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Calcitonin acutely increases tyrosyl-phosphorylation of proteins in human osteosarcoma (SaOS-2) cells

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Abstract

In order to test the hypothesis that salmon calcitonin has direct effects to modulate tyrosyl-protein phosphorylation in human osteosarcoma cells, SaOS-2 cells (with very high steady-state levels of skeletal alkaline phosphatase) were exposed to calcitonin, in duplicate serum-free cultures, at concentrations ranging from 10-13 to 10-9 mol/liter, for 0–60 minutes at 37°C. Phospho-tyrosyl proteins were identified by autoradiography of Western blots after incubation with 125I-labeled antiphosphotyrosine antibodies (or with unlabeled antibodies and 125I-labeled protein A) and quantitated by laser densitometry. The results of these studies revealed (1) time-dependent effects of salmon calcitonin (sCt) (at 3×10-12 mol/liter) to increase the level of tyrosyl-phosphorylation of at least six proteins, with apparent molecular weights of 20, 25, 27, 41, 48, and 135 kD (P<0.05 for each); and (2) dose-dependent effects of sCt (during 15 minutes of exposure) to increase the level of tyrosyl-phosphorylation of at least 10 proteins with apparent molecular weights of 19, 20, 27, 35, 41, 102, 135, 195, 220, and 244 kD (P<0.05 for each). A supplementary study of calcitonin effects on tyrosyl-protein phosphorylation in a subpopulation of SaOS-2 cells with very low steady-state levels of skeletal alkaline activity revealed similar responses—time and dose-dependent increases in the tyrosyl-phosphorylation of at least seven proteins with apparent molecular weights of 44, 48, 57, 62, 101, 244, and 280 kD (P<0.05 for each). Together, these studies demonstrate that sCt can have direct effects to modulate the level of tyrosyl-protein phosphorylation in human osteosarcoma cells, presumably by activation of tyrosyl-kinase activity and/or inhibition of phospho-tyrosyl-protein phosphatase activity.

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Authors and Affiliations

  1. Department of Medicine andf Biochemistry, Loma Linda University, Loma Linda, California, USA

    S. L. Hall, K. -H. William Lau & J. R. Farley

  2. Research Service (151), Jerry L. Pettis Memorial Veterans' Hospital, 11201 Benton Street, 92357, Loma Linda, California, USA

    V. Nicolas, K. -H. William Lau & J. R. Farley

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  1. A. Thomas
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  2. S. L. Hall
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  3. V. Nicolas
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  4. K. -H. William Lau
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  5. J. R. Farley
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Thomas, A., Hall, S.L., Nicolas, V. et al. Calcitonin acutely increases tyrosyl-phosphorylation of proteins in human osteosarcoma (SaOS-2) cells. Calcif Tissue Int 56, 268–273 (1995). https://doi.org/10.1007/BF00318045

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  • DOI: https://doi.org/10.1007/BF00318045

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