Effect of protein synthesis inhibition on gene expression during early development of Dictyostelium discoideum.

Singleton, Charles K.; Manning, Suzanne; Feng, Yi

doi:10.1128/mcb.8.1.10

Cited by 19 publications

(9 citation statements)

References 34 publications

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“…Recently, Singleton and co-workers showed that a set of vegetatively expressed genes which are normally repressed during early differentiation can be divided into two groups. One group is induced in response to cycloheximide, whereas the second group is unaffected by the antibiotic (24). Furthermore, Firtel has shown that the gene F9 that is normally induced during early development is superinduced in response to cycloheximide (16).…”

Section: Resultsmentioning

confidence: 99%

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Regulation of DdrasG gene expression during Dictyostelium development.

Khosla¹,

Robbins²,

Spiegelman³

1990

Mol. Cell. Biol.

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DdrasG gene expression during the early development of Dictyostelium discoideum has been examined in detail. The amount of DdrasG-specific mRNA increased approximately twofold during the first 2 to 3 h of development and then declined rapidly, reaching negligible levels by the aggregation stage. The increase in mRNA levels that occurred during the first 2 to 3 h of development also occurred during differentiation in cell suspensions and was enhanced when cells were shaken rapidly. This initial increase was unaffected by cell density. When cells were set up to differentiate on filters, the addition of a glucose-amino acid mixture slightly delayed differentiation and had a similar effect on the expression of the gene. The decline in DdrasG expression during development did not occur when cells were treated with cycloheximide, suggesting that the expression of a developmentally regulated gene product is essential for the reduction of DdrasG gene mRNA. There was no decrease in DdrasG mRNA level during differentiation in shake suspension, but the decrease did occur upon application of pulses of cyclic AMP to shaking cultures. The application of a continuously high level of cyclic AMP delayed the increase in expression of the gene and did not result in the subsequent decline. These results suggest that the induction of a functional cyclic AMP relay system is important in reducing DdrasG gene mRNA levels.

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Section: Resultsmentioning

confidence: 99%

“…Cycloheximide is a potent inhibitor of protein synthesis in D. discoideum (24). Recently, Singleton and co-workers showed that a set of vegetatively expressed genes which are normally repressed during early differentiation can be divided into two groups.…”

Section: Resultsmentioning

confidence: 99%

Regulation of DdrasG gene expression during Dictyostelium development.

Khosla¹,

Robbins²,

Spiegelman³

1990

Mol. Cell. Biol.

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“…Optimal annealing temperatures to reduce secondary products were determined for each reaction by temperature gradient PCR, and melting temperature analysis was performed on real-time reactions to ensure that secondary products were not present. Relative amounts of sample RT product were standardized to H7, a control gene (Singleton et al, 1988). Annealing was performed for 5 s at 47°C, and extension time was 14 s. Primers for talA were talA3: 5Ј-CCATGGTTGCTGCAACAATCGTA-GATGC-3Ј (nucleotides 7092-7114) and talA4: 5Ј-CTCGAGTTAATTTTTAT-TATAATTTTGTTTTCTTG-3Ј (nucleotides 7648 -7676); primers for H7 were H7S: 5Ј-ACGTTCAAACTAAATACGGAGCTGGT-3Ј (nucleotides 5-30) and H7AS: 5Ј-TTTGAGTGGTTTGCCAATTTCTTTT-3Ј (nucleotides 288 -312).…”

Section: Quantitative Pcrmentioning

confidence: 99%

Talin Influences the Dynamics of the Myosin VII-Membrane Interaction

Galdeen

Stephens

Thomas³

et al. 2007

MBoC

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Myosin VII (M7) and talin are ancient and ubiquitous actin-binding proteins with conserved roles in adhesion. Talin serves to link membrane receptors to the underlying actin cytoskeleton and forms a complex with M7 in Dictyostelium. The levels of talinA are tightly linked to M7 levels in Dictyostelium. Cells lacking M7 exhibit an 80% decrease in steady-state levels of talinA, whereas increased levels of M7 result in concomitant increases in total talinA. In contrast, changes in talinA levels do not affect M7 levels. Immunoprecipitation reveals that talinA and M7 are associated with each other in membrane fractions. Fluorescence recovery after photobleaching experiments on green fluorescent protein (GFP)-M7 cells expressing different levels of the M7 and talinA show that changes in the overall amounts of these two proteins influences the dynamics of membrane-associated M7. The recovery of GFP-M7 on the membrane is faster in cells lacking talinA and limited in the presence of excess amounts of talinA and M7. These results establish that M7 stabilizes talinA in the cytosol and, in return, talinA regulates the residence time of M7 at the plasma membrane, suggesting that these two proteins are both part of the same dynamic adhesion complex on the plasma membrane.

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“…For the BS153 panel, the band corresponding to H7 mRNA (lower band) is shown as an internal control. The H7 band increases with time in the actD + cyc lanes as H7 transcription is induced by cycloheximide [53]. …”

Section: Resultsmentioning

confidence: 99%

IfkA, a presumptive eIF2α kinase of Dictyostelium, is required for proper timing of aggregation and regulation of mound size

2003

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BackgroundThe transition from growth to development in Dictyostelium is initiated by amino acid starvation of growing amobae. In other eukaryotes, a key sensor of amino acid starvation and mediator of the resulting physiological responses is the GCN2 protein, an eIF2α kinase. GCN2 downregulates the initiation of translation of bulk mRNA and enhances translation of specific mRNAs by phosphorylating the translation initiation factor eIF2α. Two eIF2α kinases were identified in Dictyostelium and studied herein.ResultsNeither of the eIF2α kinases appeared to be involved in sensing amino acid starvation to initiate development. However, one of the kinases, IfkA, was shown to phosphorylate eIF2α from 1 to 7 hours after the onset of development, resulting in a shift from polysomes to free ribosomes for bulk mRNA. In the absence of the eIF2α phosphorylation, ifkA null cells aggregated earlier than normal and formed mounds and ultimately fruiting bodies that were larger than normal. The early aggregation phenotype in ifkA null cells reflected an apparent, earlier than normal establishment of the cAMP pulsing system. The large mound phenotype resulted from a reduced extracellular level of Countin, a component of the counting factor that regulates mound size. In wild type cells, phosphorylation of eIF2α by IfkA resulted in a specific stabilization and enhanced translational efficiency of countin mRNA even though reduced translation resulted for bulk mRNA.ConclusionsIfkA is an eIF2α kinase of Dictyostelium that normally phosphorylates eIF2α from 1 to 7 hours after the onset of development, or during the preaggregation phase. This results in an overall reduction in the initiation of protein synthesis during this time frame and a concomitant reduction in the number of ribosomes associated with most mRNAs. For some mRNAs, however, initiation of protein synthesis is enhanced or stabilized under the conditions of increased eIF2α phosphorylation. This includes countin mRNA.

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Effect of protein synthesis inhibition on gene expression during early development of Dictyostelium discoideum.

Cited by 19 publications

References 34 publications

Regulation of DdrasG gene expression during Dictyostelium development.

Regulation of DdrasG gene expression during Dictyostelium development.

Talin Influences the Dynamics of the Myosin VII-Membrane Interaction

IfkA, a presumptive eIF2α kinase of Dictyostelium, is required for proper timing of aggregation and regulation of mound size

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