Regulatory signals affecting a selective loss of mRNA in Dictyostelium discoideum

1989 ◽  
Vol 94 (3) ◽  
pp. 501-509
Author(s):  
H.H. Hassanain ◽  
W. Kopachik
Keyword(s):  
Cyclic Amp ◽  
Cell Surface ◽  
Dictyostelium Discoideum ◽  
Transcript Level ◽  
Cell Surface Receptor ◽  
Cell Contact ◽  
Specific Cell ◽  
Mrna Levels ◽  
Stage Of Development ◽  
Surface Receptor

We identified signals that affect mRNA levels complementary to a gene that is highly expressed in vegetative Dictyostelium discoideum cells. This gene has been cloned as cDNA in the plasmid pcD-D2. The level of transcripts homologous to pcD-D2 fell dramatically in strain XP55 during the aggregation stage of development when cells differentiate on agar. The level, however, did not fall simply as a result of starvation or aggregation-specific cell contact. Rather, before the level is reduced cells must be deprived of amino acids and cyclic AMP administered in amounts and at intervals in pulses to mimic cyclic AMP signal-relay in aggregation. This effect can be blocked either with cyclic AMP-S (a non-hydrolysable cyclic AMP analogue) or adenosine, both of which prevent cyclic AMP binding to the cyclic AMP cell surface receptor. It is also blocked in ‘frigid’ aggregation-deficient mutants HC85 and HC112 known to be defective in a G alpha protein. We conclude that the transcript level is balanced by positive nutritional signals acting against negative signals transduced in part through a cell surface cyclic AMP receptor.

Pharmacological characterization of cyclic AMP receptors mediating gene regulation in Dictyostelium discoideum

1986 ◽  
Vol 6 (7) ◽  
pp. 2402-2408
Author(s):  
B Haribabu ◽  
R P Dottin
Keyword(s):  
Gene Expression ◽  
Cyclic Amp ◽  
Cell Surface ◽  
Dictyostelium Discoideum ◽  
Cell Surface Receptor ◽  
Intracellular Camp ◽  
Response Curves ◽  
Pharmacological Characterization ◽  
Surface Receptor ◽  
Extracellular Molecules

Extracellular molecules regulate gene expression in eucaryotes. Exogenous cyclic AMP (cAMP) affects the expression of a large number of developmentally regulated genes in Dictyostelium discoideum. Here, we determine the specificity of the receptor(s) which mediates gene expression by using analogs of cAMP. The order of potency with which these analogs affect the expression of specific genes is consistent with the specificity of their binding to a cell surface receptor and is distinct from their affinity for intracellular cAMP-dependent protein kinase. Dose-response curves with cAMP and adenosine 3',5'-monophosphorothioate, a nonhydrolyzable analog, revealed that the requirement for high concentrations of exogenous cAMP for regulating gene expression is due to the rapid degradation of cAMP by phosphodiesterase. The addition of low concentrations of cAMP (100 nM) or analogs in pulses also regulates gene expression. Both the genes that are positively regulated by exogenous cAMP and the discoidin gene, which is negatively regulated, respond to cAMP analogs to the same degree. Genes expressed in prespore or prestalk cells are also similarly regulated. These data suggest that the effects are mediated through the same receptor. The specificity of this receptor is indistinguishable from that of the well-characterized cell surface cAMP receptor.

scholarly journals Pharmacological characterization of cyclic AMP receptors mediating gene regulation in Dictyostelium discoideum.

1986 ◽  
Vol 6 (7) ◽  
pp. 2402-2408 ◽  
Author(s):  
B Haribabu ◽  
R P Dottin
Keyword(s):  
Gene Expression ◽  
Cyclic Amp ◽  
Cell Surface ◽  
Dictyostelium Discoideum ◽  
Cell Surface Receptor ◽  
Intracellular Camp ◽  
Response Curves ◽  
Pharmacological Characterization ◽  
Surface Receptor ◽  
Extracellular Molecules

