scholarly journals Dominant Negative Forms of Akt (Protein Kinase B) and Atypical Protein Kinase Cλ Do Not Prevent Insulin Inhibition of Phosphoenolpyruvate Carboxykinase Gene Transcription

1999 ◽  
Vol 274 (30) ◽  
pp. 21305-21312 ◽  
Author(s):  
Ko Kotani ◽  
Wataru Ogawa ◽  
Yasuhisa Hino ◽  
Tadahiro Kitamura ◽  
Hikaru Ueno ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Gene Transcription ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Protein Kinase B ◽  
Dominant Negative ◽  
Atypical Protein Kinase ◽  
Insulin Inhibition ◽  
Phosphoenolpyruvate Carboxykinase Gene

Expression of a prenylation-deficient Rab4 inhibits the GLUT4 translocation induced by active phosphatidylinositol 3-kinase and protein kinase B

2001 ◽  
Vol 356 (1) ◽  
pp. 143-149 ◽  
Author(s):  
Mireille CORMONT ◽  
Nadine GAUTIER ◽  
Karine ILC ◽  
Yannick Le MARCHAND-BRUSTEL
Keyword(s):  
Protein Kinase ◽  
Protein Kinase B ◽  
Active Form ◽  
Small Gtpase ◽  
Dominant Negative ◽  
Glut4 Translocation ◽  
Phosphatidylinositol 3 Kinase ◽  
Isolated Adipocytes ◽  
Phosphatidylinositol 3

The small GTPase Rab4 has been shown to participate in the subcellular distribution of GLUT4 under both basal and insulin-stimulated conditions in adipocytes. In the present work, we have characterized the effect of Rab4 ΔCT, a prenylation-deficient and thus cytosolic form of Rab4, in this process. We show that the expression of Rab4 ΔCT in freshly isolated adipocytes inhibits insulin-induced GLUT4 translocation, but only when this protein is in its GTP-bound active form. Further, it not only blocks the effect of insulin, but also that of a hyperosmotic shock, but does not interfere with the effect of zinc ions on GLUT4 translocation. Rab4 ΔCT was then shown to prevent GLUT4 translocation induced by the expression of an active form of phosphatidylinositol 3-kinase or of protein kinase B, without altering the activities of the enzymes. Our results are consistent with a role of Rab4 ΔCT acting as a dominant negative protein towards Rab4, possibly by binding to Rab4 effectors.

Control of CCK gene transcription by PACAP in STC-1 cells

2000 ◽  
Vol 279 (3) ◽  
pp. G605-G612 ◽  
Author(s):  
Damian G. Deavall ◽  
Raktima Raychowdhury ◽  
Graham J. Dockray ◽  
Rod Dimaline
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Gene Transcription ◽  
Transcriptional Start Site ◽  
Dominant Negative ◽  
Activator Protein 1 ◽  
Mobility Shift ◽  
Creb Phosphorylation ◽  
Mobility Shift Assay ◽  
Kinase A

The mechanisms by which neuroendocrine stimulants regulate CCK gene transcription are unclear. We examined promoter activation by pituitary adenylate cyclase-activating polypeptide (PACAP), a known CCK secretagogue, in the enteroendocrine cell line STC-1. The promoter region from −70 to −87 bp, relative to the transcriptional start site, contains a composite calcium/cyclic AMP response element (CRE)/activator protein 1 (AP1) site that may bind CRE binding protein (CREB) and AP1. PACAP (with IBMX) stimulated expression of an 87-bp construct 3.35 ± 0.36-fold but had no effect on a −70 construct. The effect was blocked by the protein kinase A inhibitor H-89 and by a dominant-negative CREB plasmid. Mutation of the CRE/AP1 site to a canonical CRE site did not affect the response to PACAP, but mutation to a canonical AP1 site prevented it. CREB phosphorylation was increased after PACAP treatment. Electrophoretic mobility shift assay and supershift analysis revealed that CREB and not AP1 bound to the CRE/AP1 site and that PACAP increased the proportion of phosphorylated CREB that was bound. We conclude that PACAP increases CCK gene expression via a cAMP-mediated pathway involving CREB phosphorylation by protein kinase A and activation of a composite CRE/AP1 site.

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