CLONING AND CHARACTERIZATION OF A NOVEL A-KINASE ANCHORING PROTEIN - AKAP-220, ASSOCIATION WITH TESTICULAR PEROXISOMES

Compartmentalization of the type II cyclic AMP-dependent kinase (PKA) is achieved through association of the regulatory subunit (RII) with A -kinase anchoring proteins (AKAPs), Using an interaction cloning strat egy with RII alpha as a probe, we have isolated cDNAs encoding a novel 1129-amino acid protein that contains both a PKA binding region and a peroxisome targeting motif. Northern analysis detected mRNAs of 9.7 a nd 7.3 kb in several rat tissues with the highest levels present in th e brain and testis, Western analysis and RII overlay experiments showe d that the protein is approximately 220 kDa and was, therefore, named AKAP 220. Immunoprecipitation of AKAP 220 from rat testis extracts res ulted in co-purification of the type II PKA holoenzyme, The specific a ctivity of PKA increased 458-fold from 7.2 pmol/min/mg in the cell lys ate to 3.3 nmol/min/mg in the immunoprecipitate. Immunohistochemical a nalysis of rat testicular TM4 cells showed that AKAP 220 and a proport ion of RII were co-localized in microbodies that appear to be a subset of peroxisomes. Collectively, these results suggest that AKAP 220 may play a role in targeting type II PKA for cAMP-responsive peroxisomal events.