caa a22 4500 477120423 CHVBK 20180405111635.0 cr unu---uuuuu 170330e19961201xx s 000 0 eng 10.1007/BF00229472 doi (NATIONALLICENCE)springer-10.1007/BF00229472 Phosphorylation and activation of the intestinal guanylyl cyclase receptor for Escherichia coli heat-stable toxin by protein kinase C [Elektronische Daten] [John Crane, Kathryn Shanks] The heat-stable enterotoxin STa of E. coli causes diarrhea by binding to and stimulating intestinal membrane-bound guanylyl cyclase, triggering production of cyclic GMP. Agents which stimulate protein kinase C (PKC), including phorbol esters, synergistically enhance STa effects on cGMP and secretion. We investigated whether PKC causes phosphorylation of the STa receptor in vivo and in vitro. Immunoprecipitation of the STa receptor-guanylyl cyclase was carried out from extracts of T84 colon cells metabolically labelled with [32P]-phosphate using polyclonal anti-STa receptor antibody. The STa receptor was phosphorylated in its basal state, and 32P content in the 150 kDa holoreceptor band increased 2-fold in cells exposed to phorbol ester for 1 h. In vitro, immunopurified STa receptor was readily phosphorylated by purified rat brain PKC. Phosphorylation was inhibited 40% by 5 μM of a synthetic peptide corresponding to the sequence around Ser1029 of the STa receptor, a site previously proposed as a potential PKC phosphorylation site. Treatment of the immunopurified STaR/GC with purified PKC increased STa-stimulated guanylyl cyclase activity 2-fold. We conclude that PKC phosphorylates and activates the STa receptor/guanylyl cyclase in vitro and in vivo; Ser1029 of the STaR/GC remains a candidate phosphorylation site by PKC. Kluwer Academic Publishers, 1996 guanylyl cyclase (or guanylate cyclase) nationallicence protein kinase C nationallicence heat-stable enterotoxin nationallicence Escherichia coli nationallicence infectious diarrhea nationallicence STa : the heat-stable enterotoxin of E. coli, which has also been called ST-I and STp. The 18 amino acid variant was used throughout nationallicence PBS : phosphate-buffered saline nationallicence PDB : 4-β-12, 13-phorbol dibutyrate nationallicence ANP : atrial natriuretic peptide nationallicence STaR/GC : STa receptor/guanylyl cyclase, also called GC-C nationallicence PKC : protein kinase C nationallicence Crane John Division of Infectious Diseases, State University of New York at Buffalo, Room 109 Medical Research Bldg., 3435 Main St., 14214, Buffalo, NY, USA aut Shanks Kathryn Division of Infectious Diseases, State University of New York at Buffalo, Room 109 Medical Research Bldg., 3435 Main St., 14214, Buffalo, NY, USA aut Molecular and Cellular Biochemistry Kluwer Academic Publishers; http://www.wkap.nl 165/2(1996-12-01), 111-120 0300-8177 165:2<111 1996 165 11010 https://doi.org/10.1007/BF00229472 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/BF00229472 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Crane John Division of Infectious Diseases, State University of New York at Buffalo, Room 109 Medical Research Bldg., 3435 Main St., 14214, Buffalo, NY, USA aut NATIONALLICENCE 700 1- Shanks Kathryn Division of Infectious Diseases, State University of New York at Buffalo, Room 109 Medical Research Bldg., 3435 Main St., 14214, Buffalo, NY, USA aut NATIONALLICENCE 773 0- Molecular and Cellular Biochemistry Kluwer Academic Publishers; http://www.wkap.nl 165/2(1996-12-01), 111-120 0300-8177 165:2<111 1996 165 11010 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer