Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The present investigations on rat lung show that metabolic changes occurring around the 20th gestational day are accompanied by multiple alterations in the quantitative pattern of enzymes. This involves increases in two lysosomal enzymes (N-acetyl beta-glucosaminidase and beta-galactosidase) and a rise and fall in pyruvate kinase and alpha-glucosidase. The striking transient upsurge of adenylate kinase, however, is postponed until after birth. The normal diminution of thymidine kinase and peptidylproline hydroxylase is drastically enhanced by an injection of cortisol to fetal rats. Studies on human pulmonary tissues consisted in determining enzyme concentration from the ninth to the 21st week of gestation and an histologically normal adult lungs. The results show that the 15th to the 21st week of gestation is the period of increase in pyruvate kinase, adenylate kinase and alpha-glucosidase. The rise during the development of several enzymes (e.g., 5'-nucleotidase, alkaline phosphatase, and gamma-glutamyl transpeptidase) and the decline in thymidine kinase and peptidylproline hydroxylase, however, dose not begin until after the 21st week of gestation.
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PMID:Phosphotransferases and lysosomal enzymes in fetal human and rat lung. 626 41

We have developed an Escherichia coli plasmid vector for the identification and expression of foreign DNA segments that are open reading frames (ORFs). The 5' end of ompF, an E. coli gene encoding an abundant outer membrane protein, is used to provide a strong, regulated promoter, translation initiation site, and signal sequence for export from the cytoplasm. This sequence is coupled to the lacZ gene of E. coli so that expression of beta-galactosidase requires ompF transcription and translation signals. However, this hybrid gene is LacZ- because lacZ is out of frame with respect to ompF. Restriction enzyme recognition sites are located between ompF and lacZ to allow convenient insertion of DNA fragments. If an insert is an ORF of the correct length, ompF and lacZ become realigned in frame, resulting in a LacZ+ gene that produces a tribrid protein with the translation product of the insert sandwiched between OmpF and beta-galactosidase. The LacZ+ phenotype thus identifies clones containing an expressed ORF. To demonstrate the vector's utility we inserted a fragment from the herpes virus thymidine kinase gene and used the resulting tribrid protein to raise antibodies that precipitate thymidine kinase from herpes virus-infected cells. We also inserted a fragment from the E. coli lexA gene to produce a tribrid protein that is precipitated by antiserum raised with LexA protein. Thus, tribrid fusion proteins can be used to produce or detect antibodies and also to identify the product of a cloned gene.
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PMID:Open reading frame expression vectors: a general method for antigen production in Escherichia coli using protein fusions to beta-galactosidase. 630 25

We have previously demonstrated that retrovirus-mediated genes were transferred to mouse glioma cells in a meningeal gliomatosis model (Yamada et al.: Japanese Journal of Cancer Research 83:1244-1247, 1992). This retrovirus vector contains the Escherichia coli. beta-galactosidase (beta-gal) gene as a marker for integration of the lacZ gene, which is controlled by the SV40 early promoter. We investigated whether lacZ genes could be specifically controlled in mouse glioma cells by glial-specific promoters, including the 2.5 kb 5' flanking region of the mouse glial fibrillary acidic protein (GFAP) gene, the 1.3 kb 5' flanking region of the myelin basic protein (MBP) gene, and the 1.5 kb 5' flanking region of the myelin proteolipid protein (PLP) gene. Psi-2 packaging cells were transfected with each retrovirus vector (GFAP promoter-, MBP promoter-, and PLP promoter-lacZ) and the infectious virus particles were recovered from the supernatants. Blue staining for beta-gal was detected in various fibroblast, myeloma, and glioma cell lines transduced with the retrovirus BAG vector. On the other hand, blue staining was only detected in glioma cells after transduction with the lacZ gene-bearing retrovirus controlled by glial-specific promoters. The strongest promoter activity was detected after transduction with the retrovirus in which the MBP promoter controlled the lacZ gene. Mouse glioma cells transduced with retrovirus containing the MBP promoter directing the herpes simplex virus type 1 thymidine kinase (HTK) gene were extremely sensitive to ganciclovir, while the parental cells and cells transduced with retrovirus containing the lacZ gene were not sensitive to ganciclovir.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Selective expression of foreign genes in glioma cells: use of the mouse myelin basic protein gene promoter to direct toxic gene expression. 750 43

