Insertion of a crude preparation of cyclic AMP (cAMP)-dependent protein kinase inhibitor (PKI) into a cloned mouse anterior pituitary cell line (AtT-20/D16-16) blocked cAMP-mediated hormone release. This was accomplished by developing a technique to incorporate PKI into multicellular cultures. The technique involved the encapsulation of the PKI into liposomes coupled to Protein A (a bacterial protein that binds to the Fc portion of antibodies). Application of such liposomes to AtT-20 cells targeted by pre-treatment with an antiserum against neural cell adhesion molecule (a cell surface glycoprotein expressed by these cells) resulted in the attachment of the liposomes onto the cell surface followed by the delivery of the liposome content into the cells. The AtT-20 cells respond to cAMP-promoting agents such as forskolin by secreting the hormone adrenocorticotropin (ACTH). Liposomes containing PKI and coupled to protein A specifically blocked cAMP-mediated ACTH release from cells treated with anti-N-CAM antibodies. In contrast, the ACTH release response to K+ or phorbol esters does not appear to involve cAMP and was not reduced by such manipulations. The specificity of PKI to block hormone release initiated by one but not by other secretagogues directly links cAMP-dependent protein kinase with the ACTH release process but suggests that there are other mechanisms also involved in stimulus-secretion coupling in corticotrophs.
Liposome delivery of cyclic AMP-dependent protein kinase inhibitor into intact cells: specific blockade of cyclic AMP-mediated adrenocorticotropin release from mouse anterior pituitary tumor cells.
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T Reisine, G Rougon, J Barbet; Liposome delivery of cyclic AMP-dependent protein kinase inhibitor into intact cells: specific blockade of cyclic AMP-mediated adrenocorticotropin release from mouse anterior pituitary tumor cells.. J Cell Biol 1 May 1986; 102 (5): 1630–1637. doi: https://doi.org/10.1083/jcb.102.5.1630
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