Activation of Mitogen-Activated Protein kinase/ERK-2 in Phytohaemagglutin in Blasts by Recombinant interleukin-2: Contrasting Features With CD3 Activation

Immunology. 1993 May;79(1):112-8.

Abstract

We investigated activation of mitogen-activated protein (MAP) kinase, also known as microtubule associated protein-2 kinase (MAP-2K), by recombinant interleukin-2 (rIL-2) in phytohaemagglutinin (PHA)-induced peripheral blood lymphoblasts (PBL). MAP-kinase activation has been implicated in growth of lymphocytes and other cell types. Enzyme activity was purified from cell lysates by ion-exchange chromatography and activity measured by the ability to phosphorylate the substrates MAP-2 and myelin basic protein peptide (APRTPGGRR) in vitro. Recombinant IL-2 stimulated a variable (two-to 10-fold) and evanescent MAP-2K response which was dose dependent over the range 0-50 U/ml. In contrast to MAP-kinase activation by the CD3 receptor, activation by the IL-2 receptor (IL-2R) proceeded independently from protein kinase C (PKC) and extracellular-free Ca2+. MAP-kinase activation by CD3 involves an activation cascade which depends on Ca2+ influx and PKC activation. These events culminate in tyrosine phosphorylation and activation of MAP kinase. Recombinant IL-2 induced tyrosine phosphorylation of several intracellular proteins, including a 40,000 MW substrate which co-electrophoresed with ERK-2 on SDS-PAGE. The ERK-2 gene encodes a 41,000 MW MAP-2K and is subject to regulation by a variety of mitogens and growth factors in lymphocytes and non-lymphoid cells. MAP-kinase activation by rIL-2 was abrogated when PHA blasts were pretreated with the tyrosine protein kinase (TPK) inhibitor, methyl-2,5-dihydroxy-cinnamate. Although the TPK, p56lck, has been implicated in the activation of MAP kinase and the function of IL-2R, we found no mobility shift from a 56,000 to a 60,000 MW position as seen during PKC activation. Together these data suggest that tyrosine phosphorylation is critical to IL-2-mediated signal transduction and that MAP kinase is one of the cellular intermediates involved in this pathway.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • CD3 Complex / immunology*
  • Calcium-Calmodulin-Dependent Protein Kinases
  • Cells, Cultured
  • Humans
  • Immunoblotting
  • Interleukin-2 / immunology*
  • Mitogen-Activated Protein Kinase 1
  • Phosphorylation
  • Phytohemagglutinins / immunology
  • Protein Kinases / blood*
  • Protein-Serine-Threonine Kinases / blood
  • Recombinant Proteins / immunology
  • T-Lymphocytes / enzymology
  • T-Lymphocytes / immunology*
  • Tyrosine / blood

Substances

  • CD3 Complex
  • Interleukin-2
  • Phytohemagglutinins
  • Recombinant Proteins
  • Tyrosine
  • Protein Kinases
  • Protein-Serine-Threonine Kinases
  • Calcium-Calmodulin-Dependent Protein Kinases
  • Mitogen-Activated Protein Kinase 1