Elsevier

Cellular Signalling

Volume 23, Issue 5, May 2011, Pages 785-793
Cellular Signalling

Alteration of Golgi structure in senescent cells and its regulation by a G protein γ subunit

https://doi.org/10.1016/j.cellsig.2011.01.001Get rights and content

Abstract

Cellular senescence is a process wherein proliferating cells undergo permanent cell cycle arrest while remaining viable. Senescence results in enhanced secretion of proteins that promote cancer and inflammation. We report here that the structure of the Golgi complex which regulates secretion is altered in senescent cells. In cells where senescence is achieved by replicative exhaustion or in cells wherein senescence has been induced with BrdU treatment dependent stress, the Golgi complex is dispersed. The expression of a G protein γ subunit, γ11, capable of translocation from the plasma membrane to the Golgi complex on receptor activation increases with senescence. Knockdown of γ11 or overexpression of a dominant negative γ3 subunit inhibits Golgi dispersal induced by senescence. Overall these results suggest that in cellular senescence an upregulated G protein gamma subunit mediates alterations in the structure of the Golgi.

Introduction

Replicatively senescent cells or cells that have undergone genotoxic stress exhibit increased secretion of a number of factors including cytokines, growth factors, metalloproteinases and extracellular matrix proteins [1]. The enhanced secretion of these factors is known to induce inflammation and has been demonstrated to facilitate epithelial mesenchymal transition which promotes tumorigenesis [1]. Since cellular secretion is mediated by the Golgi complex, we examined the status of the Golgi in senescent cells resulting from stress or replicative exhaustion. Based on previous reports, 5-bromo 2-deoxyuridine (BrdU) exposure to cells was used as a stress induced model for senescence [2], [3], [4] which mimics the properties of replicative senescence. It has been shown that BrdU treatment induces cellular senescence likely by inducing the DNA-damage response [5]. DNA damage has been shown to trigger senescence [6]. It has been shown that it induces senescence in stem cells and inhibits proliferation of cancer cells [15], [16]. We also confirmed a previous finding that a heterotrimeric G protein subunit, γ11 (GNG11) is upregulated in senescent cells [7]. The γ11 subunit is capable of translocation from the plasma membrane to the Golgi on receptor activation as a βγ complex [8], [9] and regulates the structure of the Golgi [10]. We therefore examined the possibility that the G protein γ11 subunit plays a role in the regulation of the Golgi structure in senescence.

Section snippets

Constructs, cell lines and chemicals

The tagged and untagged G protein constructs, various Golgi markers and PH-mCh used in this study have been previously described [9], [10], [11], [12]. Mammalian expression vector containing cDNA encoding γ11 shRNA and control scrambled shRNAs were from the TRC library of Broad Institute (Sigma) and CFP-tubulin was from E. Bertrand (CNRS, Montpellier, France). The HeLa cell line was from ATCC; the WI38 and IMR90 cell lines were from NIA Aging Cell Repository at Coriell Institute for Medical

BrdU (5-bromo deoxyuridine) induces cellular senescence

We used BrdU treated HeLa cells as a rapidly assayable model for cellular senescence. It has been observed that BrdU induces cellular senescence by activating the DNA damage response [5], [14]. Consistent with previous reports [2], [4], [5], [14], [15], [17] we found that BrdU induces cellular senescence. HeLa cells treated with 200 μM BrdU for 24–48 h showed several characteristics similar to replicatively senescent IMR90 cells. BrdU treated cells slowed down in growth, changed shape (Fig. 1A

Discussion

The structure of the Golgi complex in senescent cells has not been examined before to our knowledge. Results here show that the Golgi complex is compact in non-senescent or pre-senescent cells but is dispersed in senescent cells. The consistently altered appearance of the Golgi in both stress induced senescent cells and replicatively senescent cells suggests that the Golgi structure could serve as a useful marker for cellular senescence. Senescent cells show significantly increased secretion of

Acknowledgements

This work was supported by NIH grants (GM69027 and GM080558) to N.G. and an AHA postdoctoral fellowship (D.K.S.).

References (30)

  • T. Suzuki et al.

    Exp. Gerontol.

    (2001)
  • S. Minagawa et al.

    Exp. Cell Res.

    (2005)
  • M.N. Hossain et al.

    Biochem. Biophys. Res. Commun.

    (2006)
  • M. Akgoz et al.

    J. Biol. Chem.

    (2004)
  • D.K. Saini et al.

    J. Biol. Chem.

    (2007)
  • M. Chisari et al.

    J. Biol. Chem.

    (2007)
  • L.H. Levkoff et al.

    Neoplasia

    (2008)
  • I.B. Roninson

    Cancer Lett.

    (2002)
  • T. Kumazaki et al.

    Exp. Cell Res.

    (1991)
  • J.C. Acosta et al.

    Cell

    (2008)
  • B. Storrie et al.

    Biochim. Biophys. Acta

    (1998)
  • J. Hoebeke et al.

    Biochem. Biophys. Res. Commun.

    (1976)
  • E. Ruiz-Ballesteros et al.

    Blood

    (2005)
  • J.P. Coppe et al.

    Annu. Rev. Pathol.

    (2010)
  • E. Michishita et al.

    J. Biochem.

    (1999)
  • Cited by (32)

    • G protein gamma subunit, a hidden master regulator of GPCR signaling

      2022, Journal of Biological Chemistry
      Citation Excerpt :

      Though the βγ complex in a reconstitution assay has been shown to stimulate Golgi fragmentation via a PKD- and PLCβ-mediated pathway (107) and Golgi localized βγ complex can regulate protein transport from the trans-Golgi network to the cell surface (108) it had earlier been unclear how the βγ complex reached the Golgi. A follow-up study further demonstrated that γ11 subunit also regulates cellular senescence by acting on the Golgi structure in response to GPCR activation (109). Similarly, subtype-dependent βγ complex translocation to the Golgi complex regulates the ERK pathway and cancer metastasis through PI3Kγ activation (87).

    • Subtype-dependent regulation of Gβγ signalling

      2021, Cellular Signalling
      Citation Excerpt :

      We have discussed above the enhanced Golgi fragmentation in the A569 cell line, which shows an abundant endogenous expression of Gγ11 [204]. Cellular senescence is another cellular physiological function potentially regulated by Gγ signalling at the Golgi complex, and the increased expression of Gγ11 in senescent cells was reported in another study [207]. Consistent with all the above data, the downregulation of cellular functions driven by signalling at the Golgi complex was observed due to expression of low translocating/dominant-negative Gγ3 [204,207].

    • A G-protein subunit translocation embedded network motif underlies GPCR regulation of calcium oscillations

      2014, Biophysical Journal
      Citation Excerpt :

      PH-mCh (17) was used to detect inositol trisphosphate (IP3) production during Gi activation in HeLa cells. A γ11 knockdown stable HeLa cell line was used where γ11 was knocked down using shRNA in a lentivirus resulting in ∼70–80% reduction of γ11 expression (34). Another stable HeLa cell line was created using the non-Target shRNA control virus.

    View all citing articles on Scopus
    1

    These authors contributed equally to this work.

    View full text