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:3.2.1.23 (beta-galactosidase)
14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The extracellular spore coat of Dictyostelium discoideum is composed of three major proteins, SP96, SP70, and SP60, encoded by the cotA, cotB, and cotC genes, respectively. The spore coat proteins are coordinately synthesized in prespore cells shortly after aggregation, stored in prespore vesicles during the slug stage, and secreted during encapsulation of spores. We have ligated various portions of the upstream region of cotB to lacZ such that a protein consisting of the first nine amino acids of SP70 fused to beta-galactosidase is synthesized in prespore cells. Individual cells that accumulate the enzyme can be observed in situ during early aggregation due to the sensitivity of the assay. We have found that prespore cells first appear in a random distribution throughout the aggregates with no indication of spatial localization. They subsequently sort out from prestalk cells that form a tip on the aggregates. The cotB regulatory region was subdivided into a proximal and a distal region, each of which could independently direct proper temporal and cell-type control. Transcriptional activity directed by these two regions appears to be additive in the full-length regulatory region. The proximal region was shown to be complex in that removal of certain portions partially reduced transcriptional activity while removal of other portions abolished all activity. Nevertheless, cells transformed with constructs showing attenuated activity expressed the fusion gene at the proper time in development and the activity was localized to prespore cells. The cis-acting regions responsible for all aspects of cotB regulation appear to be closely opposed within the minimal essential sequence of the proximal region.
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PMID:Enhancer regions responsible for temporal and cell-type-specific expression of a spore coat gene in Dictyostelium. 848 18

Signal transduction via a family of cAMP receptor subtypes (cARs) is critical for proper development in the cellular slime mold Dictyostelium. Genes encoding four related subtypes have been cloned and their expression, based on RNA accumulation, has been previously reported. Here we report the differential spatial and temporal distribution of cAR2 and cAR3 proteins, based on indirect double immunofluorescence. Cells were transformed with a carB::lacZ construct, and an antibody against beta-galactosidase was used to visualize cAR2 expression. Simultaneously, a cAR3-specific antibody was used to identify cAR3-expressing cells. Results indicate that by the time of tip formation (12-14 hr) both receptors are expressed and distribute in a virtually nonoverlapping pattern, with cAR2 being expressed on anterior, prestalk cells and cAR3 present in the rest of the organism. Differential distribution of these two receptor subtypes may result in distinct cAMP signaling mechanisms in the two major regions of the organism.
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PMID:Differential distribution of cAMP receptors cAR2 and cAR3 during Dictyostelium development. 857 36

Dictyostelium discoideum is a simple eukaryote that lives as an amoeba until starvation triggers aggregation. The aggregate forms a slug which then develops into a fruiting body with two main cell types, stalk and spore cells. Two mechanisms have been proposed to explain cell-type differentiation. Studies using expression of the ecmA gene as a prestalk cell marker indicated that gradients of morphogens determine cell fate in the slug. However, studies using dyes or the cysteine proteinase 2 (CP2) gene product as a prestalk cell marker indicated that cell autonomous factors such as cell-cycle phase at the time of starvation cause an initial choice of cell fate. To help resolve these differences, we have used transformed cells containing the promoter of the prestalk gene ecmA fused to beta-galactosidase (Jermyn and Williams, 1991) to study the differentiation of Dictyostelium cells at low cell density, at which cell-to-cell interactions and morphogen gradients are minimal. We find that under all conditions of low cell density in which express ion the ecmA fusion gene occurs, it is invariably detected in less than 25% of the cells from a clonal population. This suggests that a cell-autonomous mechanism is involved in ecmA expression. We then used double-labeled immunofluorescence to examine the ontogeny of the CP2-positive and the ecmA-positive cells. In developing aggregates, 9 to 12% of the cells are CP2-positive from 12 to 24 hr of development. The ecmA-positive cells are first detected at 16 hr as a subset of the CP2-positive cells and then increase in number. At approximately 20 hr, the CP2-positive cells and the ecmA-positive cells are almost completely overlapping sets. By late development, all of the CP2-positive cells are ecmA-positive and an additional 10% of the CP2-negative cells are also ecmA-positive. This indicates that up to 20 hr development, ecmA is expressed only in CP2-positive cells. The data thus suggest that cell-cycle phase at the time of starvation causes an initial choice of cell type and that during later development other factors influence cell fate.
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PMID:Initial cell-type choice in a simple eukaryote: cell-autonomous or morphogen-gradient dependent? 861 80

We have analysed expression of the ecmA and ecmB genes of Dictyostelium by enzymatic double staining using beta-galactosidase and beta-glucuronidase reporter gene constructs. Cells expressing the ecmA gene first appear as scattered cells at the mound stage of development and we show that this is also true for cells expressing the ecmB gene. During tip formation the ecmA-expressing cells move to the apex of the mound, while the ecmB-expressing cells accumulate in the base. The ecmB-expressing cells constitute part of the basal disc if the culminant is formed in situ but are discarded if a migratory slug is formed. During slug migration they are replaced by a band of ecmB-expressing cells, situated in the front half of the prespore zone and tightly apposed to the substratum. When culmination is triggered these cells rapidly move to the back half of the prestalk zone, possibly acting as a point of attachment to the substratum. Ultimately, they are joined by cells at the back of the slug, the rearguard cells, to form the basal disc. Thus, contrary to previous belief, basal disc formation is initiated very early during culmination and occurs by the forward movement of cells located in the anterior of the prespore zone.
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PMID:The initiation of basal disc formation in Dictyostelium discoideum is an early event in culmination. 863 Dec 53

Dictyostelium discoidium cells express a family of cell surface cAMP receptors, and these G-protein-coupled receptors are each expressed with unique spatial and temporal patterns. One of these receptors, cAR2, is present during the postaggregative stages of development and our previous work suggests that it is preferentially expressed in prestalk cells. We report here the isolation of the promoter for carB, the gene which encodes cAR2. Using this fragment to generate a carB::lacZ, gene fusion construct, we investigated carB expression in detail. Expression is first detected at the tight aggregate stage and subsequently in a pattern reminiscent of the prestalk-specific gene ecmA. There are subtle differences, however, with, ecmA being expressed significantly in the anterior-like cells of the migrating pseudoplasmodium and in the basal disc and lower cup supporting the sorus during terminal development. carB is not expressed in any of these places. The presence of these different prestalk cell subtypes was confirmed by double indirect immunofluorescence using anti-cAR2 and anti-beta-galactosidase antibodies. While virtually all cAR2-expressing cells also express ecmA::lacZ, a substantial fraction of ecmA::lacZ-positive cells do not express cAR2. We also found the regulation of carB gene expression to differ from that of ecmA. carB expression is induced in vitro by extracellular cAMP, but surprisingly, not by DIF-1, a soluble molecule thought to be essential for the initiation of prestalk differentiation. Thus, cAR2 appears to be a cAMP receptor present on a restricted subset of prestalk cells and whose expression does not respond typically to the prestalk inducer DIF-1. DIF-1 sensitivity may, therefore, not be characteristic of all early prestalk differentiation.
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PMID:The cAMP receptor subtype cAR2 is restricted to a subset of prestalk cells during Dictyostelium development and displays unexpected DIF-1 responsiveness. 863 93

Prespore and prestalk cells can be distinguished within aggregates of Dictyostelium by the expression of well-characterized cell type-specific genes. Fusion of the tagB regulatory region to Escherichia coli beta-galactosidase revealed that this prestalk specific gene marks the differentiation of the initial prestalk cell population, PST-1. The reporter gene was expressed normally in tagB- mutant cells despite the fact that they do not accumulate measurable levels of DIF-I, a morphogen that was previously implicated in prestalk differentiation. In an independent experimental system, wild-type cells respond to the addition of DIF-I by induction of the prestalk marker ecmA and repression of the prespore marker cotB. We found that DIF-1 did not affect the expression of the tagB or carB genes, both of which are prestalk specific and essential for PST-A cell differentiation. We conclude that the initiation of prestalk development is not dependent on DIF-1 and suggest that the morphogen participates mainly at later stages.
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PMID:Initial cell type divergence in Dictyostelium is independent of DIF-1. 863 94

