title: Identification of the major site of in vitro PKA phosphorylation in the polycystin-1 C-terminal cytosolic domain.
abstract: Sequence analysis of the C-terminal cytosolic domain of human and mouse polycystin-1 has identified three RxS consensus protein kinase A (PKA) phosphorylation motifs. GST-fusion proteins containing the full-length and truncated C-terminal cytosolic domain of murine polycystin-1 were phosphorylated in vitro by the purified catalytic subunit of PKA. This identified a sequence of 25 amino acids, immediately downstream of a previously identified heterotrimeric G-protein activation sequence, as the major site of PKA phosphorylation. Phosphorylation of wild-type and alanine substituted synthetic peptides containing this motif demonstrated that alanine substitution of serine 4159 largely eliminated phosphorylation. Mutation of this residue in the fusion protein reduced phosphorylation by about 70%, whereas mutation of the other two conserved phosphorylation motifs had little effect. We conclude that serine 4159 is the major site of PKA phosphorylation in the C-terminal cytosolic domain of murine polycystin-1.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, 66160-7421, USA.
authors.forename: S C
authors.initials: SC
authors.lastname: Parnell
authors.forename: B S
authors.initials: BS
authors.lastname: Magenheimer
authors.forename: R L
authors.initials: RL
authors.lastname: Maser
authors.forename: J P
authors.initials: JP
authors.lastname: Calvet
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D003600:Cytosol / Q000737:chemistry*; D051379:Mice; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017354:Point Mutation; D011506:Proteins / Q000737:chemistry*; D011993:Recombinant Fusion Proteins; D050396:TRPP Cation Channels
chemical_list: D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D011506:Proteins; D011993:Recombinant Fusion Proteins; D050396:TRPP Cation Channels