The murine homeobox-containing gene Hoxa-7 is expressed in restricted patterns during embryogenesis and plays an important role in the control of region-specific differentiation. Previous studies have shown that separate elements specify lineage restriction and expression boundaries of Hoxa-7. In particular 3.6 kb of 5' flanking sequences were sufficient to establish an anterior boundary of Hoxa-7 gene expression. To identify the minimal regulatory element specifying the anterior boundary of expression, transgenic mice were generated carrying chimeric constructs with deletions of 5' flanking sequences fused to a thymidine kinase minimal promoter/E. coli lacZ reporter construct. By deletion analysis, a 470 bp long control element (AX 470) located 1.6 kb upstream of the transcription start site was identified that directed expression of the beta-galactosidase protein in a pattern reflecting the anterior boundary of expression of the endogenous Hoxa-7 gene. This element was active in either orientation and conferred region-specific expression to unrelated promoters, thereby behaving like an enhancer element. In contrast, transgenic mice carrying further 5' and 3' deletions of the 470 bp long element did not exhibit an anterior boundary of Hoxa-7 expression. Based on these results the minimal control element (AX 470) specifying the anterior boundary of Hox expression was designated as Hoxa-7 enhancer. Furthermore, 3 kb of the human HOXA7 upstream region were sequenced and compared to its mouse homologue in order to identify conserved regions. Sequence comparison revealed motifs that were strongly conserved between both species. The human homologue of the mouse Hoxa-7 enhancer was 70% identical at the nucleotide level and was also capable of directing an anterior boundary in transgenic mice. Using transgenic lines a detailed analysis of the Hoxa-7 enhancer-directed expression during embryogenesis was performed. lacZ expression was first detected in the allantois at day 7.5 p.c. and in mesoderm and ectoderm at day 8.5 of gestation. Between gestational ages E8.5 to E12.5 beta-gal expression was observed in the somites, spinal cord, spinal ganglia and paraxial mesoderm as well as in mesenchymal layers of the kidney. A distinct anterior limit of expression was noted in transgenic lines at level C4 (neural tube) and C5 (spinal ganglia). Our deletion experiments defined a minimal enhancer element specifying the anterior boundary of Hox gene expression in early and late phases of development. Further studies aim at characterizing the trans-acting factors that mediate the spatial and temporal expression of Hox genes in the developing embryo.
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PMID:A conserved enhancer of the human and murine Hoxa-7 gene specifies the anterior boundary of expression during embryonal development. 753 68

The Copper isolate of bovine herpesvirus 1 (BHV-1) was used to produce a thymidine kinase-negative (TK-) recombinant by insertion of a beta-galactosidase (bgal) expression cassette into the TK coding region. The recombinant virus (TK- bgal+) was tested for abortifacient activity in cattle by inoculation of 5 pregnant heifers at 25 to 29 weeks gestation. Five additional heifers were inoculated with the Cooper TK-positive (TK+) virus to serve as controls. After inoculation, both groups of heifers developed similar febrile responses and neutralizing antibody titers. Virus was isolated from blood of all heifers during the first postinoculation (PI) week, and isolation frequencies were similar for both groups. In contrast, whereas virus was isolated from many of the nasal and vaginal swab specimens of heifers inoculated with TK+ virus, only rare virus isolations were made from the heifers given TK- bgal+ virus. All heifers inoculated with TK+ virus aborted between PI days 19 and 35. The finding of characteristic microscopic lesions and viral antigen in fetal tissues indicated that the abortions were caused by BHV-1 infection. Virus was isolated from 3 fetuses, and all isolates were TK+ virus. Two heifers inoculated with TK- bgal+ virus aborted at PI days 25 and 39. Fetal tissues had typical BHV-1 microscopic lesions and viral antigen. Virus was isolated from blood of both fetuses, and the isolates were TK- bgal+. Results of this study indicate that inactivation of the TK gene reduces, but does not eliminate, the abortifacient activity of BHV-1.
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PMID:Abortion in heifers inoculated with a thymidine kinase-negative recombinant of bovine herpesvirus 1. 757 53

The use of intrathecal, retroviral-mediated transfer of the herpes simplex thymidine kinase (HStk) gene and subsequent ganciclovir (GCV) administration has recently been shown to improve survival in a rat model of leptomeningeal carcinomatosis. Clinical application of this approach is attractive because access to the cerebrospinal fluid (CSF) space is relatively noninvasive and distribution of producer cells and vectors may be facilitated by circulation of CSF, overcoming distribution problems inherent in solid tumors. However, meningeal inflammation, transduction and injury to normal CNS tissue, proliferation of the xenogeneic producer cells in the subarachnoid space, immune-mediated injury, and development of hydrocephalus are possible complications of intraventricular or intrathecal administration of vector-producer cells. In addition, the dynamics of producer cell and vector distribution in the CSF are unknown. To address these issues, we evaluated the safety of this approach for gene delivery and assessed the dynamics of distribution of producer cells and retroviral vectors in rats and non-human primates. In rats, transduction of normal central nervous system (CNS) structures surrounding the subarachnoid space was evaluated after intrathecal and intraventricular injections of beta-galactosidase and HStk vector-producer cells, with and without GCV. In primates, beta-galactosidase and HStk vector-producer cells were injected intraventricularly and GCV was administered either intrathecally or intravenously. Toxicity was evaluated by neurologic examination, serial gadolinium-enhanced MRI scans of the brain, and blood and CSF profiles. A subgroup of monkeys received repeated intraventricular injection of vector-producer cells and intravenous GCV. The titer of retroviral-vector was measured in cisternal and lumbar CSF samples after repeated producer cell injection.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Toxicity studies and distribution dynamics of retroviral vectors following intrathecal administration of retroviral vector-producer cells. 762 Dec 61

Current vaccines for the avian respiratory disease infectious laryngotrachetitis consist of naturally attenuated strains of the causative agent--the herpesvirus infectious laryngotracheitis virus (ILTV). Due to the dissemination of these viruses from vaccinated chickens as well as their possible reversion to more pathogenic forms, the use of genetically engineered viral vaccines lacking virulence factors while retaining antigenicity is being considered. Since the thymidine kinase (TK) activity of herpesviruses has been associated with virulence, inactivation of the encoding gene in the ILTV genome should attenuate the virus. Moreover, by analogy to other TK- herpesviruses, the ability of such ILTV mutants to induce a protective response in chickens should not be compromised. Therefore, the deliberate genetic alteration of ILTV was attempted. In order to prevent reversion and also to enable identification of the modified virus, a "marker" transcriptional unit (Escherichia coli lacZ gene fused to a SV-40 3'-polyadenylation signal sequence and regulated by the pseudorabies virus gX gene promoter) was inserted via homologous recombination at one of two loci within the ILTV TK gene. Recombinant viruses were identified and plaque-purified on the basis of their ability to produce beta-galactosidase. Retention of the foreign DNA at the predicted sites in the genomes of the recombinant ILTV was verified by Southern hybridization. Since their replication was unaffected by the thymine analog 1-(2-fluoro-2-deoxy-beta-D-arabinofuranosyl)-5-methyluracil, the recombinants appeared to have a TK- phenotype. Despite this apparent deficiency, prior inoculation of either recombinant virus into chickens afforded the birds protection against a lethal challenge of virulent ILTV. Moreover, the degree of respiratory distress in the chickens vaccinated with the recombinants was relatively mild compared to the severe reaction in birds receiving the parental virus. Thus, ILTV can be genetically attenuated without an accompanying loss of immunogenicity.
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PMID:Generation of thymidine kinase-deficient mutants of infectious laryngotracheitis virus. 777 65

