Cancer Chemotherapy and Pharmacology

, Volume 75, Issue 2, pp 411–420 | Cite as

Cks1 proteasomal degradation is induced by inhibiting Hsp90-mediated chaperoning in cancer cells

  • Vinayak Khattar
  • Joshua Fried
  • Bo Xu
  • Jaideep V. Thottassery
Original Article

Abstract

Purpose

Cks1, a conformationally heterogenous 9 kDa protein, is markedly overexpressed in cancer cells and contributes to tumor development. Cks1 is an essential component of the SCF-Skp2 ubiquitin ligase complex that targets the Cdk inhibitors p27Kip1 and p21Cip1. Cks1 is known to interact with the Hsp90-Cdc37 chaperone machinery, although whether this facilitates its conformational maturation and stability is not known. To test whether abrogating the chaperone function of Hsp90 could destabilize Cks1, we examined the effects of treating different cancer cell lines with the benzoquinone ansamycin 17-allylamino geldanamycin (17-AAG), a compound that selectively binds Hsp90 and potently inhibits its ATP-dependent chaperone activity.

Methods

The effect of Hsp90 inhibition using 17-AAG on Cks1 protein and associated cell cycle proteins including Skp2, p27Kip1, p21Cip1, and Cdk1 in cancer cells was determined by Western blotting. Ubiquitination analysis was carried out by transfecting cells with an HA-ubiquitin plasmid and specifically immunoprecipitating Cks1 to examine polyubiquitinated species. Flow cytometry was utilized to examine the effects of Hsp90 inhibition on cell cycle profiles.

Results

Here, we demonstrate for the first time that inhibition of Hsp90 utilizing 17-AAG destabilizes Cks1 in cancer cells by promoting its ubiquitination and proteasomal degradation. 17-AAG-induced Cks1 depletion was accompanied by concomitant decreases in Skp2 and Cdk1. 17-AAG treatment also induced G2/M accumulation in MCF-7 breast carcinoma cells, and G1 accumulation in the colon carcinoma lines HCT116 and SW620.

Conclusions

We conclude that perturbing the Hsp90 pathway could provide a useful therapeutic strategy in tumors driven by Cks1 overexpression.

Keywords

Cks1 17-AAG Skp2 p27Kip1 Hsp90 Geldanamycin Proteasome 

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Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Vinayak Khattar
    • 1
    • 2
  • Joshua Fried
    • 1
    • 2
  • Bo Xu
    • 1
    • 2
  • Jaideep V. Thottassery
    • 1
    • 2
    • 3
  1. 1.Department of OncologySouthern Research InstituteBirminghamUSA
  2. 2.Cancer Cell Biology Program UAB Comprehensive Cancer CenterBirminghamUSA
  3. 3.Department of Oncology, Drug Discovery DivisionSouthern Research InstituteBirminghamUSA

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