Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1996 Jun 15;316(Pt 3):865–871. doi: 10.1042/bj3160865

Protein kinase C isoforms epsilon, eta, delta and zeta in murine adipocytes: expression, subcellular localization and tissue-specific regulation in insulin-resistant states.

E U Frevert 1, B B Kahn 1
PMCID: PMC1217430  PMID: 8670164

Abstract

The Ca(2+)-insensitive protein kinase C (PKC) isoforms epsilon, eta, delta and zeta are possible direct downstream targets of phosphatidylinositol 3-kinase (P13-K), and might therefore be involved in insulin signalling. Although isoform-specific changes in PKC expression have been reported for skeletal muscle and liver in insulin-resistant states, little is known about these isoforms in adipocytes. Therefore we studied (1) expression and subcellular localization of these isoforms in murine adipocytes, (2) translocation of specific isoforms to membranes in response to treatment with insulin and phorbol 12-myristate 13-acetate (PMA) and (3) regulation of expression in insulin-resistant states. The PKC isoforms epsilon, eta, delta and zeta are expressed in adipocytes. Immunoreactivity for all isoforms is higher in the membranes than in the cytosol, but subcellular fractionation by differential centrifugation shows an isoform-specific distribution within the membrane fractions. PMA treatment of adipocytes induces translocation of PKC-epsilon and -delta from the cytosol to the membrane fractions. Insulin treatment does not alter the subcellular distribution of any of the isoforms. 3T3-L1 adipocytes express PKC-epsilon and -zeta, and PKC-epsilon expression increases with differentiation from preadipocytes to adipocytes. PKC-epsilon expression decreases in an adipose-specific and age/obesity-dependent manner in two insulin-resistant models, the brown-adipose-tissue-deficient mouse and db/db mouse compared with control mice. We conclude that, although none of the isoforms investigated seems to be activated by insulin, the decrease in PKC-epsilon expression might contribute to metabolic alterations in adipocytes associated with insulin resistance and obesity.

Full Text

The Full Text of this article is available as a PDF (369.0 KB).

