cAMP oscillations restrict protein kinase A redistribution in insulin-secreting cells.
Activation of hormone receptors was recently found to evoke oscillations of the cAMP concentration ([cAMP]) beneath the plasma membrane of insulin-secreting cells. Here we investigate how different time courses of cAMP signals influence the generation of cytoplasmic Ca(2+) signals and nuclear translocation of the PKA (protein kinase A) catalytic subunit in individual INS-1 beta-cells. [cAMP] was measured with a fluorescent translocation biosensor and ratiometric evanescent wave microscopy. Analysis of PKA nuclear translocation was performed with epifluorescence microscopy and FlAsH (fluorescein arsenical helix binder) labelling of tetracysteine-tagged PKA-Calpha subunit. Both oscillatory and stable elevations of [cAMP] induced by intermittent or constant inhibition of phosphodiesterases with isobutylmethylxanthine evoked Ca(2+) spiking. During [cAMP] oscillations, the Ca(2+) spiking was restricted to the periods of elevated [cAMP]. In contrast, only stable [cAMP] elevation induced nuclear entry of FlAsH-labelled PKA-Calpha. These results indicate that oscillations of [cAMP] lead to selective target activation by restricting the spatial redistribution of PKA.
Dyachok
O
O
Department of Medical Cell Biology, Uppsala University, Biomedical Centre, Box 571, SE-75123 Uppsala, Sweden.
Sågetorp
J
J
Isakov
Y
Y
Tengholm
A
A
D021581:Active Transport, Cell Nucleus; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020134:Catalytic Domain; D045744:Cell Line, Tumor; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D051381:Rats
D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D002118:Calcium