Extracellular molecules regulate gene expression in eucaryotes. Exogenous cyclic AMP (cAMP) affects the expression of a large number of developmentally regulated genes in Dictyostelium discoideum. Here, we determine the specificity of the receptor(s) which mediates gene expression by using analogs of cAMP. The order of potency with which these analogs affect the expression of specific genes is consistent with the specificity of their binding to a cell surface receptor and is distinct from their affinity for intracellular cAMP-dependent protein kinase. Dose-response curves with cAMP and adenosine 3',5'-monophosphorothioate, a nonhydrolyzable analog, revealed that the requirement for high concentrations of exogenous cAMP for regulating gene expression is due to the rapid degradation of cAMP by phosphodiesterase. The addition of low concentrations of cAMP (100 nM) or analogs in pulses also regulates gene expression. Both the genes that are positively regulated by exogenous cAMP and the discoidin gene, which is negatively regulated, respond to cAMP analogs to the same degree. Genes expressed in prespore or prestalk cells are also similarly regulated. These data suggest that the effects are mediated through the same receptor. The specificity of this receptor is indistinguishable from that of the well-characterized cell surface cAMP receptor.

Inositol 1,4,5-trisphosphate and calcium stimulate actin polymerization in Dictyostelium discoideum

1986 ◽  
Vol 82 (1) ◽  
pp. 41-51
Author(s):  
G.N. Europe-Finner ◽  
P.C. Newell
Keyword(s):  
Cyclic Amp ◽  
Dictyostelium Discoideum ◽  
Actin Polymerization ◽  
Cell Surface Receptor ◽  
Receptor Stimulation ◽  
Normal Cells ◽  
Cellular Slime Mould ◽  
Inositol 1,4,5 Trisphosphate ◽  
Surface Receptor ◽  
Cellular Slime

The effect of chemoattractants such as cyclic AMP and folate on amoebae of the cellular slime mould Dictyostelium discoideum is to cause a series of rapid intracellular responses. One of the most rapid of these responses is the polymerization of actin associated with the cytoskeleton, an event correlated with pseudopodium formation, which occurs within 3–5 s of chemotactic receptor stimulation. We report that this response can be mimicked by addition of 5 microM-inositol 1,4,5-triphosphate (IP3) or by addition of 100 microM-Ca2+ to saponin-permeabilized amoebae. The data suggest that cytoskeletal actin polymerization occurs in normal cells as a result of IP3 formation in response to cell surface receptor stimulation and the consequent release of Ca2+ from internal stores.

scholarly journals Decreased urokinase receptor expression by overexpression of the plasminogen activator in a colon cancer cell line

1992 ◽  
Vol 285 (2) ◽  
pp. 629-634 ◽  
Author(s):  
W Hollas ◽  
E Soravia ◽  
A Mazar ◽  
J Henkin ◽  
F Blasi ◽  
...  
Keyword(s):  
Cell Surface ◽  
Binding Site ◽  
Cell Line ◽  
Marker Gene ◽  
Receptor Expression ◽  
Cell Surface Receptor ◽  
Scatchard Analysis ◽  
Specific Cell ◽  
Ample Evidence ◽  
Surface Receptor

There is now ample evidence that the proteolytic action of urokinase (UK) is potentiated by a specific cell surface receptor. The present study was undertaken to determine the role of UK as a modulator of its binding site. GEO colonic cells, which secrete low levels of UK (approximately 2.5 ng/ml per 72 h per 10(6) cells) and display approx. 10(4) receptors per cell, the majority of which are vacant, were transfected with an exogenous UK gene driven by the RSV long terminal repeat (LTR) promoter (pRSVUK). Several UK-overexpressing pRSVUK clones were identified by an e.l.i.s.a., Northern blotting and Southern blotting, and analysed for receptor numbers after an acid pretreatment which dissociates receptor-bound UK. pRSVUK GEO clones, expressing high levels of UK, consistently bound 50-75% less radioactive di-isopropylfluorophosphate (DFP)-UK than clones harbouring the selectable marker gene neo only or control GEO cells. Cross-linking experiments with a radioactive N-terminal fragment of UK which binds to the receptor showed a decreased amount of a binding protein of approx. 51 kDa in representative pRSVUK-transfected cells. Saturation and Scatchard analysis indicated that this reduction in radioligand binding reflected a 40-70% decrease in the number of UK receptors, rather than a change in the dissociation constant. The reduction in receptor display could be accounted for by a decrease in the amount of steady-state mRNA encoding the receptor. Radioactive DFP-UK binding to pRSVUK GEO clones, which display two-thirds less receptors than their neo counterparts, could be restored to control levels (untreated cells harbouring neo) by cultivating them in the presence of an antibody which inhibits the interaction of UK with its receptor. These data suggest that for one colonic cell line at least, UK reduces the expression of its own binding site via an autocrine stimulation of its cell surface receptor.

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