Selected References

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

  1. Akita Y., Ohno S., Yajima Y., Konno Y., Saido T. C., Mizuno K., Chida K., Osada S., Kuroki T., Kawashima S. Overproduction of a Ca(2+)-independent protein kinase C isozyme, nPKC epsilon, increases the secretion of prolactin from thyrotropin-releasing hormone-stimulated rat pituitary GH4C1 cells. J Biol Chem. 1994 Feb 11;269(6):4653–4660. [PubMed] [Google Scholar]
  2. Blackshear P. J., Haupt D. M., Stumpo D. J. Insulin activation of protein kinase C: a reassessment. J Biol Chem. 1991 Jun 15;266(17):10946–10952. [PubMed] [Google Scholar]
  3. Bogoyevitch M. A., Parker P. J., Sugden P. H. Characterization of protein kinase C isotype expression in adult rat heart. Protein kinase C-epsilon is a major isotype present, and it is activated by phorbol esters, epinephrine, and endothelin. Circ Res. 1993 Apr;72(4):757–767. doi: 10.1161/01.res.72.4.757. [DOI] [PubMed] [Google Scholar]
  4. Cheatham B., Vlahos C. J., Cheatham L., Wang L., Blenis J., Kahn C. R. Phosphatidylinositol 3-kinase activation is required for insulin stimulation of pp70 S6 kinase, DNA synthesis, and glucose transporter translocation. Mol Cell Biol. 1994 Jul;14(7):4902–4911. doi: 10.1128/mcb.14.7.4902. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chida K., Sagara H., Suzuki Y., Murakami A., Osada S., Ohno S., Hirosawa K., Kuroki T. The eta isoform of protein kinase C is localized on rough endoplasmic reticulum. Mol Cell Biol. 1994 Jun;14(6):3782–3790. doi: 10.1128/mcb.14.6.3782. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Considine R. V., Caro J. F. Protein kinase C: mediator or inhibitor of insulin action? J Cell Biochem. 1993 May;52(1):8–13. doi: 10.1002/jcb.240520103. [DOI] [PubMed] [Google Scholar]
  7. Considine R. V., Nyce M. R., Allen L. E., Morales L. M., Triester S., Serrano J., Colberg J., Lanza-Jacoby S., Caro J. F. Protein kinase C is increased in the liver of humans and rats with non-insulin-dependent diabetes mellitus: an alteration not due to hyperglycemia. J Clin Invest. 1995 Jun;95(6):2938–2944. doi: 10.1172/JCI118001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cooper D. R., Watson J. E., Dao M. L. Decreased expression of protein kinase-C alpha, beta, and epsilon in soleus muscle of Zucker obese (fa/fa) rats. Endocrinology. 1993 Nov;133(5):2241–2247. doi: 10.1210/endo.133.5.8404676. [DOI] [PubMed] [Google Scholar]
  9. Cushman S. W., Salans L. B. Determinations of adipose cell size and number in suspensions of isolated rat and human adipose cells. J Lipid Res. 1978 Feb;19(2):269–273. [PubMed] [Google Scholar]
  10. Cushman S. W. Structure-function relationships in the adipose cell. I. Ultrastructure of the isolated adipose cell. J Cell Biol. 1970 Aug;46(2):326–341. doi: 10.1083/jcb.46.2.326. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. DOLE V. P. A relation between non-esterified fatty acids in plasma and the metabolism of glucose. J Clin Invest. 1956 Feb;35(2):150–154. doi: 10.1172/JCI103259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Farese R. V., Standaert M. L., Francois A. J., Ways K., Arnold T. P., Hernandez H., Cooper D. R. Effects of insulin and phorbol esters on subcellular distribution of protein kinase C isoforms in rat adipocytes. Biochem J. 1992 Nov 15;288(Pt 1):319–323. doi: 10.1042/bj2880319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Goodnight J. A., Mischak H., Kolch W., Mushinski J. F. Immunocytochemical localization of eight protein kinase C isozymes overexpressed in NIH 3T3 fibroblasts. Isoform-specific association with microfilaments, Golgi, endoplasmic reticulum, and nuclear and cell membranes. J Biol Chem. 1995 Apr 28;270(17):9991–10001. doi: 10.1074/jbc.270.17.9991. [DOI] [PubMed] [Google Scholar]
  14. Goodnight J., Kazanietz M. G., Blumberg P. M., Mushinski J. F., Mischak H. The cDNA sequence, expression pattern and protein characteristics of mouse protein kinase C-zeta. Gene. 1992 Dec 15;122(2):305–311. doi: 10.1016/0378-1119(92)90219-f. [DOI] [PubMed] [Google Scholar]
  15. Gschwendt M., Leibersperger H., Rincke G., Marks F. Immunological demonstration of epsilon PKC. Murine tissue distribution, ontogeny, cellular localization and translocation. FEBS Lett. 1991 Sep 23;290(1-2):115–118. doi: 10.1016/0014-5793(91)81239-5. [DOI] [PubMed] [Google Scholar]
  16. Hamann A., Flier J. S., Lowell B. B. Decreased brown fat markedly enhances susceptibility to diet-induced obesity, diabetes, and hyperlipidemia. Endocrinology. 1996 Jan;137(1):21–29. doi: 10.1210/endo.137.1.8536614. [DOI] [PubMed] [Google Scholar]
  17. Hashimoto Y., Osada S., Ohno S., Kuroki T. A Ca(2+)-independent protein kinase C, nPKC eta: its structure, distribution and possible function. Tohoku J Exp Med. 1992 Oct;168(2):275–278. doi: 10.1620/tjem.168.275. [DOI] [PubMed] [Google Scholar]
  18. Heidenreich K. A., Toledo S. P., Brunton L. L., Watson M. J., Daniel-Issakani S., Strulovici B. Insulin stimulates the activity of a novel protein kinase C, PKC-epsilon, in cultured fetal chick neurons. J Biol Chem. 1990 Sep 5;265(25):15076–15082. [PubMed] [Google Scholar]
  19. Hug H., Sarre T. F. Protein kinase C isoenzymes: divergence in signal transduction? Biochem J. 1993 Apr 15;291(Pt 2):329–343. doi: 10.1042/bj2910329. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. James D. E., Brown R., Navarro J., Pilch P. F. Insulin-regulatable tissues express a unique insulin-sensitive glucose transport protein. Nature. 1988 May 12;333(6169):183–185. doi: 10.1038/333183a0. [DOI] [PubMed] [Google Scholar]
  21. Kahn B. B. Facilitative glucose transporters: regulatory mechanisms and dysregulation in diabetes. J Clin Invest. 1992 May;89(5):1367–1374. doi: 10.1172/JCI115724. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kikkawa R., Haneda M., Uzu T., Koya D., Sugimoto T., Shigeta Y. Translocation of protein kinase C alpha and zeta in rat glomerular mesangial cells cultured under high glucose conditions. Diabetologia. 1994 Aug;37(8):838–841. doi: 10.1007/BF00404342. [DOI] [PubMed] [Google Scholar]
  23. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  24. Lehel C., Olah Z., Jakab G., Anderson W. B. Protein kinase C epsilon is localized to the Golgi via its zinc-finger domain and modulates Golgi function. Proc Natl Acad Sci U S A. 1995 Feb 28;92(5):1406–1410. doi: 10.1073/pnas.92.5.1406. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lowell B. B., S-Susulic V., Hamann A., Lawitts J. A., Himms-Hagen J., Boyer B. B., Kozak L. P., Flier J. S. Development of obesity in transgenic mice after genetic ablation of brown adipose tissue. Nature. 1993 Dec 23;366(6457):740–742. doi: 10.1038/366740a0. [DOI] [PubMed] [Google Scholar]
  26. Mischak H., Goodnight J. A., Kolch W., Martiny-Baron G., Schaechtle C., Kazanietz M. G., Blumberg P. M., Pierce J. H., Mushinski J. F. Overexpression of protein kinase C-delta and -epsilon in NIH 3T3 cells induces opposite effects on growth, morphology, anchorage dependence, and tumorigenicity. J Biol Chem. 1993 Mar 25;268(9):6090–6096. [PubMed] [Google Scholar]
  27. Mizuno K., Kubo K., Saido T. C., Akita Y., Osada S., Kuroki T., Ohno S., Suzuki K. Structure and properties of a ubiquitously expressed protein kinase C, nPKC delta. Eur J Biochem. 1991 Dec 18;202(3):931–940. doi: 10.1111/j.1432-1033.1991.tb16453.x. [DOI] [PubMed] [Google Scholar]
  28. Mochly-Rosen D. Localization of protein kinases by anchoring proteins: a theme in signal transduction. Science. 1995 Apr 14;268(5208):247–251. doi: 10.1126/science.7716516. [DOI] [PubMed] [Google Scholar]
  29. Nakanishi H., Brewer K. A., Exton J. H. Activation of the zeta isozyme of protein kinase C by phosphatidylinositol 3,4,5-trisphosphate. J Biol Chem. 1993 Jan 5;268(1):13–16. [PubMed] [Google Scholar]
  30. Nishimura H., Simpson I. A. Staurosporine inhibits phorbol 12-myristate 13-acetate- and insulin-stimulated translocation of GLUT1 and GLUT4 glucose transporters in rat adipose cells. Biochem J. 1994 Aug 15;302(Pt 1):271–277. doi: 10.1042/bj3020271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Okada T., Kawano Y., Sakakibara T., Hazeki O., Ui M. Essential role of phosphatidylinositol 3-kinase in insulin-induced glucose transport and antilipolysis in rat adipocytes. Studies with a selective inhibitor wortmannin. J Biol Chem. 1994 Feb 4;269(5):3568–3573. [PubMed] [Google Scholar]
  32. Osada S., Mizuno K., Saido T. C., Suzuki K., Kuroki T., Ohno S. A new member of the protein kinase C family, nPKC theta, predominantly expressed in skeletal muscle. Mol Cell Biol. 1992 Sep;12(9):3930–3938. doi: 10.1128/mcb.12.9.3930. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Pessin J. E., Bell G. I. Mammalian facilitative glucose transporter family: structure and molecular regulation. Annu Rev Physiol. 1992;54:911–930. doi: 10.1146/annurev.ph.54.030192.004403. [DOI] [PubMed] [Google Scholar]
  34. RODBELL M. METABOLISM OF ISOLATED FAT CELLS. I. EFFECTS OF HORMONES ON GLUCOSE METABOLISM AND LIPOLYSIS. J Biol Chem. 1964 Feb;239:375–380. [PubMed] [Google Scholar]
  35. Schonhorn J. E., Akompong T., Wessling-Resnick M. Mechanism of transferrin receptor down-regulation in K562 cells in response to protein kinase C activation. J Biol Chem. 1995 Feb 24;270(8):3698–3705. doi: 10.1074/jbc.270.8.3698. [DOI] [PubMed] [Google Scholar]
  36. Simpson I. A., Yver D. R., Hissin P. J., Wardzala L. J., Karnieli E., Salans L. B., Cushman S. W. Insulin-stimulated translocation of glucose transporters in the isolated rat adipose cells: characterization of subcellular fractions. Biochim Biophys Acta. 1983 Dec 19;763(4):393–407. doi: 10.1016/0167-4889(83)90101-5. [DOI] [PubMed] [Google Scholar]
  37. Smas C. M., Sul H. S. Control of adipocyte differentiation. Biochem J. 1995 Aug 1;309(Pt 3):697–710. doi: 10.1042/bj3090697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Toker A., Meyer M., Reddy K. K., Falck J. R., Aneja R., Aneja S., Parra A., Burns D. J., Ballas L. M., Cantley L. C. Activation of protein kinase C family members by the novel polyphosphoinositides PtdIns-3,4-P2 and PtdIns-3,4,5-P3. J Biol Chem. 1994 Dec 23;269(51):32358–32367. [PubMed] [Google Scholar]
  39. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Zang R., Müller H. J., Kielbassa K., Marks F., Gschwendt M. Partial purification of a type eta protein kinase C from murine brain: separation from other protein kinase C isoenzymes and characterization. Biochem J. 1994 Dec 1;304(Pt 2):641–647. doi: 10.1042/bj3040641. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES