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The significance of Ser 1029 of the heat‐stable enterotoxin receptor (STaR): Relation of STa‐mediated guanylyl cyclase activation and signaling by phorbol myristate acetate
Author(s) -
Ian Padilla Philip
Publication year - 1996
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(96)00284-0
Subject(s) - phosphorylation , chemistry , transfection , amino acid , microbiology and biotechnology , mutant , protein kinase c , immunoprecipitation , signal transduction , receptor , phorbol , biochemistry , biology , gene
To characterize Ser 1029 in STaR at a consensus sequence of phosphorylation site by PKC, two mutants of mS1029A with replacement of Ser 1029 to Ala 1029 and CΔ1029 lacking 22 amino acids including Ser 1029 were prepared. Preincubation of the wild type‐STaR (wt‐STaR) transfectant with 1 μM PMA caused additional STa‐mediated guanylyl cyclase (GC) activation compared to control, whereas the mS1029A‐ and CΔ1029‐transfected cells did not show a similar enhanced GC activation by PMA. After metabolic labeling with [ 32 P]phosphate, transfected cells with wt‐STaR and mutants were incubated with 1 μM PMA. Subsequent 32 P‐radiolabeled proteins were immunoprecipitated using anti‐STaR antibody, and analyzed by autoradiography after separation on SDS‐PAGE. The immunoprecipitated wt‐STaR but not mS1029A and CΔ1029 had a significant radioactivity. These results suggest that the effect of PMA on wt‐STaR transfectants may be caused by phosphorylation of Ser 1029 . The CΔ1012 mutant, with further truncation (Gln 1012 ‐Phe 1050 ) of the carboxy terminus, did not show STa‐mediated GC activation. Based on these data, these 17 amino acids (Gln 1012 ‐Ala 1028 ), essential for signaling of GC activation by STa, have an abundance of basic amino acids which might be functionally influenced by phosphorylation of Ser 1029 .