Human myometrial quiescence and activation during gestation and parturition involve dramatic changes in expression and activity of particulate type II (RII alpha) protein kinase A holoenzyme.
There are substantial data indicating that components of the cAMP-signaling pathway are differentially expressed in the human myometrium during pregnancy. The effects of cAMP in most tissues and cell types are mainly modulated via protein kinase A, a heterotetrameric protein complex consisting of two regulatory (R) and two catalytic (C) subunits. In the studies presented here, we used specific antibodies in Western blotting/immunoprecipitation, RT-PCR, and functional protein kinase A (PKA) phosphorylation assays to determine the PKA holoenzymes that are expressed in the human myometrium throughout pregnancy and labor. We report that as early as the second trimester of pregnancy, there is a significant increase in expression of the regulatory RII alpha protein subunit of PKA in the myometrium. This increase in protein expression is also mirrored at the mRNA level, indicating transcriptional control throughout pregnancy, whereas during parturition both transcript and protein are significantly decreased. This increase in RII alpha protein also resulted in increased particulate PKA activity in the myometrium during gestation, which was subsequently decreased during labor. Two specific A kinase anchoring proteins, AKAP95 and AKAP79, which have high binding affinities for RII alpha subunits, were found to form complexes with myometrial RII alpha species employing immunoprecipitation assays, but their levels of expression remained uniform in all myometrial tissue samples investigated. Our findings indicate that increased particulate type II PKA activity occurs throughout pregnancy, therefore directing the cAMP quiescence signal to specific subcellular loci within myometrial smooth muscle cells including the contractile machinery at the cytoskeleton; this effect is then removed during parturition.
Department of Obstetrics and Gynaecology, School of Surgical and Reproductive Sciences, University of Newcastle upon Tyne, Royal Victoria Infirmary, Newcastle upon Tyne NE1 4LP, United Kingdom.
Malcolm W J
MW
MacDougall
G Nicholas
GN
Europe-Finner
Stephen C
SC
Robson
D054758:A Kinase Anchor Proteins; D048868:Adaptor Proteins, Signal Transducing; D000328:Adult; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism; D054754:Cyclic AMP-Dependent Protein Kinase RIIalpha Subunit; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D005865:Gestational Age; D006801:Humans; D007150:Immunohistochemistry; D007163:Immunosorbent Techniques; D047908:Intracellular Signaling Peptides and Proteins; D008875:Middle Aged; D009215:Myometrium / Q000201:enzymology*; D009687:Nuclear Proteins / Q000378:metabolism; D036801:Parturition; D010766:Phosphorylation; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction
D054758:A Kinase Anchor Proteins; C487775:AKAP5 protein, human; C490117:AKAP8L protein, human; D048868:Adaptor Proteins, Signal Transducing; C415703:Akap5 protein, rat; D002352:Carrier Proteins; D054754:Cyclic AMP-Dependent Protein Kinase RIIalpha Subunit; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; C517819:PRKAR2A protein, human; C517822:Prkar2a protein, rat; D012333:RNA, Messenger; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases