Differential localization of protein kinase a type II isozymes in the Golgi-centrosomal area

Selectivity in the action of cAMP may be mediated by compartmentalized pools of cyclic AMP-dependent protein kinase (PKA), PKA type II is directed to different subcellular loci by interaction of the type II regulatory subunits (RII alpha, RII beta) with A-kinase anchoring proteins. In order to separately investigate the subcellular localization of PKA type II isozymes, monospecific antibodies to human RII alpha and RII alpha subunits of PHA were developed. We demonstrate that centrosomes bind both RII alpha and RII beta. Centrosomes were the preferred intracellular anchoring site for RII beta. However, centrosomal localization of RII beta was observed only in some highly differentiated cells such as keratinocytes, granulosa cells, and macrophages and in all neoplastic cell lines examined. Centrosomal localization of RII beta was not observed in normal undifferentiated cells such as fibroblasts, myoblasts, and T and B cells. In contrast, RII alpha was abundant in the Golgi area and in the trans-Golgi network (TGN). Furthermore, although RIIa appeared to colocalize with microtubules in the Golgi/TGN, extractions with nonionic detergent demonstrated that RII alpha was mainly membrane-associated. In addition, alterations of microtubule dynamics with Nocodazole or Taxol affected the distribution of the detergent-extractable pool of RII alpha, indicating that RII alpha may localize with microtubule-associated vesicles, Thus, RII alpha and RII beta clearly localize differently in the Golgi-centrosomal region. This indicates specific roles for PKA isozymes containing either RII alpha or RII beta. (C) 1999 Academic Press.