Reduced Expression of Transforming Growth Factor β Type I Receptor Contributes to the Malignancy of Human Colon Carcinoma Cells

doi:10.1074/jbc.271.29.17366

Reduced Expression of Transforming Growth Factor β Type I Receptor Contributes to the Malignancy of Human Colon Carcinoma Cells*

From the ^‡ Department of Biochemistry and Molecular Biology, Medical College of Ohio, Toledo, Ohio 43699, the
^§Department of Medicine and CWRU/Ireland Cancer Center, Case Western Reserve University, Cleveland, Ohio 44106, the
^¶Department of Pharmacology, Duke University Medical Center, Durham, North Carolina 27710, and the
^∥Department of Pharmacology, University of Kentucky College of Medicine, Lexington, Kentucky 40536

” To whom correspondence should be addressed. Tel.: 419-381-4324; Fax: 419-382-7395.

Abstract

Transforming growth factor β (TGFβ) type I (RI) and type II (RII) receptors are essential for TGFβ signal transduction. A human colon carcinoma cell line, designated GEO, is marginally responsive to TGFβ and expresses a low level of RI mRNA relative to colon carcinoma cells, which are highly responsive to TGFβ. Hence, the role of RI as a limiting factor for TGFβ sensitivity and the contribution of low RI levels to the malignant phenotype of GEO cells were examined. Stable transfection of a tetracycline-regulatable rat RI cDNA increased TGFβ₁ binding to RI and resulted in increased growth inhibition by exogenous TGFβ₁. In contrast, although stable transfection of an RII expression vector into the same GEO cells increased TGFβ₁ binding to RII, growth inhibition by exogenous TGFβ₁ was not altered. This indicated that the low level of RI is a limiting factor for the growth-inhibitory effects of TGFβ in GEO cells. RI-transfected cells were growth-arrested at a lower saturation density than GEO control cells. They also showed reduced growth and clonogenicity in plating efficiency and soft agarose assays, whereas RII-transfected cells did not show any differences from the NEO control cells in these assays. Tetracycline repressed RI expression in transfected cells and reversed the reduction in plating efficiency of RI-transfected clones, confirming that growth effects were due to increased RI expression in transfected cells. TGFβ₁ neutralizing antibody stimulated the proliferation of RI-transfected cells but had little effect on GEO control cells, indicating that increased autocrine-negative TGFβ activity also resulted from increased RI expression. Tumorigenicity in athymic nude mice was significantly delayed in RI-transfected cells. These results indicate that low RI expression can be a limiting factor for response to exogenous TGFβ, as well as TGFβ autocrine-negative activity, and that reduction of RI expression can contribute to malignant progression.

Footnotes

↵* This work was supported by National Institutes of Health Grants CA38173 and CA50457 (to M. G. B.), CA63480 (to L. Z. S.), CA68316 (to J. K. V. W.), and P30 CA4370301 (to the Case Western Reserve University/Ireland Cancer Center), and by United States Army Grant DAMD17-94-J-4065 (to X.F. Wang). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

TGFβ

transforming growth factor β

RI, RII, RIII

receptor types I, II, III, respectively

tTA

tetracycline-controlled transactivator

MTT

(3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide)

ECM

extracellular matrix.
- Received December 18, 1995.