Author of

• 11481

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  P2.02 - Poster Session 2 - Novel Cancer Genes and Pathways (ID 148)

  • Event: WCLC 2013
  • Type: Poster Session
  • Track: Biology
  • Presentations: 1
  • Moderators:
  • Coordinates: 10/29/2013, 09:30 - 16:30, Exhibit Hall, Ground Level

  • 11483

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    P2.02-002 - Stratifin expressed in early invasive lung adenocarcinoma functionally enhances tumor progression and down-regulates SCF^Fbw7ubiquitin ligase activity (ID 659)

    09:47 - 10:04  |  Author(s): S. Iemura
    • Abstract

    Background
    Adenocarcinoma in situ (AIS) of the lung shows a very favorable prognosis, with a 5-year survival rate of 100%. However, early but invasive adenocarcinoma (eIA) sometimes has a fatal outcome. In order to elucidate the key molecule that affects the malignant progression of adenocarcinoma at an early stage, we previously compared the expression profiles of AIS with those of eIA, and found that stratifin (SFN, 14-3-3 sigma) is one of the differentially expressed genes related to tumor progression (Aya Shiba-Ishii, IJC. 2011). Immunohistochemistry (IHC) with anti-SFN antibody revealed that more than 95% of eIAs were SFN-positive, in comparison with only 13% of AISs. We also found that promoter demethylation triggered aberrant SFN overexpression in eIAs (Aya Shiba-Ishii, AJP. 2012). Here, we performed functional analysis of SFN and identification of SFN binding protein (SBP) to clarify how SFN affects the progression of lung adenocarcinoma.

    Methods
    For in vitro functional analysis, we performed RNA interference and expression vector transfection analyses and subsequent cell proliferation assays or cell cycle phase assay using a lung adenocarcinoma cell line (A549). An in vivo animal study was also performed using siSFN-transfected A549. Additionally, in order to identify SBP, SFN vector-transfected A549 was subjected to pull-down assay and subsequent LC-MS analysis. Interaction of SFN and SBP was examined using co-IP and western western blotting.

    Results
    Suppression of SFN expression by siSFN significantly reduced cell proliferation activity and the S-phase subpopulation. Transfection of the SFN expression vector led to a significant increase in cell proliferation. A549 treated with siSFN showed reduced tumor development relative to the controls. Pull-down assay and LC-MS analysis revealed that SKP1 is one of the SBPs. SKP1 is a component of the SKP1-Cullin1-F-box containing complex (SCF complex). We found that Fbw7, one of the substrate recognition subunits of SCF complex, also binds to the SFN-SKP1 complex, resulting in up-regulation of cyclin E1 phosphorylation by SFN.

    Conclusion
    Although SFN was originally identified as a negative regulator of the cell cycle, especially in response to p53-sensitive DNA damage, subsequent reports indicated that it is a positive mediator of cell proliferation. In breast cancer, SFN induces G1/S progression by increasing the expression of cyclin D1. Here, we demonstrated that SFN enhanced the proliferative capacity of lung adenocarcinoma cells both in vitro and in vivo. We also revealed that SFN down-regulated expression of the SCF[Fbw7] complex. As cyclin E1 is a common target of SCF[Fbw7] ubiquitination, we predict that SCF[Fbw7] down-regulation by SFN might induce stabilization of cyclin E1. Since we also found that the S-phase subpopulation was decreased after siSFN treatment, SFN might induce G1/S progression through an increase of cyclin E1 phosphorylation in lung adenocarcinoma. In conclusion, SFN facilitates cell proliferation and malignant progression of lung adenocarcinoma, and its mechanism of action is thought to be associated with inhibition of SCF[Fbw7] ubiquitination and degradation.