Topology of the signal transduction of the G protein-coupled somatostatin receptor sst2 in human glioma cells (original) (raw)

Abstract.

By a dual approach, using electron microscopy and biochemical techniques, we investigated the topology of the somatostatin receptor sst2 with its inhibitory G protein Giα after ligand-induced stimulation and internalization in human glioma cells. On intact cells, the sst2 was labeled at 8°C by an antibody directed to its extracellular sequence followed by a 15-nm gold-labeled secondary antibody. In the presence of the ligand, internalization was induced by exposure to 37°C for 5–10 min. Then, cells were either fixed for immunoelectron-microscopic analysis or homogenized for density gradient separation. After post-embedding staining of the sst2-labeled sections with anti-Giα1–3 or anti-caveolin, a co-localization of sst2, Giα and caveolin was detected in endosomal vesicles after 5 min of internalization, but not after 10 min. Furthermore, the gold-labeled organelles containing the internalised receptor were separated from the non-labeled ones on sucrose gradients (density shift separation) and analyzed by Western blotting. Also here, in fractions with higher densities, sst2 could be co-stained with Giα and caveolin after 5 min. From these congruent results from both methods, it can be concluded that, in human glioma cells, the receptor sst2 (1) is internalised in caveolin-positive vesicles and (2) is neighboured to its Giα proteins at the plasma membrane and early endosomes.

Access this article

Log in via an institution

Subscribe and save

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

Author information

Authors and Affiliations

  1. Anatomisches Institut der Universität Kiel, Olshausenstrasse 40, 24098 Kiel, Germany, , , , ,
    Rolf Mentlein, Janka Held-Feindt & Brigitte Krisch

Authors

  1. Rolf Mentlein
    You can also search for this author inPubMed Google Scholar
  2. Janka Held-Feindt
    You can also search for this author inPubMed Google Scholar
  3. Brigitte Krisch
    You can also search for this author inPubMed Google Scholar

Additional information

Electronic Publication

Rights and permissions

About this article

Cite this article

Mentlein, R., Held-Feindt, J. & Krisch, B. Topology of the signal transduction of the G protein-coupled somatostatin receptor sst2 in human glioma cells.Cell Tissue Res 303, 27–34 (2001). https://doi.org/10.1007/s004410000302

Download citation