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Molecular cloning and expression of the major protein kinase C substrate of platelets

Nature volume 333, pages 470–473 (1988)Cite this article

Abstract

In platelets, agonists that stimulate phosphoinositide turnover cause the rapid phosphorylation of a protein of apparent relative molecular mass (Mr) 40–47,000, called P47, by protein kinase C (PKC)1–4. Diverse identities have been ascribed to P47 including lipocortin5, inositol 1,4,5-trisphosphate 5-phosphomonoesterase6, pyruvate dehydrogenase α subunit7 and an actin regulatory protein8. We have isolated human P47 clones by immunological screening of a λgtll complementary DNA library from HL-60 cells, a human promyelocytic leukaemia cell line9. P47 recom-binants thus identified hybridized to a 3.0 kilobase (kb) messenger RNA in mature white blood cell lines; the same mRNA was induced in HL-60 cells during differentiation. A 1,050 base pair (bp) open reading frame that could encode a protein of Mr 40,087 was confirmed by comparison with peptide sequences from platelet P47, and by expression of the putative recombinant P47 in E. coli and in vitro. The P47 sequence appears to have been conserved throughout vertebrate evolution, and is not similar to any other known sequence including human lipocortin and the α subunit of pyruvate dehydrogenase. The P47 protein contains a potential Ca2+-binding 'EF-hand' structure and a region that strongly resembles known PKC phosphorylation sites.

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Authors and Affiliations

  1. Department of Biochemistry, McMaster University, Hamilton, Ontario, Canada, L8N 3Z5

    Michael Tyers, Richard A. Rachubinski, Richard J. Haslam & Calvin B. Harley

  2. Department of Pathology, McMaster University, Hamilton, Ontario, Canada, L8N 3Z5

    Mary I. Stewart & Richard J. Haslam

  3. Smith Kline and French Laboratories, PO Box 1539, King of Prussia, Pennsylvania, 19406, USA

    Angela M. Varrichio & Robert G. L. Shorr

Authors
  1. Michael Tyers
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  2. Richard A. Rachubinski
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  4. Angela M. Varrichio
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  5. Robert G. L. Shorr
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  6. Richard J. Haslam
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  7. Calvin B. Harley
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Tyers, M., Rachubinski, R., Stewart, M. et al. Molecular cloning and expression of the major protein kinase C substrate of platelets. Nature 333, 470–473 (1988). https://doi.org/10.1038/333470a0

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