Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1989 Sep 1;262(2):569–574. doi: 10.1042/bj2620569

Presence of haemin-controlled eIF-2 alpha kinases in both undifferentiated and differentiating mouse erythroleukaemia cells.

T F Sarre 1
PMCID: PMC1133306  PMID: 2803269

Abstract

In rabbit reticulocytes, globin synthesis is regulated by a haemin-controlled translational inhibitor (HCI) which acts by phosphorylating the alpha-subunit of eukaryotic initiation factor 2 (eIF-2). With purified eIF-2 as substrate, haemin-controlled eIF-2 alpha kinases could be partially purified from cultured mouse erythroleukaemia cells (MEL cells), which can be induced in vivo to erythroid differentiation. The eIF-2 alpha kinases from both uninduced and induced MEL cells are clearly distinct from the double-stranded-RNA-activated eIF-2 alpha kinase described for many mammalian cell types. A rough quantitative estimation indicates that, on a per-cell basis, induced MEL cells contain the same amount of haemin-controlled eIF-2 alpha kinase activity as rabbit reticulocytes, whereas uninduced MEL cells contain about one-tenth as much. As to their chromatographic behavior on CM-Sephadex and DEAE-cellulose and their sensitivity towards physiological concentrations of haemin (5-10 microM), the eIF-2 alpha kinases from MEL cells are indistinguishable from HCI. They differ from HCI with respect to their response towards activating stimuli such as prolonged incubation at 37 degrees C or brief exposure to the thiol reagent N-ethylmaleimide.

Full text

PDF
569

Images in this article

571Fig. 1.
on p.571
571Fig. 2.
on p.571
572Fig. 3.
on p.572
573Fig. 4.
on p.573
573Fig. 5.
on p.573

Selected References

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

  1. Austin S. A., Clemens M. J. The effects of haem on translational control of protein synthesis in cell-free extracts from fed and lysine-derived Ehrlich ascites tumour cells. Eur J Biochem. 1981 Jul;117(3):601–607. doi: 10.1111/j.1432-1033.1981.tb06380.x. [DOI] [PubMed] [Google Scholar]
  2. Bader M., Sarre T. F. A (re)initiation-dependent cell-free protein-synthesis system from mouse erythroleukemia cells. Eur J Biochem. 1986 Nov 17;161(1):103–109. doi: 10.1111/j.1432-1033.1986.tb10129.x. [DOI] [PubMed] [Google Scholar]
  3. Beuzard Y., London I. M. The effects of hemin and double-stranded RNA on alpha and beta globin synthesis in reticulocyte and Krebs II ascites cell-free systems and the relationship of these effects to an initiation factor preparation. Proc Natl Acad Sci U S A. 1974 Jul;71(7):2863–2866. doi: 10.1073/pnas.71.7.2863. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  5. Clemens M. J., Galpine A., Austin S. A., Panniers R., Henshaw E. C., Duncan R., Hershey J. W., Pollard J. W. Regulation of polypeptide chain initiation in Chinese hamster ovary cells with a temperature-sensitive leucyl-tRNA synthetase. Changes in phosphorylation of initiation factor eIF-2 and in the activity of the guanine nucleotide exchange factor GEF. J Biol Chem. 1987 Jan 15;262(2):767–771. [PubMed] [Google Scholar]
  6. De Benedetti A., Baglioni C. Activation of hemin-regulated initiation factor-2 kinase in heat-shocked HeLa cells. J Biol Chem. 1986 Jan 5;261(1):338–342. [PubMed] [Google Scholar]
  7. Delaunay J., Ranu R. S., Levin D. H., Ernst V., London I. M. Characterization of a rat liver factor that inhibits initiation of protein synthesis in rabbit reticulocyte lysates. Proc Natl Acad Sci U S A. 1977 Jun;74(6):2264–2268. doi: 10.1073/pnas.74.6.2264. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Duncan R. F., Hershey J. W. Translational repression by chemical inducers of the stress response occurs by different pathways. Arch Biochem Biophys. 1987 Aug 1;256(2):651–661. doi: 10.1016/0003-9861(87)90622-9. [DOI] [PubMed] [Google Scholar]
  9. Farrell P. J., Balkow K., Hunt T., Jackson R. J., Trachsel H. Phosphorylation of initiation factor elF-2 and the control of reticulocyte protein synthesis. Cell. 1977 May;11(1):187–200. doi: 10.1016/0092-8674(77)90330-0. [DOI] [PubMed] [Google Scholar]
  10. Galabru J., Hovanessian A. Autophosphorylation of the protein kinase dependent on double-stranded RNA. J Biol Chem. 1987 Nov 15;262(32):15538–15544. [PubMed] [Google Scholar]
  11. Grosfeld H., Ochoa S. Purification and properties of the double-stranded RNA-activated eukaryotic initiation factor 3 kinase from rabbit reticulocytes. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6526–6530. doi: 10.1073/pnas.77.11.6526. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gross M., Kaplansky D. A. Differential effect of Mn2+ on the hemin-controlled translational repressor and the double-stranded RNA-activated inhibitor. Biochim Biophys Acta. 1983 Aug 2;740(3):255–263. doi: 10.1016/0167-4781(83)90134-3. [DOI] [PubMed] [Google Scholar]
  13. Gross M., Rabinovitz M. Control of globin synthesis by hemin: factors influencing formation of an inhibitor of globin chain initiation in reticulocyte lysates. Biochim Biophys Acta. 1972 Dec 6;287(2):340–352. doi: 10.1016/0005-2787(72)90383-8. [DOI] [PubMed] [Google Scholar]
  14. Jackson R. J., Hunt T. Preparation and use of nuclease-treated rabbit reticulocyte lysates for the translation of eukaryotic messenger RNA. Methods Enzymol. 1983;96:50–74. doi: 10.1016/s0076-6879(83)96008-1. [DOI] [PubMed] [Google Scholar]
  15. Ochoa S. Regulation of protein synthesis initiation in eucaryotes. Arch Biochem Biophys. 1983 Jun;223(2):325–349. doi: 10.1016/0003-9861(83)90598-2. [DOI] [PubMed] [Google Scholar]
  16. Orkin S. H., Harosi F. I., Leder P. Differentiation in erythroleukemic cells and their somatic hybrids. Proc Natl Acad Sci U S A. 1975 Jan;72(1):98–102. doi: 10.1073/pnas.72.1.98. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Pain V. M. Initiation of protein synthesis in mammalian cells. Biochem J. 1986 May 1;235(3):625–637. doi: 10.1042/bj2350625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Pinphanichakarn P., Kramer G., Hardesty B. Partial purification and characterization of a translational inhibitor from Friend leukemia cells. J Biol Chem. 1977 Mar 25;252(6):2106–2112. [PubMed] [Google Scholar]
  19. Proud C. G., Pain V. M. Purification and phosphorylation of initiation factor eIF-2 from rabbit skeletal muscle. FEBS Lett. 1982 Jun 21;143(1):55–59. doi: 10.1016/0014-5793(82)80273-1. [DOI] [PubMed] [Google Scholar]
  20. Ranu R. S. Isolation of a translational inhibitor from wheat germ with protein kinase activity that phosphorylates initiation factor eIF-2. Biochem Biophys Res Commun. 1980 Dec 16;97(3):1124–1132. doi: 10.1016/0006-291x(80)91492-8. [DOI] [PubMed] [Google Scholar]
  21. Reuben R. C., Rifkind R. A., Marks P. A. Chemically induced murine erythroleukemic differentiation. Biochim Biophys Acta. 1980 Sep 22;605(3):325–346. doi: 10.1016/0304-419x(80)90015-3. [DOI] [PubMed] [Google Scholar]
  22. Scorsone K. A., Panniers R., Rowlands A. G., Henshaw E. C. Phosphorylation of eukaryotic initiation factor 2 during physiological stresses which affect protein synthesis. J Biol Chem. 1987 Oct 25;262(30):14538–14543. [PubMed] [Google Scholar]
  23. Staehelin T., Erni B., Schreier M. H. Purification and characterization of seven initiation factors for mammalian protein synthesis. Methods Enzymol. 1979;60:136–165. doi: 10.1016/s0076-6879(79)60013-7. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES