 years, showed younger donor kidneys had a significantly lower chronic loss rate compared with each older donor category. Apparently the younger donor kidneys have a resiliency and nephron reserve that provides better long-term function. However, they may have lower short-term (1-yr) graft survival rates, possibly due to their small size. 3. Black and White donor kidneys had similar one-year graft survival rates; however, in every age group, recipients of White donor kidneys had significantly better 5-year graft survival rates than Black donor kidneys. There was also a noticeably lower chronic loss rate among recipients of White than Black donor kidneys. 4. HLA-matched White donor kidneys had better one- and 5-year graft survival rates and lower chronic loss rates than HLA-mismatched kidneys. The matching effect was lost when the donor age increased beyond age 40. PRA had an effect both at one and 5 years after transplantation. The chronic loss rate was similar with high and low PRA. Therefore, PRA had a relatively short-term effect. 5. Cold ischemia time had a modest effect after 35 hours both at one and 5 years. However, the chronic loss rate was unaffected by CIT, suggesting prolonged ischemia time had a relatively short-term effect. 6. More focused attention on sensitization and lowered CIT can both have a significant effect on short-term graft survival rates. However, both matching and younger donor organs provide the best opportunity for better long-term graft success rates.
Adolescent↗
The COOH-terminal tyrosine phosphorylation sites on IRS-1 bind SHP-2 and negatively regulate insulin signaling.
Activation of tyrosine kinases by numerous growth factor and cytokine receptors leads to tyrosine phosphorylation of the insulin receptor substrate (IRS)-proteins. Tyrosine-phosphorylated motifs on the IRS proteins bind to the SH2 domains in proteins that mediate downstream signals, including phosphatidylinositol 3'-kinase, GRB-2, and SHP-2. We investigated the function of the two SHP-2 binding COOH-terminal tyrosines of IRS-1 by replacing them with phenylalanine (IRS-1(FCT)). IRS-1(FCT) failed to bind SHP-2 or mediate its tyrosine phosphorylation during insulin stimulation. Although several reports suggest a critical role for SHP-2 in insulin stimulated mitogen-activated protein kinase activation and cell proliferation, IRS-1(FCT) mediated these effects normally in 32D cells. Indeed, IRS-1(FCT) exhibited increased tyrosine phosphorylation, phosphatidylinositol 3'-kinase binding and activation of protein synthesis in response to insulin. These results suggest that SHP-2 attentuates the phosphorylation and downstream signal transmission of IRS-1 and that the interaction of IRS-1 and SHP-2 is an important regulatory event which attenuates insulin metabolic responses.
Animals↗
Results of the double-blind, randomized, multicenter, phase III clinical trial of Thymoglobulin versus Atgam in the treatment of acute graft rejection episodes after renal transplantation.
BACKGROUND: Thymoglobulin, a rabbit anti-human thymocyte globulin, was compared with Atgam, a horse anti-human thymocyte globulin for the treatment of acute rejection after renal transplantation. METHODS: A multicenter, double-blind, randomized trial with enrollment stratification based on standardized histology (Banff grading) was conducted. Subjects received 7-14 days of Thymoglobulin (1.5 mg/kg/ day) or Atgam (15 mg/kg/day). The primary end point was rejection reversal (return of serum creatinine level to or below the day 0 baseline value). RESULTS: A total of 163 patients were enrolled at 25 transplant centers in the United States. No differences in demographics or transplant characteristics were noted. Intent-to-treat analysis demonstrated that Thymoglobulin had a higher rejection reversal rate than Atgam (88% versus 76%, P=0.027, primary end point). Day 30 graft survival rates (Thymoglobulin 94% and Atgam 90%, P=0.17), day 30 serum creatinine levels as a percentage of baseline (Thymoglobulin 72% and Atgam 80%; P=0.43), and improvement in posttreatment biopsy results (Thymoglobulin 65% and Atgam 50%; P=0.15) were not statistically different. T-cell depletion was maintained more effectively with Thymoglobulin than Atgam both at the end of therapy (P=0.001) and at day 30 (P=0.016). Recurrent rejection, at 90 days after therapy, occurred less frequently with Thymoglobulin (17%) versus Atgam (36%) (P=0.011). A similar incidence of adverse events, post-therapy infections, and 1-year patient and graft survival rates were observed with both treatments. CONCLUSIONS: Thymoglobulin was found to be superior to Atgam in reversing acute rejection and preventing recurrent rejection after therapy in renal transplant recipients.
Acute Disease↗
Incidence of new-onset hypercholesterolemia in renal transplant patients treated with FK506 or cyclosporine.
In this study, we compare cholesterol levels during the first year after renal transplantation in FK506 (Prograf)- and cyclosporine-treated patients matched for cumulative first-year steroid dose and hypercholesterolemia risk factors. All patients had pretransplant cholesterol levels < 200 mg/dl. At 3 months posttransplant, 68% of the cyclosporine-treated patients had at least one cholesterol level greater than 200 mg/dl compared with 30% of the FK506-treated patients (P < 0.05). At the end of the year, 26% of FK506- and 67% of cyclosporine-treated patients remained hypercholesterolemic (P < 0.05). We conclude that cyclosporine has inherently more effect on cholesterol levels than FK506 during the first year after kidney transplantation.
Adult↗
In vitro study of BCX-34: a new human T-lymphocyte-specific purine phosphorylase inhibitor.
Explore the source record for details and available documents.
Animals↗