Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.31 (beta-glucuronidase)
7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The abundant multifunctional protein ABF1 of Saccharomyces cerevisiae binds to the upstream region of several genes, including some ribosomal protein genes like the one encoding protein S33. Deletion of the ABF1-binding sequence lowers the transcription of these genes three- to more than ten-fold. We have isolated the S33 genes of two related yeast species, Kluyveromyces lactis and Kluyveromyces marxianus. Comparison of the nucleotide sequences of these S33 genes with their counterpart from S. cerevisiae shows a strong sequence similarity covering the whole of the coding regions. In contrast, little or no sequence similarity is found in the 5'-flanking regions of the three genes. Also the trailer regions differ considerably in both length and sequence from one species to another. An ABF1-binding site is present in the upstream region of the S33 gene of K. marxianus. Retardation analyses showed that this sequence is able to bind a protein present in Kluyveromyces cells with a molecular mass somewhat lower than that of S. cerevisiae ABF1. Functional analyses, using a beta-glucuronidase reporter system, showed that the ABF1-binding site is indeed involved in transcription activation of the K. marxianus S33 gene in Kluyveromyces cells. A S. cerevisiae ABF1-gene-specific probe showed only weak hybridization with Kluyveromyces DNA and Northern blots did not show a signal. These results indicate that S. cerevisiae and Kluyveromyces contain functionally related but structurally dissimilar ABF1-type proteins.
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PMID:Structure and expression of the ABF1-regulated ribosomal protein S33 gene in Kluyveromyces. 148 71

In order to study the regulation of nuclear genes coding for plastid ribosomal proteins, we have analysed the promoter region of spinach rps22 using both in vitro and in vivo approaches. By footprinting analyses, we have identified eight DNA elements interacting with spinach leaf nuclear factors in the 300 bp promoter region upstream of the transcription start site. Among these elements, four are short AT-rich sequences and one is identical to the Hex motif characterized initially in wheat histone genes. In transgenic tobacco plants, the reporter gene coding for the beta-glucuronidase (GUS) directed by a 1.2 kb upstream region of rps22 was expressed in several plant organs, with high levels in leaf mesophyll, embryo cotyledons and root meristematic cells and very low levels in other cell types. Interestingly, when deleted to -295, the promoter, which contained all the foot-printed elements, was still able to confer the same expression pattern, although the activity was relatively lower than with the 1.2 kb promoter. When deleted further to -154, the promoter, from which the AT-rich elements were eliminated, loses its activity almost completely, suggesting that these AT-rich elements are important for the rps22 promoter activity. Altogether, our results show that rps22 gene expression is controlled by specific cis elements not present in other nuclear-encoded plastid ribosomal protein genes studied so far.
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PMID:Cis-acting elements and expression pattern of the spinach rps22 gene coding for a plastid-specific ribosomal protein. 764 93

Lateral roots can be synchronously induced in Arabidopsis by a brief auxin treatment. An early event in the development of a lateral root primordium is the accumulation of mRNAs encoding ribosomal proteins. In situ hybridizations show that mRNA encoding one ribosomal protein, L16, accumulates in all rapidly proliferating tissues including the shoot and root apical meristems and lateral root primordia. To understand further the mechanisms by which ribosomal proteins are coordinately synthesized, two genes encoding the ribosomal protein L16 were isolated from Arabidopsis thaliana. Promoter sequences from each RPL16A and RPL16B were fused to the beta-glucuronidase reporter gene GUS. The promoter of RPL16B(from -848 to -19) conferred X-Gluc staining in proliferating tissues including the shoot and root apical meristems. When GUS was expressed from the RPL16A promoter (from -875 to -22), X-Gluc staining was observed in cells in the root stele and in anthers. When seedlings transformed with either promoter construct were treated with auxin to induce lateral roots, X-Gluc staining accumulated in the lateral root primordia by 16 h after induction. Transcription of the RPL16B promoter appears to be correlated with cell division, while transcription of the RPL16A promoter is very cell specific. Expression of two genes encoding L16 during the early phase of lateral root initiation and in developing pollen may serve to increase levels of ribosomal proteins during the rapid growth of these tissues.
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PMID:Developmental regulation of ribosomal protein L16 genes in Arabidopsis thaliana. 765 8

All ribosomal protein (rp) gene promoters from Saccharomyces cerevisiae studied so far contain either (usually two) binding sites for the global gene regulator Rap1p or one binding site for another global factor, Abf1p. Previous analysis of the rpS33 and rpL45 gene promoters suggested that apart from the Abf1 binding site, additional cis-acting elements play a part in transcription activation of these genes. We designed a promoter reconstruction system based on the beta-glucuronidase reporter gene to examine the role of the Abf1 binding site and other putative cis-acting elements in promoting transcription. An isolated Abf1 binding site turned out to be a weak activating element. A T-rich sequence derived from the rpS33 proximal promoter was found to be stronger, but full transcription activation was only achieved by a combination of these elements. Both in the natural rpL45 promoter and in the reconstituted promoter, a Rap1 binding site could functionally replace the Abf1 binding site. Characteristic rp gene nutritional control of transcription, evoked by a carbon source upshift or by nitrogen re-feeding to nitrogen starved cells, could only be mediated by the combined Abf1 (or Rap1) binding site and T-rich element and not by the individual elements. These results demonstrate that Abf1p and Rap1p do not activate rp genes in a prototypical fashion, but rather may serve to potentiate transcription activation through the T-rich element.
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PMID:Transcription activation of yeast ribosomal protein genes requires additional elements apart from binding sites for Abf1p or Rap1p. 778 99

A genomic clone encoding the potato homolog of the yeast ubiquitin-ribosomal protein fusion gene ubi3 was isolated and characterized. Chimeric genes containing the ubi3 promoter (920 bp of 5' to the ubiquitin start codon) were constructed in which the reporter gene beta-glucuronidase (GUS) was either fused directly to the promoter, or introduced as a translational fusion to the ubiquitin-coding region. After introduction into the potato by Agrobacterium-mediated transformation, GUS activities were measured in leaves and in tubers of transgenic clones. GUS activity was 5- to 10-fold higher in clones expressing the ubiquitin-GUS translational fusion than in clones containing GUS fused directly to the ubi3 promoter. For both types of constructs, GUS activity was highest in meristematic leaves and declined during leaf expansion, then rose again to near the meristematic levels during senescence. GUS activity in tubers was similar to that in young leaves. In contrast to the native ubi3 genes, the chimeric ubi3-GUS transgenes were not activated in the tuber by wounding.
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PMID:Isolation of a ubiquitin-ribosomal protein gene (ubi3) from potato and expression of its promoter in transgenic plants. 811 Oct 11

The rpL34 gene, which encodes a cytoplasmic ribosomal protein with a high homology to the rat 60S r-protein L34, was isolated from a genomic library of tobacco (Nicotiana tabacum L. cv. Xanthi-nc). A 1500 bp upstream promoter fragment was fused to the chloramphenicol acetyltransferase (CAT) reporter gene or beta-glucuronidase (GUS) reporter gene and transferred into tobacco plants by the Agrobhacterium-mediated leaf disk transformation method. Analysis of CAT activity in leaf tissues showed that mechanical wounding increased the rpL34 promoter activity about 5 times as compared to untreated controls and that the promoter activity was further enhanced by plant growth regulators, 2,4-dichlorophenoxyacetic acid and benzyladenine. Histochemical GUS staining patterns of the transgenic plants showed that the rpL34 promoter activity is high in actively growing tissues, including various meristems, floral organs, and developing fruits. A series of 5' deletion analyses of the rpL34 promoter indicated that a 50 bp region located between -179 and -129 is essential for wound, auxin and cytokinin responses. Deletion of this region reduced the promoter activity to an undetectable level. Insertion of the 50 nucleotide sequence into a minimal promoter restored the promoter activity and the promoter strength was proportional to the copy number of the upstream sequence. The role of TATA and CAAT box regions was studied by a series of 3' deletion analyses. A 3' deletion up to -28 did not significantly affect the promoter strength. However deletion of the promoter up to 70 bp, which deleted the TATA box region, significantly reduced promoter activity. Further deletion of the promoter up to - 104. eliminating the CAAT box region, abolished the promoter activity. These results suggest that the TATA box and CAAT box regions are also important for the rpL34 promoter activity in addition to the 50 bp upstream region.
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PMID:Promoter elements controlling developmental and environmental regulation of a tobacco ribosomal protein gene L34. 900 4

Due to constraints in vector construction, reporter polypeptides often carry N-terminal sequences of extraneous origin. Since protein half-life can be influenced by small determinants in the N-terminus, such foreign sequences can destabilize proteins and compromise results of reporter-based studies. We provide a real-life example of this problem (destabilizing sequences derived from a ribosomal protein) and show that it can be solved with the ubiquitin fusion technique, in which ubiquitin sequences are placed upstream of the reporter, in our case beta-glucuronidase. Post-translational processing by characterized pathways removes the ubiquitin together with destabilizing sequences, generating a stable reporter whose N-terminus is constant.
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PMID:UGUS, a reporter for use with destabilizing N-termini. 946 79

Quantitative gene expression protocols require adequate controls to monitor intersample variation. Quantitative approaches to describe relative changes in gene expression use endogenous controls--"housekeeping" genes. Given the low amounts of mRNA in fat cells, RT-PCR is the method of choice, and housekeeping genes are widely used as endogenous controls. However, literature reports suggest changes in gene expression of typical housekeeping genes (e. g. GAPDH, beta-actin, 18S rRNA) upon hormonal stimulation or during adipogenic differentiation. Thus, we tested the influence of 6 hormones and adipogenic differentiation on gene expression levels of 11 commonly used housekeeping genes in primary cultured mature human adipocytes and preadipocytes. Using the TaqMan RT-PCR technique and "Human Endogenous Control Assays" (PE Biosystems), we found several housekeeping genes with at least twice the difference in expression levels between stimulated and unstimulated cells (such as acidic ribosomal protein, beta-actin, beta(2)-microglobulin and beta-glucuronidase). Only GAPDH and transferrin receptor gene expression levels did not change under any of the stimuli tested, thus appeared best suited for gene expression studies in human adipose cells across a wide range of experimental settings.
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PMID:Validation of endogenous controls for gene expression studies in human adipocytes and preadipocytes. 1160 84

For interpretation of quantitative gene expression measurements in clinical tumor samples, a normalizer is necessary to correct expression data for differences in cellular input, RNA quality, and RT efficiency between samples. In many studies, a single housekeeping gene is used for normalization. However, no unequivocal single reference gene (with proven invariable expression between cells) has been identified yet. As the best alternative, the mean expression of multiple housekeeping genes can be used for normalization. In this study, no attempt was made to determine the gold-standard gene for normalization, but to identify the best single housekeeping gene that could accurately replace the measurement of multiple genes. Expression patterns of 13 frequently used housekeeping genes were determined in 80 normal and tumor samples from colorectal, breast, prostate, skin, and bladder tissues with real-time quantitative RT-PCR. These genes included, large ribosomal protein, beta-actin, cyclophilin A, glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerokinase 1, beta-2-microglobin, beta-glucuronidase, hypoxanthine ribosyltransferase (HPRT), TATA-box-binding protein, transferrin receptor, porphobilinogen deaminase, ATP synthase 6, and 18S ribosomal RNA. Principal component analysis was used to analyze these expression patterns, independent of the level of expression. Our approach identified HPRT as the single best reference gene that could be used as an accurate and economic alternative for the measurement of multiple housekeeping genes. We recommend this gene for future studies to standardize gene expression measurements in cancer research and tumor diagnostics until a definite gold standard has been determined.
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PMID:Normalization of gene expression measurements in tumor tissues: comparison of 13 endogenous control genes. 1554 3

Real-time reverse transcription PCR has greatly improved the ease and sensitivity of quantitative gene expression studies. However, measurement of gene expression generally requires selection of a valid reference (housekeeping gene) for data normalization to compensate for inherent variations. Given the dynamic nature of early embryonic development, application of this technology to studies of oocyte and early embryonic development is further complicated due to limited amounts of starting material and a paucity of information on constitutively expressed genes for data normalization. We have validated quantitative procedures for real-time reverse transcription polymerase chain reaction (RT-PCR) analysis of mRNA abundance during bovine meiotic maturation and early embryogenesis and utilized this technology to determine temporal changes in mRNA abundance for ribosomal protein L-15, cyclophilin-A, phosphoglycerokinase, beta-glucuronidase, glyceraldehyde-3-phosphate dehydrogenase, beta-actin, and histone H2A. Quantification of amounts of specific exogenous RNAs added to samples revealed acceptable rates of RNA recovery and efficiency of reverse transcription with minimal variation. Progression of bovine oocytes to metaphase II resulted in reduced abundance of polyadenylated, but not total transcripts for majority of above genes; however phosphoglycerokinase exhibited a significant decline in both RNA populations. Abundance of mRNAs for above genes in early embryos generally remained low until the blastocyst stage, but abundance of ribosomal protein L-15 mRNA was increased at the morula stage and histone H2A mRNA showed dynamic changes prior to embryonic genome activation. Results demonstrate a valid approach for quantitative analysis of mRNA abundance in oocytes and embryos, but do not support constitutive expression of above genes during early embryonic development.
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PMID:Quantitative analysis of messenger RNA abundance for ribosomal protein L-15, cyclophilin-A, phosphoglycerokinase, beta-glucuronidase, glyceraldehyde 3-phosphate dehydrogenase, beta-actin, and histone H2A during bovine oocyte maturation and early embryogenesis in vitro. 1626 7


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