Ca2+ and Zn2+ dependence of DNA synthesis in untransformed and in Ha-ras(val-12)-expressing NIH 3T3 cells

C J Back, L Sistonen, M O Enkvist, J E Heikkilä, K E Akerman

Research output: Contribution to journalArticleScientificpeer-review

13 Citations (Scopus)

Abstract

The role of Ca2+ and Zn2+ in the initiation of DNA synthesis in NIH 3T3 fibroblasts and c-Ha-ras(val-12) oncoprotein-expressing (NIH 3T3) cells has been studied. Entrapment of the Ca2+ chelator, BAPTA (30 microM), into the cells totally blocked a serum-induced rise in cytosolic free Ca2+ ([Ca2+]i) as determined with fura-2. Serum starvation for 24 h considerably reduced DNA synthesis in control NIH 3T3 fibroblasts. BAPTA treatment reduced serum-induced DNA synthesis and totally inhibited platelet-derived growth factor-induced DNA synthesis in these cells. DNA synthesis of the c-Ha-ras(val-12)-expressing fibroblasts was little affected by serum starvation and unaffected by entrapment of BAPTA into the cells. Intracellular Zn2+ was measured using the fluorescent probe TSQ in intact cells. As determined using image analysis the TSQ fluorescence was distributed throughout the cytoplasm and concentrated around the nucleus. The permeable Zn2+ chelator, TPEN, at a concentration of 10 microM, caused a maximal reduction in TSQ-available Zn2+. This concentration of TPEN totally blocked DNA synthesis both in control and c-Ha-ras(val-12)-expressing fibroblasts. Upon addition of 11 microM Zn2+ DNA synthesis was restored even after TPEN addition. [3H]Thymidine incorporation itself was also sensitive to TPEN treatment. The results suggest that c-Ha-ras(val-12)-induced proliferation is independent of changes in [Ca2+]i. A specific role of Zn2+ in c-Ha-ras-induced proliferation is unlikely since ras-expressing and control cells reacted similarly to Zn2+ deprivation. There seems to be a constant requirement for the presence of Zn2+ in cell proliferation.

Original languageEnglish
Pages (from-to)303-10
Number of pages8
JournalExperimental Cell Research
Volume208
Issue number1
DOIs
Publication statusPublished - Sept 1993
MoE publication typeA1 Journal article-refereed

Keywords

  • 3T3 Cells
  • Animals
  • Calcium/pharmacology
  • Cations, Divalent
  • Cell Cycle/drug effects
  • Cell Transformation, Neoplastic/metabolism
  • Chelating Agents/pharmacology
  • Cytosol/physiology
  • DNA/biosynthesis
  • DNA Replication/drug effects
  • In Vitro Techniques
  • Mice
  • Protein Biosynthesis
  • Proto-Oncogene Proteins p21(ras)/physiology
  • Transfection
  • Zinc/pharmacology

Fingerprint

Dive into the research topics of 'Ca2+ and Zn2+ dependence of DNA synthesis in untransformed and in Ha-ras(val-12)-expressing NIH 3T3 cells'. Together they form a unique fingerprint.

Cite this