Changes of p 27<sup>Kip1</sup>  in rat tongue carcinoma cells induced by 4-nitroquinoline 1-oxide

Aoyagi Nobuyoshi, Ueno Shigeru

doi:10.18905/shikaigaku.66.1_32

We investigated changes in the cell-cycle regulatory proteins in malignant cells from rat tongue carcinoma that had been chemically induced by 4-nitroquinoline 1-oxide. We found a decreased expression of p 27^Kip1 in carcinoma using immunohistochemistry and immunoblotting. Lysate from carcinoma cells showed an induced expression of cyclin E, cyclin D₁ and CDK 4 compared with lysate from normal cells. We found that fragments of 22 kD were produced by breakdown of p 27^Kip1 at the N-terminus when we used immunoblotting with two types of polyclonal anti-p 27^Kip1 antibodies, C-19 and N-20, which recognize the C-terminus and the N-terminus, respectively. In order to demonstrate the involvement of ubiquitin-dependent proteolysis, we incubated tumor cells with ubiquitin and proteasome inhibitors and examined the lysates by immunoblotting. We found a ladder pattern of multiple C-19 positive bands, which indicates polyubiquitination. Since these multibands were not seen in the samples incubated without proteasome inhibitors, we concluded that the ubiquitin-proteasome proteolytic system contributes to the breakdown of p 27^Kip1 in this carcinoma model. A decrease of p 27^Kip1 and an increase of cyclin E, cyclin D₁, and CDK 4 in the cells may contribute to the transformation from cellular quiescence that could result in carcinogenesis. Shika Igaku (J Osaka Odontol Soc) 2003 Mar; 66(1): 32-38.

Changes of p 27<sup>Kip1</sup> in rat tongue carcinoma cells induced by 4-nitroquinoline 1-oxide

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