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LPS-induced down-regulation of NO-sensitive guanylyl cyclase in astrocytes occurs by proteasomal degradation in nuclear bodies

Background

We have previously shown that inflammatory agents (LPS, IL-1β, β-amyloid peptides) that induce reactivity and NOS-2 expression in glial cells down-regulate astroglial soluble guanylyl cyclase (sGC) in vitro and in vivo [1, 2].

Results

Here we show that the decrease in sGC activity and β1 subunit protein induced by LPS (10 ng/ml, 24 h) in cultured rat cerebellar astrocytes is prevented by inhibitors of proteasome activity (MG132 5 μM; lactacystin 10 μM) whereas other protease inhibitors (calpain inhibitor 25 μM; ICE inhibitor II 100 μM and leupeptin 5 μM) were not effective. Furthermore, immunocytochemistry and confocal laser microscopy revealed that in LPS-treated cells a strong sGC β1 immunorreactivity is evident in aggregates in the cell nuclei where it co-localizes with 20S proteasomes and ubiquitin in clastosomes, nucleoplasmic substructures involved in ubiquitin-proteasome-dependent nuclear proteolysis, but do not colocalize with others proteasome-enriched structures include promyelocytic leukaemia bodies and splicing speckles. In contrast, in untreated astrocytes clastosomes are scarce and sGC β1 immunorectivity shows a diffuse cytoplasmic pattern, while in the nucleus it is very weak. A similar distribution is observed when cells are treated with LPS and the proteasome inhibitor MG132 or the protein synthesis inhibitor cycloheximide.

Conclusion

LPS orchestrates the recruitment of sGC-β1 protein and components of the ubiquitin-proteasome system to specialized nuclear bodies, clastosomes, suggesting a mechanism for inflammation-induced down-regulation of sGC in astrocytes.

References

  1. 1.

    Baltrons MA, Pedraza CE, Heneka MT, García A: β-Amyloid peptides decrease soluble guanylyl cyclase expression in astroglial cells. Neurobiol Dis. 2002, 10: 39-149. 10.1006/nbdi.2002.0492.

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  2. 2.

    Pedraza CE, Baltrons MA, Heneka MT, García A: Interleukin-1β and lipopolysaccharide decrease soluble guanylyl cyclase in cells of the CNS: NO-independent destabilization of protein and NO-dependent decrease of mRNA. J Neuroimmunol. 2003, 144: 80-90. 10.1016/j.jneuroim.2003.08.034.

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Acknowledgements

This work was supported by a SAF2004-01717 grant (Spain).

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Correspondence to María Antonia Baltrons.

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Open Access This article is published under license to BioMed Central Ltd. This is an Open Access article is distributed under the terms of the Creative Commons Attribution 2.0 International License (https://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Baltrons, M.A., Pifarré, P., Berciano, M.T. et al. LPS-induced down-regulation of NO-sensitive guanylyl cyclase in astrocytes occurs by proteasomal degradation in nuclear bodies. BMC Pharmacol 7, P3 (2007). https://doi.org/10.1186/1471-2210-7-S1-P3

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Keywords

  • Guanylyl Cyclase
  • Nuclear Body
  • Calpain Inhibitor
  • Soluble Guanylyl Cyclase
  • Proteasome Inhibitor MG132