Membrane depolarization was required to induce DNA synthesis in murine macrophage cell line PU5-1.8.
The role of membrane potential (Em) on the initiation of DNA synthesis in murine macrophage cell line PU5-1.8 was investigated with fluorescent probes bis-oxonol and diS-C3-(5). Incubation of PU5-1.8 cells in high K(+)-HEPES buffer or with gramicidin at 37 degrees C for 1h that depolarized the membrane induced [3H]-thymidine incorporation and expression of early response gene such as c-myc and c-fos. When PU5-1.8 cells were treated with a number of agents including fetal calf serum (FCS), lipopolysaccharide (LPS), epidermal growth factor (EGF), N-formyl-methionyl-leucyl-phenylalanine (FMLP) and bradykinin (BK), only FCS caused DNA synthesis and membrane depolarization. Other agents had no effect on these events. The FCS-mediated DNA synthesis in PU5-1.8 cells was inhibited by clamping the membrane potential with valinomycin. Moreover, intracellular alkalinization induced by nigericin at pH 7.9, which is believed to be a permissive signal for mitogenesis, caused membrane depolarization. On the other hand, challenge of cells with phorbol 12-myristate 13 acetate (PMA) suppressed the K(+)-mediated DNA synthesis. However, the treatment of cells with PMA did not change the membrane potential but suppressed the gramicidin-mediated membrane depolarization. These observations suggest that there is a correlation between membrane depolarization and initiation of DNA synthesis in PU5-1.8 cells. PKC may be acting as a modulator in this transducing pathway.
Kong
S K
SK
Department of Biochemistry, Chinese University of Hong Kong, Shatin.
Suen
Y K
YK
Choy
Y M
YM
Fung
K P
KP
Lee
C Y
CY
D000818:Animals; D002460:Cell Line; D004247:DNA / Q000096:biosynthesis*; D005312:Fetal Blood / Q000502:physiology; D016762:Genes, fos; D016259:Genes, myc; D006863:Hydrogen-Ion Concentration; D008264:Macrophages / Q000378:metabolism*; D008564:Membrane Potentials; D051379:Mice; D008807:Mice, Inbred BALB C; D011188:Potassium / Q000494:pharmacology; D011493:Protein Kinase C / Q000502:physiology
D004247:DNA; D011493:Protein Kinase C; D011188:Potassium