We have isolated the gene. rnrB, that encodes the ribonucleotide reductase small subunit of Dictyostelium discoideum. The deduced amino acid sequence of rnrB exhibits about 60% sequence identity with its homologues in other eukaryotes. As demonstrated by RNA blot analysis the rnrB transcript is detected in growing cells and decreases dramatically at the onset of development. The rnrB transcript reappears after the cells have formed multicellular aggregates. To further examine the pattern of expression, we have fused the rnrB promoter and part of its coding sequence to lacZ. The transgenic strain bearing such a reporter construct expresses the fusion gene with a biphasic profile, which is indistinguishable from that of the endogenous rnrB. The multicellular aggregates of Dictyostelium are differentiated along the anterior-posterior axis. Cells in the anterior give rise to the stalk of the fruiting body while cells in the posterior are precursors of spores. Results from histochemical staining show that beta-galactosidase activity is detected exclusively in the posterior two-thirds of the aggregates. These data suggest that rnrB is expressed in prespore cells during postaggregative development and in vegetative cells.
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PMID:A prespore-specific gene of Dictyostelium discoideum encodes the small subunit of ribonucleotide reductase. 895 Jan 85

Almost all methods for transformation of the social ameba Dictyostelium discoideum rely on axenic growth, that is, growth in a synthetic medium, for at least part of the procedure. Axenic growth requires several mutations. Here we describe a procedure that can be used to transform wild-type strains which are able to grow only on the natural food source, bacteria. The method relies on a new selection cassette driven by the V18 promoter, a promoter that we show is substantially more active during growth on bacteria than the actin-6 promoter, which is widely used for axenic transformation. The procedure gives transformation frequencies of about 10(-5) with both strains Ax2 (capable of axenic growth) and NC4 (capable of growth only on bacteria). Using this vector, we have obtained NC4 strains carrying several beta-galactosidase reporter cassettes. Our vector can also be used in axenic transformations.
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PMID:Wild-type strains of Dictyostelium discoideum can be transformed using a novel selection cassette driven by the promoter of the ribosomal V18 gene. 900 12

We have examined the promoter of rnrB, the gene encoding the small subunit of ribonucleotide reductase of Dictyostelium discoideum, using lacZ as a reporter gene. Deletion analysis showed that expression of this gene in vegetative cells involves an A/T-rich element, whereas its expression in prespore cells during development requires a region encompassing two G/C-rich elements, designated box A and box B. Removal of boxes A and B results in very low level of activity. When either box A or box B is deleted, prestalk cells adjacent to the prespore zone also express beta-galactosidase. The behavior of these cis-regulatory elements implies that the mechanism regulating the prespore-specific expression of rnrB is different from that regulating other known prespore genes. We have used electrophoretic mobility shift assays to identify factors that interact with box A and box B. Box A interacts with a factor that is found in the nuclear fraction. While box B interacts with a factor that is present in the cytosolic fraction throughout growth and development, its presence in the nuclear fraction is developmentally regulated. Results from competition assays suggest that both box A and box B interact with transcriptional activators that have not been characterized previously.
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PMID:Identification of cis-regulating elements and trans-acting factors regulating the expression of the gene encoding the small subunit of ribonucleotide reductase in Dictyostelium discoideum. 1040 Jun 62

Starvation for amino acids initiates the developmental cycle in the cellular slime mold, Dictyostelium discoideum. Upon starvation one of the earliest developmental events is the selective loss of the ribosomal protein mRNAs from polysomes. This loss depends upon sequences in the 5' non-translated leader of the ribosomal protein (r-protein) mRNAs. Here evidence is presented which indicates that those cells which will become prestalk cells express the ribosomal protein genes during development under starvation conditions. Cells which enter the prespore pathway shut off r-protein synthesis. The promoter and 5' non-translated leader sequences from two ribosomal protein genes, the rp-L11 and the rp-S9 genes, are fused to the Escherichia coli beta-galactosidase reporter gene. While beta-galactosidase enzyme activity is detected in situ in most growing cells, by 15 h of development beta-galactosidase enzyme activity is largely lost from the prespore cells although strong beta-galactosidase enzyme activity is present in the prestalk cells. These observations suggest the possibility that the ribosomal protein mRNAs are excluded from polysomes in a cell-type-specific manner.
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PMID:Ribosomal protein gene expression is cell type specific during development in Dictyostelium discoideum. 1055 May 41


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