Calcium receptor-induced serotonin secretion by parafollicular cells: role of phosphatidylinositol 3-kinase-dependent signal transduction pathways.
Elevation of extracellular Ca2+ (increase[Ca2+]e) stimulates the Ca2+ receptor (CaR) to induce secretion of 5-hydroxytryptamine (5-HT) from the calcium-sensing parafollicular (PF) cells. The CaR has been reported to couple to Galpha(q) with subsequent activation of protein kinase C-gamma (PKCgamma). We have identified a parallel transduction pathway in primary cultures of sheep PF cells by using a combinatorial approach in which we expressed adenoviral-encoded dominant-negative signaling proteins and performed in vitro kinase assays. The role of the CaR was established by expression of a dominant-negative CaR that eliminated calcium-induced 5-HT secretion but not secretion in response to KCl or phorbol esters. The calcium-induced secretion was inhibited by a dominant-negative p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3-K). PI3-K activity was also assayed using isoform-specific antibodies. The activity of p85/p110beta (PI3-Kbeta) immunocomplexes was elevated by increase[Ca2+]e and activated by Gbetagamma subunits. In addition, secretion of 5-HT was antagonized by the expression of a minigene encoding a peptide scavenger of Gbetagamma subunits (C-terminal fragment peptide of bovine beta-adrenergic receptor kinase). One target of PI3-K activity is phosphoinositide-dependent kinase-1 (PDK1), which in turn activated PKCzeta. Expression of a dominant-negative PKCzeta in PF cells reduced 5-HT secretion. Together, these observations establish that increase[Ca2+]e evokes 5-HT secretion from PF cells by stimulating both Galpha(q)- and Gbetagamma-signaling pathways downstream of the CaR. The betagamma cascade subsequently activates PI3-Kbeta-dependent signaling that is coupled to PDK1 and the downstream effector PKCzeta, and results in an increase in 5-HT release.
Liu
Kuo-peing
KP
Division of Neuroscience, New York State Psychiatric Institute, New York, New York 10032, USA.
Russo
Andrew F
AF
Hsiung
Shu-chi
SC
Adlersberg
Mella
M
Franke
Thomas F
TF
Gershon
Michael D
MD
Tamir
Hadassah
H
D064413:3-Phosphoinositide-Dependent Protein Kinases; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002469:Cell Separation; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000096:biosynthesis / Q000235:genetics; D005110:Extracellular Space / Q000378:metabolism; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D015870:Gene Expression; D005799:Genes, Dominant; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000096:biosynthesis / Q000235:genetics; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D012701:Serotonin / Q000378:metabolism*; D012756:Sheep; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013961:Thyroid Gland / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D014162:Transfection; D051552:beta-Adrenergic Receptor Kinases; D000099221:Protein Kinase C zeta
D064413:3-Phosphoinositide-Dependent Protein Kinases; D002118:Calcium; D002135:Calcium-Binding Proteins; D017868:Cyclic AMP-Dependent Protein Kinases; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020962:Heterotrimeric GTP-Binding Proteins; D007527:Isoenzymes; D010446:Peptide Fragments; D019869:Phosphatidylinositol 3-Kinases; D011493:Protein Kinase C; D017346:Protein Serine-Threonine Kinases; D021122:Protein Subunits; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D012701:Serotonin; D051552:beta-Adrenergic Receptor Kinases; D000099221:Protein Kinase C zeta