Activation of the Ca2+ message system by parathyroid hormone is dependent on the cell cycle.
To verify whether the heterogeneous intracellular calcium ([Ca2+]i) responses to PTH observed in the UMR 106-01 osteogenic sarcoma cells are secondary to cell cycle asynchrony or to genotypic differences within the population, we synchronized cell monolayers at the G1/S boundary using a sequential thymidine-aphidicolin block. Video image analysis of fura-2-loaded cells revealed that PTH (10(-7) M) induced transient increases of [Ca2+]i preferentially in cells in S phase (82% response frequency, n = 63; 286 +/- 33% of baseline, n = 29), whereas cells in G1 phase responded poorly to PTH (10% response frequency, n = 51; 140 +/- 8% of baseline, n = 5). In contrast, cell exposure to 2% fetal calf serum was followed by [Ca2+]i transients in 83% (n = 42) of cells in G1 phase, but in only 25% (n = 63) of cells in S phase, with similar response amplitude. Hormonal responsiveness was heterogeneous in small clones obtained from single UMR 106-01 cells, with response frequency similar to that observed in nonsynchronized cultures. Pretreatment with either La3+, nifedipine, or pertussis toxin reduced both frequency and amplitude of PTH response in S phase to levels close to G1 phase, whereas there was no significant difference in inositol trisphosphate generated by PTH stimulation in either phase. Therefore, the heterogeneous [Ca2+]i responses of UMR 106-01 cells to hormonal stimulation is dependent on the phase of the cell cycle, rather than on genotypic heterogeneity. The switch from the G1 to the S phase mode of response is driven by active coupling between the PTH receptor and a Ca2+ channel through a pertussis toxin-sensitive G protein.
Bizzarri
C
C
Division of Endocrinology and Bone and Mineral Diseases, Jewish Hospital, Washington University Medical Center, St. Louis, Missouri 63110.
Civitelli
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R
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002453:Cell Cycle; D000242:Cyclic AMP / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D005312:Fetal Blood; D007295:Inositol Phosphates / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism; D009994:Osmolar Concentration; D010281:Parathyroid Hormone / Q000502:physiology*; D037342:Pertussis Toxin; D015290:Second Messenger Systems; D014407:Tumor Cells, Cultured; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D002121:Calcium Channel Blockers; D007295:Inositol Phosphates; D010281:Parathyroid Hormone; D010566:Virulence Factors, Bordetella; D004247:DNA; D000242:Cyclic AMP; D037342:Pertussis Toxin; D002118:Calcium