The fidelity of homologous recombination in vaccinia virus (VV) DNA was examined by constructing a viral recombinant whose genome contained a copy of the Escherichia coli lac z gene in which the central third of the gene was repeated on either side of the VV thymidine kinase (tk) gene. In this virus, homologous DNA recombination and consequent excision of the tk gene were necessary to restore the open reading frame for beta-galactosidase and thereby to confer a lac z+ phenotype. Imprecise recombination was predicted to increase the frequency of lac z- virus. However, after several passages during which almost every viral genome underwent homologous recombination, the frequency of lac z- plaques was indistinguishable from that of a control virus that could express beta-galactosidase without prior recombination. We conclude that homologous recombination in VV DNA occurs with perfect fidelity at least 99% of the time.
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PMID:Fidelity of homologous recombination in vaccinia virus DNA. 777 3

The development and utilization of a tissue culture system for the analysis of quiescent, nonreplicating herpes simplex virus type 1 (HSV-1) genomes is described. It was demonstrated previously that the HSV-1 Vmw65 mutant in1814, which is impaired for immediate early (IE) transcription, was retained for many days in human fetal lung (HFL) fibroblasts in a quiescent 'latent' state. Molecular analysis of the viral genome was not possible, however, due to residual expression of IE proteins and consequent cytotoxicity at high m.o.i. In the study reported here, IE transcription was reduced further by pretreatment of cells with interferon-alpha (IFN-alpha) and by the use of mutant in1820, a derivative of in1814 in which the Vmw110 promoter was replaced by the Moloney murine leukaemia virus (Momulv) enhancer. The Momulv enhancer was not expressed under IE conditions; thus in1820 was more impaired for replication than in1814 and behaved as if deficient for both Vmw65 and Vmw110. In cells pretreated with IFN-alpha and subsequently infected with in1820 cytotoxicity was overcome, enabling a tissue culture system to be developed in which all cells stably retained at least one quiescent viral genome. To assist the analysis of gene expression, in1820 was further modified by insertion of the Escherichia coli lacZ gene controlled by the human cytomegalovirus enhancer (mutant in1883) or the HSV-1 immediate early Vmw110 promoter (in1884). Expression of beta-galactosidase was not detected after infection of IFN-alpha-pretreated cells with in1883 or in1884 but could be induced in almost all cells containing a viral genome, by superinfection of cultures. In1820-derived viruses were retained for at least 9 days and were not reactivated by subculture of cells. A regular arrangement of nucleosomes, as found in cellular chromatin, was not detected on the viral genome at the thymidine kinase locus. The non-linear genome was a template for reactivation with no requirement for prior conversion to a linear form. A small number of remaining linear genomes resulted from incomplete uncoating of input virus.
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PMID:Quiescent viral genomes in human fibroblasts after infection with herpes simplex virus type 1 Vmw65 mutants. 778 70

Three independently selected spontaneous thymidine kinase-negative mutants (TK-phenotype) and a recombinant with Escherichia coli beta-galactosidase gene (LacZ+ phenotype) inserted in the viral thymidine kinase gene (tk) were derived from a plaque-cloned isolate of K-1 ectromelia virus strain (TK+ phenotype). Dramatically decreased virulence of TK- variants was observed for all routes of mouse inoculation. The kinetics of TK+ and TK- variants in various target organs indicated a significant decrease of production and dissemination of TK- mutants and recombinant in the organs of mice. In the spleen and liver of intranasally or intracerebrally infected mice TK- virus was not detected during the entire period of observation. Analysis of organs homogenates of mice intranasally infected by a mixture of recombinant with TK-LacZ+ phenotype and parental isolate with TK+LacZ- phenotype on the monolayers of TK- cells indicated that only white plaques (LacZ-) with the TK+ phenotype appeared from liver and spleen homogenates. Thus, the mouse acts as a live filter much more efficiently than any other selective systems. Ultrastructural studies showed that viral damage in animals infected by TK- variants was far less than that observed in mice, infected with wild type of ectromelia virus and pathological lessions were slight and reversible. Replication of ectromelia virus TK- variants was blocked at the viroplasma stage in cells with a high level of differentiation in contrast to TK+ variants. Most likely, such restriction of target cells assortment is the general reason of reduced virulence in the case of tk-gene inactivation.
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PMID:Fine mechanisms of ectromelia virus thymidine kinase-negative mutants avirulence. 783 64


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