Role of the prostanoid FP receptor in action potential generation and phenotypic transformation of NRK fibroblasts.
By using an shRNA approach to knockdown the expression of the prostaglandin (PG)-F(2alpha) receptor (FP-R), the role of PGF(2alpha) in the process of phenotypic transformation of normal rat kidney (NRK) fibroblasts has been studied. Our data show that PGF(2alpha) up-regulates Cox-2 expression both at the mRNA and protein level, indicating that activation of FP-R in NRK fibroblasts induces a positive feedback loop in the production PGF(2alpha). Knockdown of FP-R expression fully impaired the ability of PGF(2alpha) to induce a calcium response and subsequent depolarization in NRK cells. However, these cells could still undergo phenotypic transformation when treated with a combination of EGF and retinoic acid, but in contrast to the wild-type cells, this process was not accompanied by a membrane depolarization to -20 mV. Knockdown of FP-R expression also impaired the spontaneous firing of calcium action potentials by density-arrested NRK cells. These data show that a membrane depolarization is not a prerequisite for the acquisition of a transformed phenotype. Furthermore, our data provide the first direct evidence that activity of PGF(2alpha) by putative pacemaker cells underlies the generation of calcium action potentials in NRK monolayers.
Almirza
W H
WH
Department of Cell Biology, Faculty of Science, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands.
Dernison
M M
MM
Peters
P H J
PH
van Zoelen
E J J
EJ
Theuvenet
A P R
AP
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D002452:Cell Count; D002461:Cell Line, Transformed; D049109:Cell Proliferation / Q000187:drug effects; D002471:Cell Transformation, Neoplastic / Q000187:drug effects; D051545:Cyclooxygenase 1 / Q000096:biosynthesis; D051546:Cyclooxygenase 2 / Q000096:biosynthesis; D015237:Dinoprost / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects; D004790:Enzyme Induction / Q000187:drug effects; D005347:Fibroblasts / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000096:biosynthesis; D010641:Phenotype; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D011982:Receptors, Prostaglandin / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014212:Tretinoin / Q000494:pharmacology
D008565:Membrane Proteins; D034741:RNA, Small Interfering; D011982:Receptors, Prostaglandin; D016212:Transforming Growth Factor beta; C034671:prostaglandin F2alpha receptor; D014212:Tretinoin; D015237:Dinoprost; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; C496513:Ptgs1 protein, rat; C489691:Ptgs2 protein, rat