Different organizational states of fodrin in cultured MDCK cells are induced by treatment with low pH, calmodulin antagonist TFP, and tumor promoter PMA.
We have investigated the molecular mechanisms underlying dynamic organization of the fodrin network by treating the epithelial MDCK cells with various agents affecting intracellular pH, intracellular calcium ion concentration, intracellular calmodulin, and protein kinase C (PKC) activity. Elevation of intracellular calcium level by A23187 or treatment with trifluoperazine (TFP), a calmodulin inhibitor, did not have any drastic effect on the fodrin distribution as judged by immunofluorescence microscopy. A long-term incubation with phorbol-12-myristate-13-acetate (PMA), a protein kinase C activator, in contrast, released fodrin from the lateral walls of the MDCK cells, leading to a diffuse cytoplasmic distribution. TFP, along with PMA, accelerated destabilization of the fodrin skeleton. Treatment with TFP alone rapidly released the cells from the substratum, which, however, could be prevented by PMA. We have previously shown that lowering of intracellular pH (< 6.5) leads to a removal of fodrin from its basolateral residence (Eskelinen et al., 1992) and that this translocation is reversed upon returning normal pH. We now show that the rebuilding of the membrane skeleton can be prevented if TFP is added to the acidified cells. Moreover, in TFP-treated acidified cells, fodrin shows a clusterlike organization similar to that observed in resting lymphocytes. We also noticed that interconversions between these different organizational states of fodrin are independent of the intracellular calcium concentration. Thus manipulation of the intracellular pH and treatment with TFP and PMA reveals different organizational states of the fodrin skeleton. This suggests that fodrin may participate in PMA-, TFP- and pH-sensitive signal transduction pathways.
Huotari
V
V
Biocenter, University of Oulu, Finland.
Sormunen
R
R
Lehto
V P
VP
Eskelinen
S
S
D000143:Acids / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000037:antagonists & inhibitors*; D002273:Carcinogens / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism / Q000502:physiology*; D002460:Cell Line; D004338:Drug Combinations; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000166:cytology; D008565:Membrane Proteins / Q000378:metabolism / Q000502:physiology; D008840:Microfilament Proteins / Q000378:metabolism / Q000502:physiology*; D012016:Reference Values; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014018:Tissue Distribution; D014268:Trifluoperazine / Q000494:pharmacology*; D013049:Spectrin
D000143:Acids; D002118:Calcium; D002147:Calmodulin; D002273:Carcinogens; D002352:Carrier Proteins; D004338:Drug Combinations; D008565:Membrane Proteins; D008840:Microfilament Proteins; D013755:Tetradecanoylphorbol Acetate; D014268:Trifluoperazine; D013049:Spectrin