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

The lymphocyte cell surface receptor for the high endothelial venules (HEV's) of peripheral lymph nodes is specifically recognized by the monoclonal antibody MEL-14. Three independent complementary DNA (cDNA) clones, each of which encodes the protein ubiquitin, were detected by virtue of the expression of the MEL-14 antigenic determinant on cDNA-beta-galactosidase bacterial fusion proteins. The antigenic determinant defined by MEL-14 resides in the carboxyl terminal 13-amino-acid proteolytic peptide of ubiquitin, but is undetected in intact undenatured ubiquitin and other cellular ubiquitinated proteins. Antisera and monoclonal antibodies to ubiquitin determinants bind to the surface of both HEV-receptor positive and negative cell lines. The MEL-14-identified cDNA clones hydridize to RNA transcripts that encode tandemly repeated ubiquitins. Sequence analysis of these polyubiquitin cDNA's does not identify a leader sequence for export to the cell surface. The expression of the MEL-14 epitope of ubiquitin depends upon its local environment. The steady-state levels of expression of the ubiquitin messenger RNA's do not correlate with either the tissue derivation of the RNA or the expression of the lymphocyte HEV receptor. Regulation of the expression of the HEV receptor is not likely to reflect the transcriptional control of ubiquitin genes, but rather to reflect control of the expression of the HEV core polypeptide or its level or form of ubiquitination.
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PMID:Expression cloning of a lymphocyte homing receptor cDNA: ubiquitin is the reactive species. 300 14

In order to have a proper biosynthesis and secretion of the melanin-pigment granules (melanosomes) the melanocyte may require a melanosome-associated molecule that provides a signal for assembly and organization of melanogenic enzymes and proteins within the compartment of melanosomes. This study reports the presence of a Ca(2+)-binding phosphoprotein, p90, which can be engaged in such melanogenic function, located on the melanosomal membrane of human melanocytes. A human melanoma cDNA expression library in lambda Zap II was screened with a rabbit polyclonal antibody raised against human melanosomes isolated from cultured human melanoma cells, SK MEL 23. A cDNA encoding a melanosomal protein, M(r) 90 kDa, was identified through this immunoscreening. A partial sequencing of nucleotides (822 bp from the N-terminal domain) of this clone (3.8 kb) and predicted amino acids showed more than 90% homology with dog calnexin, a previously reported endoplasmic reticulum (ER) transmembrane protein. A fusion protein of this p90 with beta-galactosidase expressed in Escherichia coli revealed both the immuno-cross-reactivity with anti-dog calnexin and anti-human melanosome antibodies and the Ca(2+)-binding property. Upon immunohistochemistry, the anti-dog calnexin antibody revealed the positive immunoreactivities with both normal and malignant human melanocytes, showing a much higher expression of antigenic epitope than nonmelanocytic human cells. The laser scanning confocal immunofluorescence, using an antibody against a human melanosome-specific antigen (HMSA-5), and immunoelectron microscopy, using immunogold, confirmed the major localization of anti-dog calnexin antibody epitope on the melanosomes and ER.
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PMID:Identification of a cDNA coding for a Ca(2+)-binding phosphoprotein (p90), calnexin, on melanosomes in normal and malignant human melanocytes. 826 46

The beta-globin locus control region (LCR) confers high levels of position-independent, copy number-dependent expression onto globin transgenes. Here > 40 independent transgenic mouse lines and founders that carried the LCR in cis with the beta-globin gene promoter driving a lacZ reporter gene were studied. Expression of the lacZ transgene was assayed by measuring beta-galactosidase enzyme activity in fetal liver extracts, the levels of which correlated with the quantity of lacZ mRNA determined using RNase protection assays. Unexpectedly, expression of the lacZ transgene was found to show strong position effects, varying as much as 700-fold per transgene copy. These position effects occurred even if the whole beta-globin gene was incorporated as part of the lacZ reporter gene. Moreover, DNase I-hypersensitive sites appeared in the transgene LCR in high expressing but not in low expressing lines, suggesting that the LCR itself was position dependent. In contrast, MEL cell clones, in which transcriptionally active integration sites were selected for, gave < 13-fold variation in expression per copy of an LCR-lacZ construct. These results show that the lacZ reporter affects the ability of the LCR to activate chromatin in mice and that culture cells are not an adequate model for position-independent gene expression studies.
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PMID:The beta-globin locus control region enhances transcription of but does not confer position-independent expression onto the lacZ gene in transgenic mice. 867 Aug 75

