Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1989 Jun;86(12):4525–4529. doi: 10.1073/pnas.86.12.4525

Effect of cell spreading on cytoplasmic pH in normal and transformed fibroblasts.

M A Schwartz 1, G Both 1, C Lechene 1
PMCID: PMC287303  PMID: 2734302

Abstract

Growth of normal fibroblasts requires an adhesive substratum on which to spread, whereas transformed cells can grow in suspension. Since an alkaline cytoplasm has been shown to be required for growth, we measured cytoplasmic pH in individual cells as a function of spreading. The degree of spreading was controlled by coating tissue culture plastic with varying amounts of the nonadhesive polymer polyHEMA. Completely round BALB/c 3T3 cells were 0.15 pH unit more acidic than spread cells. In short-term experiments, cells were treated with the peptide Gly-Arg-Gly-Asp-Ser-Pro to induce rounding or by plating on fibronectin to induce spreading. When cells were induced to change shape, pH changed rapidly and reversibly. All of the anchorage-dependent cell lines tested behaved similarly, but 3T3 cells transformed by the plasma membrane oncogene src or ras were able to maintain a relatively alkaline pH even when completely round. Cells transformed by the nuclear oncogene myc behaved like normal cells. Our results suggest that the requirement for spreading may in part be mediated by cytoplasmic pH. Anchorage-independent growth due to oncogenes that localize to the plasma membrane is associated with loss of this control mechanism.

Full text

PDF
4525

Images in this article

4527Image
on p.4527
4527Image
on p.4527
4527Image
on p.4527
4527Image
on p.4527
4527Image
on p.4527
4528Image
on p.4528
4528Image
on p.4528

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aggeler J., Frisch S. M., Werb Z. Changes in cell shape correlate with collagenase gene expression in rabbit synovial fibroblasts. J Cell Biol. 1984 May;98(5):1662–1671. doi: 10.1083/jcb.98.5.1662. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Armelin H. A., Armelin M. C., Kelly K., Stewart T., Leder P., Cochran B. H., Stiles C. D. Functional role for c-myc in mitogenic response to platelet-derived growth factor. Nature. 1984 Aug 23;310(5979):655–660. doi: 10.1038/310655a0. [DOI] [PubMed] [Google Scholar]
  3. Ben-Ze'ev A., Amsterdam A. In vitro regulation of granulosa cell differentiation. Involvement of cytoskeletal protein expression. J Biol Chem. 1987 Apr 15;262(11):5366–5376. [PubMed] [Google Scholar]
  4. Ben-Ze'ev A., Farmer S. R., Penman S. Protein synthesis requires cell-surface contact while nuclear events respond to cell shape in anchorage-dependent fibroblasts. Cell. 1980 Sep;21(2):365–372. doi: 10.1016/0092-8674(80)90473-0. [DOI] [PubMed] [Google Scholar]
  5. Ben-Ze'ev A., Robinson G. S., Bucher N. L., Farmer S. R. Cell-cell and cell-matrix interactions differentially regulate the expression of hepatic and cytoskeletal genes in primary cultures of rat hepatocytes. Proc Natl Acad Sci U S A. 1988 Apr;85(7):2161–2165. doi: 10.1073/pnas.85.7.2161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Benecke B. J., Ben-Ze'ev A., Penman S. The control of mRNA production, translation and turnover in suspended and reattached anchorage-dependent fibroblasts. Cell. 1978 Aug;14(4):931–939. doi: 10.1016/0092-8674(78)90347-1. [DOI] [PubMed] [Google Scholar]
  7. Bishop J. M. The molecular genetics of cancer. Science. 1987 Jan 16;235(4786):305–311. doi: 10.1126/science.3541204. [DOI] [PubMed] [Google Scholar]
  8. Blum J. L., Zeigler M. E., Wicha M. S. Regulation of rat mammary gene expression by extracellular matrix components. Exp Cell Res. 1987 Dec;173(2):322–340. doi: 10.1016/0014-4827(87)90274-6. [DOI] [PubMed] [Google Scholar]
  9. Bright G. R., Fisher G. W., Rogowska J., Taylor D. L. Fluorescence ratio imaging microscopy: temporal and spatial measurements of cytoplasmic pH. J Cell Biol. 1987 Apr;104(4):1019–1033. doi: 10.1083/jcb.104.4.1019. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Doppler W., Jaggi R., Groner B. Induction of v-mos and activated Ha-ras oncogene expression in quiescent NIH 3T3 cells causes intracellular alkalinisation and cell-cycle progression. Gene. 1987;54(1):147–153. doi: 10.1016/0378-1119(87)90357-x. [DOI] [PubMed] [Google Scholar]
  11. Folkman J., Moscona A. Role of cell shape in growth control. Nature. 1978 Jun 1;273(5661):345–349. doi: 10.1038/273345a0. [DOI] [PubMed] [Google Scholar]
  12. Freedman V. H., Shin S. I. Cellular tumorigenicity in nude mice: correlation with cell growth in semi-solid medium. Cell. 1974 Dec;3(4):355–359. doi: 10.1016/0092-8674(74)90050-6. [DOI] [PubMed] [Google Scholar]
  13. L'Allemain G., Franchi A., Cragoe E., Jr, Pouysségur J. Blockade of the Na+/H+ antiport abolishes growth factor-induced DNA synthesis in fibroblasts. Structure-activity relationships in the amiloride series. J Biol Chem. 1984 Apr 10;259(7):4313–4319. [PubMed] [Google Scholar]
  14. Land H., Parada L. F., Weinberg R. A. Tumorigenic conversion of primary embryo fibroblasts requires at least two cooperating oncogenes. Nature. 1983 Aug 18;304(5927):596–602. doi: 10.1038/304596a0. [DOI] [PubMed] [Google Scholar]
  15. Moolenaar W. H. Effects of growth factors on intracellular pH regulation. Annu Rev Physiol. 1986;48:363–376. doi: 10.1146/annurev.ph.48.030186.002051. [DOI] [PubMed] [Google Scholar]
  16. Musgrove E., Seaman M., Hedley D. Relationship between cytoplasmic pH and proliferation during exponential growth and cellular quiescence. Exp Cell Res. 1987 Sep;172(1):65–75. doi: 10.1016/0014-4827(87)90093-0. [DOI] [PubMed] [Google Scholar]
  17. Noda M., Selinger Z., Scolnick E. M., Bassin R. H. Flat revertants isolated from Kirsten sarcoma virus-transformed cells are resistant to the action of specific oncogenes. Proc Natl Acad Sci U S A. 1983 Sep;80(18):5602–5606. doi: 10.1073/pnas.80.18.5602. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Ober S. S., Pardee A. B. Intracellular pH is increased after transformation of Chinese hamster embryo fibroblasts. Proc Natl Acad Sci U S A. 1987 May;84(9):2766–2770. doi: 10.1073/pnas.84.9.2766. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Oliver J. M., Berlin R. D. Mechanisms that regulate the structural and functional architecture of cell surfaces. Int Rev Cytol. 1982;74:55–94. doi: 10.1016/s0074-7696(08)61169-9. [DOI] [PubMed] [Google Scholar]
  20. Perona R., Serrano R. Increased pH and tumorigenicity of fibroblasts expressing a yeast proton pump. Nature. 1988 Aug 4;334(6181):438–440. doi: 10.1038/334438a0. [DOI] [PubMed] [Google Scholar]
  21. Pierschbacher M. D., Ruoslahti E. Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule. Nature. 1984 May 3;309(5963):30–33. doi: 10.1038/309030a0. [DOI] [PubMed] [Google Scholar]
  22. Piwnica-Worms H., Saunders K. B., Roberts T. M., Smith A. E., Cheng S. H. Tyrosine phosphorylation regulates the biochemical and biological properties of pp60c-src. Cell. 1987 Apr 10;49(1):75–82. doi: 10.1016/0092-8674(87)90757-4. [DOI] [PubMed] [Google Scholar]
  23. Pouysségur J., Sardet C., Franchi A., L'Allemain G., Paris S. A specific mutation abolishing Na+/H+ antiport activity in hamster fibroblasts precludes growth at neutral and acidic pH. Proc Natl Acad Sci U S A. 1984 Aug;81(15):4833–4837. doi: 10.1073/pnas.81.15.4833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Soltoff S. P., Cragoe E. J., Jr, Mandel L. J. Amiloride analogues inhibit proximal tubule metabolism. Am J Physiol. 1986 May;250(5 Pt 1):C744–C747. doi: 10.1152/ajpcell.1986.250.5.C744. [DOI] [PubMed] [Google Scholar]
  25. Spiegelman B. M., Ginty C. A. Fibronectin modulation of cell shape and lipogenic gene expression in 3T3-adipocytes. Cell. 1983 Dec;35(3 Pt 2):657–666. doi: 10.1016/0092-8674(83)90098-3. [DOI] [PubMed] [Google Scholar]
  26. Thomas J. A., Buchsbaum R. N., Zimniak A., Racker E. Intracellular pH measurements in Ehrlich ascites tumor cells utilizing spectroscopic probes generated in situ. Biochemistry. 1979 May 29;18(11):2210–2218. doi: 10.1021/bi00578a012. [DOI] [PubMed] [Google Scholar]
  27. Tucker R. W., Butterfield C. E., Folkman J. Interaction of serum and cell spreading affects the growth of neoplastic and non-neoplastic fibroblasts. J Supramol Struct Cell Biochem. 1981;15(1):29–40. doi: 10.1002/jsscb.1981.380150104. [DOI] [PubMed] [Google Scholar]
  28. Zanetti N. C., Solursh M. Induction of chondrogenesis in limb mesenchymal cultures by disruption of the actin cytoskeleton. J Cell Biol. 1984 Jul;99(1 Pt 1):115–123. doi: 10.1083/jcb.99.1.115. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES