Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1976 Sep 1;70(3):671–684. doi: 10.1083/jcb.70.3.671

Presence of adenylate cyclase activity in Golgi and other fractions from rat liver. II. Cytochemical localization within Golgi and ER membranes

PMCID: PMC2109840  PMID: 956270

Abstract

The presence of adenylate cyclase (AC) in liver Golgi and microsomal fractions from ethanol-treated rats was tested cytochemically using 5'- adenylyl imidodiphosphate (AMP-PNP) lead phosphate method. Parallel biochemical assays showed that rat liver Golgi AC was only partially inhibited by lead: in the presence of 1 mM Pb++ 80% of the enzyme was preserved, while when 2 mM Pb++ was used 25% remained. No cAMP was formed when the AMP-PNP medium was incubated in the presence of 1 or 2 mM Pb++ but in the absence of cell fractions, indicating that at these concentrations Pb++ does not cause the nonenzymatic hydrolysis of AMP- PNP. Therefore, the reaction product observed by cytochemical localization is not due to the nonenzymatic hydrolysis of AMP-PNP by Pb++. In Golgi subfractions, lead phosphate reaction product was widely distributed among Golgi elements: it was seen in association with the majority of the very low density lipoprotein-filled secretory droplets which predominated in the two lightest Golgi fractions (GF1 and GF2) as well as within the majority of the cisternae found in the heaviest Golgi fraction (GF3). In the latter, reaction product was heaviest along the dilated peripheral rims of the cisternae. In all cases, the reaction product was localized to the outside or cytoplasmic face of the Golgi membranes. When microsomes were incubated cytochemically for AC, deposits were found on the cytoplasmic surface of smooth endoplasmic reticulum (ER) membranes, but none were observed on rough ER membranes. The results confirm the biochemical data reported previously indicating the presence of AC in Golgi and smooth microsomal fractions from rat liver and further demonstrate that the activity is indeed indigenous to Golgi elements and not due to plasma membrane contaminants. They also indicate that AC is widely distributed among Golgi and smooth ER elements. Thus, AC is not restricted in its distribution to plasma membranes as usually assumed.

Full Text

The Full Text of this article is available as a PDF (5.4 MB).

Selected References

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

  1. Cheng H., Farquhar M. G. Presence of adenylate cyclase activity in Golgi and other fractions from rat liver. I. Biochemical determination. J Cell Biol. 1976 Sep;70(3):660–670. doi: 10.1083/jcb.70.3.660. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cutler L. S. Letter: Comments on the validity of the use of lead nitrate for the cytochemical study of adenylate cyclase. J Histochem Cytochem. 1975 Oct;23(10):786–787. doi: 10.1177/23.10.1194668. [DOI] [PubMed] [Google Scholar]
  3. Ehrenreich J. H., Bergeron J. J., Siekevitz P., Palade G. E. Golgi fractions prepared from rat liver homogenates. I. Isolation procedure and morphological characterization. J Cell Biol. 1973 Oct;59(1):45–72. doi: 10.1083/jcb.59.1.45. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Farnham C. J. Cytochemical localization of adenylate cyclase and 3',5'-nucleotide phosphodiesterase in Dictyostelium. Exp Cell Res. 1975 Mar 1;91(1):36–46. doi: 10.1016/0014-4827(75)90138-x. [DOI] [PubMed] [Google Scholar]
  5. Farquhar M. G., Bergeron J. J., Palade G. E. Cytochemistry of Golgi fractions prepared from rat liver. J Cell Biol. 1974 Jan;60(1):8–25. doi: 10.1083/jcb.60.1.8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Howell S. L., Whitfield M. Cytochemical localization of adenyl cyclase activity in rat islets of Langerhans. J Histochem Cytochem. 1972 Nov;20(11):873–879. doi: 10.1177/20.11.873. [DOI] [PubMed] [Google Scholar]
  7. Jande S. S., Robert P. Cytochemical localization of parathyroid hormone activated adenyl cyclase in rat kidney. Histochemistry. 1974;40(4):323–327. doi: 10.1007/BF00495039. [DOI] [PubMed] [Google Scholar]
  8. Lemay A., Jarett L. Pitfalls in the use of lead nitrate for the histochemical demonstration of adenylate cyclase activity. J Cell Biol. 1975 Apr;65(1):39–50. doi: 10.1083/jcb.65.1.39. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Marchesi V. T., Palade G. E. The localization of Mg-Na-K-activated adenosine triphosphatase on red cell ghost membranes. J Cell Biol. 1967 Nov;35(2):385–404. doi: 10.1083/jcb.35.2.385. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Perkins J. P. Adenyl cyclase. Adv Cyclic Nucleotide Res. 1973;3:1–64. [PubMed] [Google Scholar]
  11. Pohl S. L., Birnbaumer L., Rodbell M. The glucagon-sensitive adenyl cyclase system in plasma membranes of rat liver. I. Properties. J Biol Chem. 1971 Mar 25;246(6):1849–1856. [PubMed] [Google Scholar]
  12. Reik L., Petzold G. L., Higgins J. A., Greengard P., Barrnett R. J. Hormone-sensitive adenyl cyclase: cytochemical localization in rat liver. Science. 1970 Apr 17;168(3929):382–384. doi: 10.1126/science.168.3929.382. [DOI] [PubMed] [Google Scholar]
  13. Rodbell M., Birnbaumer L., Pohl S. L., Krans H. M. The glucagon-sensitive adenyl cyclase system in plasma membranes of rat liver. V. An obligatory role of guanylnucleotides in glucagon action. J Biol Chem. 1971 Mar 25;246(6):1877–1882. [PubMed] [Google Scholar]
  14. Rodbell M. In vitro assays of adenyl cyclase. Acta Endocrinol Suppl (Copenh) 1971;153:337–347. doi: 10.1530/acta.0.068s337. [DOI] [PubMed] [Google Scholar]
  15. Rodbell M., Krishna G. Preparation of isolated fat cells and fat cell "ghosts"; methods for assaying adenylate cyclase activity and levels of cyclic AMP. Methods Enzymol. 1974;31:103–114. doi: 10.1016/0076-6879(74)31010-5. [DOI] [PubMed] [Google Scholar]
  16. Szmigielski S. The use of dextran in phosphatase techniques employing lead salts. J Histochem Cytochem. 1971 Aug;19(8):505–507. doi: 10.1177/19.8.505. [DOI] [PubMed] [Google Scholar]
  17. Wagner R. C., Kreiner P., Barrnett R. J., Bitensky M. W. Biochemical characterization and cytochemical localization of a catecholamine-sensitive adenylate cyclase in isolated capillary endothelium. Proc Natl Acad Sci U S A. 1972 Nov;69(11):3175–3179. doi: 10.1073/pnas.69.11.3175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Widnell C. C. Cytochemical localization of 5'-nucleotidase in subcellular fractions isolated from rat liver. I. The origin of 5'-nucleotidase activity in microsomes. J Cell Biol. 1972 Mar;52(3):542–558. doi: 10.1083/jcb.52.3.542. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Yount R. G., Babcock D., Ballantyne W., Ojala D. Adenylyl imidodiphosphate, an adenosine triphosphate analog containing a P--N--P linkage. Biochemistry. 1971 Jun 22;10(13):2484–2489. doi: 10.1021/bi00789a009. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES