Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Precursor limitation”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 415 records · Page 23Linked to original sources
PubMed1989

Studies of the mechanism of tolerance induced by short-term immunosuppression with cyclosporine in high-risk corneal allograft recipients. I. Analysis of CTL precursor frequencies.

Transplantation of unmatched allogeneic corneas into highly vascularized recipient eyes under the cover of short-term immunosuppression with cyclosporine enables permanent engraftment. The aim of this study was to further elucidate the mechanism(s) underlying this tolerant state. In eight "high-risk" cornea recipients the clone sizes of donor-specific and third-party reactive cytotoxic T cell precursors were assessed by limiting dilution analyses before and at three and six months after transplantation. Acquired allograft tolerance in these patients was not accompanied by clonal reduction of donor-specific CTL-p, whereas in the case of an irreversible rejection the donor-specific CTL pool size was significantly enlarged. This donor-specific CTL-p increase could already be seen two months before clinical manifestation. These patterns differed from that of tolerant renal transplant patients, in whom marked and donor-specific reduction of CTL-p was observed. During rejection identical patterns with increasing donor-specific CTL-p frequencies were seen in both groups of patients. We conclude that induction of tolerance by short-term CsA to unmatched cornea grafts is not caused by clonal reduction of the effector precursor cell pool.

Adult↗
PubMed1990

Evidence that Leydig precursors localize in immature band two cells isolated on Percoll gradients.

The present studies examined responses to hCG and/or insulin of 3 beta-hydroxy-5-ene-steroid dehydrogenase and steroid 5----4-ene-isomerase activity (3 beta-HSD) in cultured Band 2 and Band 3 cells from 25- to 40-day-old rats isolated on Percoll gradients. In Band 2 cells, from 25-day-old rats enzyme activity increased about 3- and 2.5-fold, after 6 days of exposure to hCG or insulin, respectively. However, hCG did not stimulate enzyme activity in Band 2 cells from 30-, 35- and 40-day-old animals, and responses to insulin alone or insulin plus hCG declined with age. In Band 3 cells only insulin increased enzyme activity at each age. Neither hCG or insulin altered DNA levels in Band 2 or Band 3 cells, suggesting that increased activity in Band 2 cells from 25-day-old rats was not due to cellular replication. However, hCG increased the number of cells staining positive for 3 beta-HSD about 4-fold in Band 2 cells from 25-day-old rats. Insulin did not increase the number of positive staining cells in Band 2 and Band 3 cells from 25-day-old rats, suggesting that its major effect was to increase enzyme activity in existing cells. These results suggest that during a limited period of maturation precursor cells in Band 2, which are undetected by histochemical staining for 3 beta-HSD, can be converted to Leydig cells in culture by hCG.

Animals↗
PubMed1979

Anaerobic metabolism of immediate methane precursors in Lake Mendota.

Lake Mendota sediments and the immediate overlying water column were studied to better understand the metabolism of the methanogenic precursors H2/CO2 and acetate in nature. The pool size of acetate (3.5 microns M) was very small, and the acetate turnover time (0.22h) was very rapid. The dissolved inorganic carbon pool was shown to be large (6.4 to 8.3 mM), and the turnover time was slow (111 H.). CO2 was shown to account for 41 +/- 5.5% of the methane produced in sediment. Acetate and H2/CO2 were simultaneously converted to CH4. The addition of H2 to sediments resulted in an increase specific activity of CH4 from H(14)CO3- and a decrease in specific activity of CH4 from [2-14C]acetate. Acetate addition resulted in a decrease in specific activity of CH4 from H(14)CO3-. The metabolism of H(14)CO3- or [2-14C]acetate to 14CH4 was not inhibited by addition of acetate or H2. After greater than 99% of added [2-14C]acetate had been turned over, 42% of the label was recovered as 14CH4 20% was recovered as 14CO2 and 38% was incorporated into sediment. Inhibitor studies of [2-14C]acetate metabolism in sediments demonstrated that CHCl3 completely inhibited CH4 formation, but not CO2 production. Air and nitrate addition inhibited CH4 formation and stimulated CO2 production, whereas fluoroacetate addition totally inhibited acetate metabolism. The oxidation of [2-14C]acetate to 14CO2 was shown to decrease with time when sediment was incubated before the addition of label, suggesting depletion of low levels of an endogenous sediment electron acceptor. Acetate metabolism varied seasonally and was related to the concentration of sulfate in the lake and interstitial water. Methanogenesis occurred in the sediment and in the water immediately overlying the sediment during period of lake stratification and several centimeters below the sediment-water interface during lake turnovers. These data indicate that methanogenesis in Lake Mendota sediments was limited by "immediate" methane precursor availability (i.e., acetate and H2), by competition for these substrates by nonmethanogens, and by seasonal variations which altered sediment and water chemistry.

Acetates↗
PubMed1989

Vitamin K-dependent carboxylase: influence of the "propeptide" region on enzyme activity.

The liver microsomal vitamin K-dependent carboxylase catalyzes the post-translational conversion of specific glutamyl to gamma-carboxyglutamyl (Gla) residues in precursor forms of a limited number of proteins. These proteins contain an amino-terminal extension (propeptide) that is presumed to serve as an enzyme recognition site to assure their normal processing. The free, noncovalently bound propeptide has also been shown to stimulate the in vitro activity of this enzyme. This peptide has now been shown to lower the app Km of a low-molecular-weight Glu site substrate while having no influence on the app Km of the other substrates, vitamin KH2, O2, and CO2/HCO3-. Propeptide addition was shown to have no influence on the ratio of the two products of the enzyme, Gla and vitamin K-2,3-epoxide. Stimulation of carboxylase activity by the propeptide from human factor X was observed in a number of rat tissues and in the liver of a number of different species. Stability of the enzyme in crude microsomal preparations was greatly enhanced by the presence of propeptide. These observations are consistent with the hypothesis that this region of the protein substrates for the carboxylase not only serves an enzyme recognition or docking function but also modulates the activity of the enzyme by altering the affinity for one of its substrates.

Amino Acid Sequence↗
PubMed1990

Production of gastrin-releasing peptide-(18-27) and a stable fragment of its precursor in small cell lung carcinoma cells.

The production and postsecretory stability of gastrin-releasing peptide (GRP) and the C-terminal part of the GRP precursor were studied in small cell lung cancer cell lines using RIAs developed against these two parts of the precursor. In three otherwise different cell lines (NIC-H345, NIC-H69, and NIC-H510), similar chromatographic patterns of mainly GRP-(18-27) and some GRP-(14-27) along with large fragments of the C-terminal counterpart of the precursor were found to be stored in the cells. In tissue culture medium, gel filtration chromatography indicated that postsecretory limited proteolysis of the GRP precursor fragments occurred. The amount of accumulated immunoreactivity varied among the three cell lines and between the two parts of the precursor. In medium in which only low amounts of GRP immunoreactivity accumulated, the radiolabeled GRP was degraded rapidly. When incubated with plasma, GRP-(14-27) disappeared within a few hours, whereas the C-terminal precursor fragments were stable. It is concluded that the postsecretory stability of peptides excised from the GRP precursor in small cell lung cancer cells varies under tissue culture conditions, but epitopes in the C-terminal part of the precursor are more stable in plasma than the small GRP peptides and, thus, may serve as a better indicator than GRP itself for expression of the GRP precursor in cancer cells.

Amino Acid Sequence↗
PubMed2002

Complexities in ETS-domain transcription factor function and regulation: lessons from the TCF (ternary complex factor) subfamily. The Colworth Medal Lecture.

The ETS-domain transcription factor family can be divided into a series of subfamilies. Elk-1 represents the founding member of the ternary complex factor (TCF) subfamily. By focusing on the TCF subfamily, we can demonstrate the complexities that exist in the function and regulation of ETS-domain transcription factors. This article focuses on Elk-1 in detail and summarizes the functions of other TCFs. The key themes covered include the domain structure of the TCFs, the mechanisms of complex formation with serum response factor, regulation of TCFs by mitogen-activated protein kinase cascades, and transcriptional regulatory properties of the TCFs. Finally, the emerging role of the TCFs in vivo is discussed. A picture is developing indicating that, while these proteins exhibit significant sequence and functional conservation, key differences in their structure and regulation are being identified which may relate to unique functions of these proteins in vivo.

Amino Acid Sequence↗
PubMed1999

Subtilisin-like proprotein convertase PACE4 (SPC4) is a candidate processing enzyme of bone morphogenetic proteins during tooth formation.

The temporospatial expression of PACE4, a member of the mammalian subtilisin-like proprotein convertase family, in the developing rat molar tooth was determined by in situ hybridization. At the initiation stage of tooth development, PACE4 mRNA was weakly expressed in the dental lamina, whereas the mesenchymal cells intensely expressed the PACE4 transcript. At the bud stage, high-level expression of PACE4 mRNA was found in the dental epithelium and condensed dental mesenchyme. Its expression became more localized in the differentiating ameloblasts during cap and early bell stages. In the newborn rats, PACE4 mRNA was localized in the ameloblasts and odontoblasts, but its expression became weaker with advancing development, showing apparent association with the differentiation and establishment of functional ameloblasts and odontoblasts. These expression patterns of PACE4 were very similar to those of several bone morphogenetic proteins (BMPs) reported previously. Because BMPs, which are primarily involved in the morphogenesis in tooth formation, are synthesized as inactive precursors and activated by limited proteolysis at the consensus Arg-X-X-Arg maturation site, the present observations suggest that PACE4 is possibly a candidate proBMP convertase that acts during tooth formation.

Ameloblasts↗
PubMed1998

Development of T cell precursor activity in the murine fetal liver.

The generation of T cell precursors in the liver of murine embryos was studied. The total number of T cell precursors in the liver was measured in thymic organ cultures by a limiting dilution assay. Sixty T cell precursors were detected in the liver at day 11 of gestation. By day 12 the number of precursors showed a 20-fold increase, half of which could be explained by in situ proliferation as ascertained by a fetal liver organ culture assay. By day 13 a further 2-3-fold increase was observed. Whereas the number of total liver cells continued to increase, that of T cell precursors declined in the following days, suggesting a massive exit of these cells after day 13. The capacity to generate a TCRB repertoire in the cells was evaluated by a PCR assay. T cell precursors in day 11 fetal liver developed a TCRB repertoire at day 8 of culture. The cells from days 12-15 developed an identically diverse repertoire by day 6, suggesting that day 11 precursors are more immature than those of later days. A mechanism for yielding a single wave of T cell precursors in the fetal liver is discussed with a proposed model.

Animals↗
PubMedDate not supplied

Genomic channeling in bacterial cell division.

The bacterial dcw cluster is a group of genes involved in cell division and peptidoglycan synthesis. Comparison of the cluster across several bacterial genomes shows that its gene content and its gene order are conserved in distant bacterial lineages and, moreover, that, being most conserved in rod-shaped bacteria, the degree of conservation relates to bacterial morphology. We propose a model in which the selective pressure to maintain the cluster arises from the need to efficiently coordinate the processes of elongation and septation in rod-shaped bacteria. Gene order in the dcw cluster would be conserved as a result of mechanisms comprising: (i) a limited amount of peptidoglycan precursors required both for septation and elongation of the wall; (ii) co-translational assembly of the protein complexes involved in cell division and in the synthesis of the peptidoglycan precursors; and (iii) alternation in the cellular localization of the assembled complexes to participate either in the synthesis of the septal peptidoglycan and division, or in the synthesis of the lateral wall. The name genomic channeling is proposed for this model as it involves a genomic arrangement that could facilitate the assembly of specific protein complexes and their subsequent conveyance to specific locations in the crowded cytoplasm and the envelope.

Bacteria↗
PubMed2005

Testing the role of solvent surface tension in protein ionization by electrospray.

This work was aimed at probing the influence of solvent surface tension on protein ionization by electrospray. In particular, we were interested in testing the previously suggested hypothesis that the charge-state distributions (CSDs) of proteins in electrospray ionization mass spectrometry (ESI-MS) are controlled by the surface tension of the least volatile solvent component. In the attempt to minimize uncontrolled conformational effects, we used acid-sensitive proteins (cytochrome c and myoglobin) at low pH or highly stable proteins (ubiquitin and lysozyme) in the presence of low concentrations of organic solvents. A first set of experiments compared the effect of 1- and 2-propanol. These two alcohols have similar chemico-physical properties but values of vapor pressure below and above that of water, respectively. Both compounds have much lower surface tension than water. The solvents employed allowed testing of the influence of surface tension on protein spectra obtained from similarly denaturing solutions. The compared solvent conditions gave rise to very similar spectra for each tested protein. We then investigated the effect of the addition of dimethyl sulfoxide to acid-unfolded proteins. We observed enhanced ionization in the presence of acetic or formic acid, consistent with the previously described supercharging effect, but almost no shift of the CSD in the presence of HCl. Finally, we analyzed thermally denatured cytochrome c, to obtain reference spectra of the unfolded protein in high-surface-tension solutions. Also in this case, the CSD of the unfolded protein was shifted towards lower m/z values relative to low-surface-tension systems. In contrast to the other results reported here, this effect is consistent with an influence of solvent surface tension on CSD. The magnitude of the effect, however, is much smaller than predicted by the Rayleigh equation. The results presented here are not easy to reconcile with the hypothesis that the maximum charge state exhibited by proteins in ESI-MS reflects the Rayleigh-limit charge of the precursor droplet. The data are discussed with reference to models for the mechanism of electrospray ionization.

Ions↗
PubMed1997

Evidence for involvement of NF-kappaB in the transcriptional control of COX-2 gene expression by IL-1beta.

The cyclooxygenase (COX) isoforms COX-1 and COX-2 convert arachidonic acid to prostaglandin (PG) precursors and are a limiting step in PG production. Interleukin-1beta (IL-1beta) treatment of type II A549 cells increases PGE2 synthesis via transcription- and translation-dependent induction of COX-2. IL-1beta produces a 10-fold induction of COX-2 mRNA and an 8-fold increase in COX-2 transcription that was temporally preceded by activation of the transcription factor nuclear factor-kappaB (NF-kappaB). The protein-tyrosine phosphatase inhibitor phenylarsine oxide (PAO) prevented both NF-kappaB activation and induction of COX-2 mRNA. We show that two putative NF-kappaB motifs, kappaBu (-447/-438) and kappaBd (-224/-214), from the COX-2 promoter bind p50/p65 NF-kappaB heterodimers in an IL-1beta-dependent manner and that the upstream element has the greater affinity. Finally, we demonstrate that the two NF-kappaB subunits, p50 and p65, synergistically activate a -917/+49 COX-2 promoter construct. We conclude that IL-1beta stimulates PG production via transcriptional activation of COX-2 and provide evidence that this may involve NF-kappaB.

Arsenicals↗
PubMed1999

A bacterial signal peptidase enhances processing of a recombinant single chain antibody fragment in insect cells.

The production of an antibody single chain fragment (scFv) in insect cells was accompanied by the formation of an insoluble intracellular precursor even with the inclusion of the bee melittin signal peptide. The presence of the precursor polypeptide suggests a limitation in the processing of the signal peptide so a baculovirus containing a signal peptidase from Bacillus subtilis (SipS) was constructed for expression studies. When the wild type SipS was coexpressed with scFv, preprocessed scFv fragments were no longer detected in insect cell lysates. Conversely, coexpression of scFv alone or with an inactive mutant SipS resulted in at least 30% of the intracellular polypeptide in an unprocessed form at 3 days post infection. Production of scFv in the medium was also enhanced in the presence of SipS; however, low secretion levels indicate the presence of a post-processing bottleneck.

Animals↗
PubMed1995

Effects of ultraviolet-B irradiation on human LAK and NK cytotoxic activity.

In vitro ultraviolet-B (UVB) irradiation of murine and rodent bone marrow cells prevents GVHD without compromising engraftment while inducing tolerance to donor-type allografts. In anticipation of clinical trials of UVB-modified bone marrow grafts, we studied the in vitro effects of UVB irradiation (50-300 J/m2) on human natural killer and lymphokine activated killer cells since both types of cells influence the development of GVHD and graft-versus-tumor effect. Interleukin-2-activated and untreated human lymphocytes were used as effectors in a 51Cr release cytotoxic assay against various tumor cell lines as targets. NK-mediated lysis of K562 targets was decreased by UVB irradiation of the effector cells in a dose-dependent manner. FACS analysis of CD16+ and CD56+ cells 24 hr after UVB exposure showed a UVB-dose-dependent decrease in the number of cells expressing these surface markers. UVB irradiation of lymphocytes prior to activation with high-dose IL-2 resulted in a range of 20- to 89-fold decrease in LAK precursors as measured by limiting dilution analysis using the LAK-sensitive cell line HL60. In contrast, the LAK activity of lymphocytes that had been stimulated in vitro with high-dose IL-2 prior to UVB irradiation was preserved when assayed immediately after UVB modulation; however, there was a significant decrease in lytic activity (with most samples tested) when the assay was performed 24 hr after UVB exposure. It appears that the lymphocyte response to UVB modification is dose dependent, with some cell types displaying higher sensitivity to UVB irradiation than others. These findings suggest that prevention of GVHD by UVB is due, in part, to inhibition of NK activity, and may offer a new strategy to augment the graft versus leukemia effect of UVB-modified bone marrow grafts in clinical transplantation.

Bone Marrow Transplantation↗
PubMed1996

Metabolic fluxes regulate the success of sporulation in Saccharomyces cerevisiae.

In this work we investigated to what extent cellular metabolism and energetics regulate sporulation in Saccharomyces cerevisiae and which metabolic pathways are involved in such regulation. Sporulation, meiosis, and associated metabolic fluxes in S. cerevisiae strain CH1211 were studied in several experimental protocols involving changes of carbon source (acetate, lactate, or pyruvate) or cell density in sporulation medium, or changing the phase of batch growth at which cells were harvested before transfer to sporulation medium. In acetate-based sporulation medium, the rate at which cells utilized glyoxylate and gluconeogenic pathways correlated positively with the percentage of asci per cell at 72 h. In contrast, in lactate sporulation medium the frequency of sporulation correlated negatively with both the rate of lactate consumption and the fluxes through gluconeogenesis and the pyruvate-carboxylase catalyzed step. In the presence of lactate, the respiratory capacity did correlate positively with the percentage of asci per cell. The experimental data suggest that acetate limits fluxes to anabolic precursors during sporulation. In contrast, sporulation on lactate appears to be influenced by catabolic processes or, even more precisely, by the respiratory capacity of yeast cells. The results obtained are discussed in terms of the hypothesis that an imbalance between anabolic and catabolic fluxes may be required for an efficient sporulation.

Acetates↗
PubMed1993

Growth inhibition of ovarian cancer cells induced by antisense IL-6 oligonucleotides.

In previous work, we saw that interleukin-6 (IL-6), a multifunctional cytokine, is produced by epithelial ovarian cancer cells and that ovarian cancer cells express the IL-6 receptor. Here, we examined the possibility that IL-6 acts as an autocrine growth factor for ovarian cancer cells. Inhibition of IL-6 gene expression by exposure to IL-6 antisense oligonucleotides resulted in greatly decreased cellular proliferation. Exposure of ovarian cancer cell lines (CAOV-3, OVCAR-3, and OC-436), to 1-5 microM of a 15-base single-stranded antisense IL-6 oligodeoxynucleotide, specific for a sequence in the second coding exon of the IL-6 gene, resulted in decreased IL-6 production and a > 80-85% inhibition of cellular proliferation. However, the addition of exogenous IL-6 failed to restore the proliferation of the antisense-treated cells. Antibodies to IL-6 did not consistently inhibit cell growth nor did rIL-6 enhance precursor frequency in a limiting dilution analysis. These results suggest that IL-6 does not directly induce the proliferation of ovarian cancer cells although endogenous IL-6 production is needed for optimal cell growth. As the majority of epithelial ovarian cancers produce IL-6, the direct specific inhibition of IL-6 gene expression is of potential therapeutic value.

Base Sequence↗
PubMed1995

Cytotoxic T lymphocyte escape variants, induced mutations, and synthetic peptides define a dominant H-2Kb-restricted determinant in simian virus 40 tumor antigen.

Immunization of C57BL/6 mice with syngeneic cells transformed by simian virus 40 large T antigen (SV40 T ag) induces the generation of T antigen-specific cytotoxic T lymphocytes (CTL) which are restricted by the major histocompatibility class I antigens H-2Db and H-2Kb. Previous studies have shown that the H-2Db-restricted CTL response is directed to at least three distinct epitopes (I, II/III, and V) in the SV40 T antigen which have been precisely mapped using deletion mutagenesis and overlapping synthetic peptides. Although in vivo the CTL response to SV40 T antigen is dominated by the H-2Kb class I antigen, the precise location of the H-2Kb-restricted epitope(s) was not known, and whether there was multiplicity of H-2Kb-restricted epitopes remained unclear. In this study, we have defined the minimal recognition epitope for the SV40-specific H-2Kb-restricted CTL clone Y-4 as T antigen residues 404-411 by using T antigen deletion and point mutants and synthetic peptides. DNA sequence analysis of the region encoding residues 404-411 from the T antigens expressed in three independently isolated CTL clone Y-4 escape variants identified inactivating mutations capable of abrogating CTL recognition. Estimation of CTL precursor (CTLp) frequencies by limiting dilution analysis revealed that CTLp specific for epitope IV represent a large percentage of the total CTL response elicited by the intact T antigen in H-2b mice. Immunization of B6 mice with cells expressing a T antigen derivative deleted of residues 404-411 revealed that site IV represents the only immunodominant H-2Kb-restricted epitope within T antigen.

Animals↗