Translocation of cytosolic annexin 2 to a Triton-insoluble membrane subdomain upon nicotine stimulation of chromaffin cultured cells.
To gain a better understanding of the function of annexin 2, we have investigated the subcellular distribution of the monomeric and heterotetrameric forms of annexin 2 and their relationship to the cytoskeleton upon stimulation of chromaffin cells. Quantitative immunoblotting has revealed that in resting cells a large amount of annexin 2 is monomeric and cytosolic. Upon nicotine stimulation 80% of total annexin 2 becomes associated with a Triton-X100-insoluble fraction where the monomeric and the heterotetrameric forms are found. The translocation of monomeric annexin 2 is Ca2+-dependent and complete at 1 microM free Ca2+. We have shown that about 66% of the annexin 2 associated with the Triton-X100-insoluble fraction is soluble in octylglucoside while the remnants are insoluble in the detergent and remain likely associated with actin filaments and associated cytoskeleton proteins. The octylglucoside-soluble fraction contains integral proteins from the plasma membrane and from granule membrane, but does not contain caveolin. Moreover, upon nicotine stimulation, a redistribution of proteins was detected in this fraction. These dynamic processes appear concomitantly with the phosphorylation of annexin 2 in this compartment and with catecholamine release. It is suggested that the soluble octylglucoside fraction may represent a special lipidic membrane compartment where the NSF attachment proteins and the cytosolic proteins like annexin 2 and rab3a may become concentrated upon stimulation of the cell. The presence of annexin 2 is consistent with its proposed function on granule and target membrane proteins required for the close apposition of two distinct membranes and supports its functional role in the regulated exocytosis/endocytosis process.
Sagot
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Centre National de la Recherche Scientifique, U.P.R. 9071 de Neurobiologie Physicochimique, Institut de Biologie Physico-Chimique, Paris, France.
Regnouf
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Henry
J P
JP
Pradel
L A
LA
D000818:Animals; D017306:Annexin A2 / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000187:drug effects / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D005960:Glucosides / Q000494:pharmacology*; D008565:Membrane Proteins / Q000378:metabolism; D009538:Nicotine / Q000494:pharmacology; D017830:Octoxynol / Q000494:pharmacology; D010766:Phosphorylation
D017306:Annexin A2; D003902:Detergents; D005960:Glucosides; D008565:Membrane Proteins; C018619:octyl-beta-D-glucoside; D009538:Nicotine; D017830:Octoxynol