Relevance of the palatal protein kinase A pathway to the pathogenesis of cleft palate by secalonic acid D in mice.
Secalonic acid-D (SAD) is a teratogenic mycotoxin inducing cleft palate (CP) in the offspring of the exposed mice by reducing palatal shelf size secondary to reduced proliferation of the palatal mesenchymal (PM) cells. Co-administration of dimethylsulfoxide (DMSO) reversed the CP-inducing effect of SAD. Although SAD has been shown to affect both protein kinases A (PKA) and C (PKC) pathways, the relevance of each of these pathways to its CP induction is unknown. The present studies were designed to test the hypothesis that the protective effect of DMSO is mediated by its specific reversal of the effect(s) of SAD on one of these two pathways using ELISA-based activity assays, Western blot analysis, electrophoretic mobility shift assays (EMSA), and murine embryonic PM (MEPM) cell growth in culture. Within the PKA pathway, SAD inhibited the activity of the catalytic subunit of PKA and its migration into the nucleus, elevated phosphorylated cyclic AMP (cAMP) response element (CRE)-binding protein (pCREB) level, and reduced the binding of CREB to CRE. In the PKC pathway, SAD reduced the activity of PKC and the binding of transcription factors (TF) to 12-O-tetradecanoate-13 phorbol acetate-response element (TRE). SAD also inhibited MEPM cell growth and the expression of the CRE- and TRE-containing gene, proliferating cell nuclear antigen (PCNA). Reversal, by DMSO, of the effects of SAD on MEPM cell growth, on PCNA expression and on all components of the PKA, but not of PKC, pathway suggests that the perturbation of the PKA pathway by SAD is relevant to its induction of CP in mice.
Dhulipala
Vamsidhara C
VC
Department of Veterinary Biomedical Sciences, University of Missouri, 1600 East Rollins Street, Columbia, MO 65211, USA.
Hanumegowda
Umesh M
UM
Balasubramanian
Ganesh
G
Reddy
Chada S
CS
D000818:Animals; D015153:Blotting, Western; D002384:Catalysis; D002455:Cell Division / Q000187:drug effects; D002467:Cell Nucleus / Q000378:metabolism; D002972:Cleft Palate / Q000139:chemically induced* / Q000201:enzymology* / Q000473:pathology; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D024202:Electrophoretic Mobility Shift Assay; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D051379:Mice; D010159:Palate / Q000166:cytology / Q000201:enzymology*; D010766:Phosphorylation; D011247:Pregnancy; D018809:Proliferating Cell Nuclear Antigen / Q000096:biosynthesis / Q000235:genetics; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D013723:Teratogens / Q000633:toxicity*; D014157:Transcription Factors / Q000378:metabolism; D044004:Xanthones / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000633:toxicity*
D017362:Cyclic AMP Response Element-Binding Protein; D004791:Enzyme Inhibitors; D018809:Proliferating Cell Nuclear Antigen; D013723:Teratogens; D014157:Transcription Factors; D044004:Xanthones; C009218:secalonic acid; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D004121:Dimethyl Sulfoxide