Loss of nuclear cyclic AMP binding in cyclic AMP-unresponsive Walker 256 mammary carcinoma.

A marked increase of cyclic AMP-binding and protein kinase activities occurs in the nuclei of N6,02’-dibutyryl adenosine 3’:5’-monophosphate (Bt,cAMP)-responsive Walker 256 mammary carcinoma (W256) following incubation of the tumor slices with CAMP in uitro. The macromolecular fraction containing [“HIcAMP in the nuclei can be extracted with 1.0 M KC1 and identified by acrylamide gel electrophoresis. Cytoplasmic origin of these increased nuclear CAMP-binding and protein kinase activities is suggested by the following observations: (a) cytoplasmic and nuclear CAMP-binding and protein kinase activities are inversely related during the CAMP stimulation of tumor slices; (b) the sequential transfer of CAMP-binding proteins and protein kinase to the nucleus is a temperature-dependent process; and (c) an initial interaction of CAMP with cytoplasm is an absolute prerequisite for the nuclei binding in vitro. The nuclear translocation of CAMP-binding proteins and protein kinase is greatly diminished in the other type (Bt,cAMP-unresponsive) of W256, which grows during the administration of B&CAMP in duo. The experiments using a cell-free system show that cytoplasmic CAMP-binding protein . CAMP complex from responsive W256 binds to isolated nuclei from both responsive and unresponsive tumors, whereas the complex from the unresponsive tumor binds neither nuclei. These results suggest that the lack of nuclear accumulation of CAMP-binding proteins and protein kinase observed in unresponsive W256 could have been due to a defect in cytoplasmic CAMP-binding proteins which fail to interact with nuclear components. Cyclic AMP-binding proteins of unresponsive W256 also fail to respond to endogenously generated CAMP: e.g. when tumor slices are incubated with prostaglandin E, (PGE,) in vitro CAMP-binding proteins in unresponsive W256 do not respond to the PGE, stimulus as do the binding proteins in responsive W256, despite a significant elevation of the CAMP level in the tumor slices. These results suggest that a molecular lesion in CAMP-binding proteins can be a cause of B&CAMP unre-