One problem limiting the development of long-term gene replacement therapy is gene silencing. A variety of experiments have implicated DNA methylation and histone deacetylation in gene silencing and shown that the agents 5-azacytidine (5-Aza) and trichostatin A (TSA) are able to reverse these effects. To begin to investigate clinically relevant strategies to reverse silencing with these drugs, we transduced the MEL and FDCP-1 hematopoietic cell lines with Moloney murine leukemia virus (MMLV) and Harvey murine sarcoma virus (HMSV)-based retroviral vectors carrying the beta-galactosidase/neomycin resistance fusion gene (beta-geo). Fifty-one clones were isolated under G418 selection over 2 weeks and then allowed to grow without selection as beta-gal activity was monitored over time. More than 80% of these clones showed significant silencing over a period of 70-80 days. The clones were then exposed to a wide range of 5-Aza and TSA concentrations, both alone and in combination, in an effort to reverse silencing. Despite demonstration that the agents were able to decrease DNA methylation and increase histone acetylation, significant reversal of long-term silencing was not seen under any experimental condition. These results suggest that long-term retroviral silencing involves mechanisms in addition to DNA methylation and histone acetylation and that new pharmacologic strategies are needed to overcome the silencing process.
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PMID:Long-term silencing of retroviral vectors is resistant to reversal by trichostatin A and 5-azacytidine. 1080 88

Melanoma has proven to be resistant to conventional chemotherapy; however,the mechanism of chemoresistance is still unclear. Recent reports show that the transcription factor, E2F-1, may play a role in mediating cytotoxicity of certain chemotherapeutic agents. We have shown in a previous study that adenovirus-mediated overexpression of E2F-1 can efficiently induce apoptosis in melanoma cells. In the present study, the effect of E2F-1 expression on drug sensitivity of melanoma cells was evaluated. Two human melanoma cell lines, SK-MEL-28 and SK-MEL-2, were treated with drugs (etoposide, Adriamycin, roscovitine, cisplatin, 5-fluorouracil, or cycloheximide), alone or in combination with adenoviral vectors expressing beta-galactosidase (Ad-LacZ) or E2F-1 (Ad-E2F-1) at a multiplicity of infection of 1 in vitro. E2F-1 expression was confirmed by Western blot analysis. Sublethal concentrations of each drug alone or infection with Ad-E2F-1 alone produced <5% apoptosis by 3 days posttreatment. Conversely, cotreatment with Ad-E2F-1 and low concentrations of etoposide or Adriamycin markedly sensitized melanoma cells to apoptotic cell death. A slight enhancement of the cytotoxicity of roscovitine was demonstrated in combination with E2F-1 overexpression, but not to cisplatin, 5-fluorouracil, or cycloheximide. Ad-LacZ infection showed no obvious effects on drug sensitivity. Overexpression of p21 can block apoptosis induced by the combination chemogene therapy of Ad-E2F-1 and topoisomerase II poisons and does not require its proliferating cell nuclear antigen-binding ability. The protein synthesis inhibitor cycloheximide also has a cytotoxicity-protective effect against topoisomerase II inhibitor/E2F-1-induced apoptosis and suggests that new protein synthesis is required for this process. Topoisomerase II inhibitors also cooperated with Ad-E2F-1 to enhance antitumor activity in an in vivo model using xenografts in nude mice. When combined with Adriamycin or etoposide, E2F-1 adenovirus therapy resulted in an 87% or 91% decrease in tumor size, respectively, compared with controls (P < 0.002). Our results show that adenovirus-mediated E2F-1 gene transfer can sensitize melanoma cells to some chemotherapeutic agents, particularly topoisomerase II poisons, in vitro and in vivo. These results suggest a new chemosensitization strategy for melanoma gene therapy.
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PMID:Adenovirus-mediated E2F-1 gene transfer sensitizes melanoma cells to apoptosis induced by topoisomerase II inhibitors. 1191 54