process. However, the existence ofTGF-Punresponsive MvlLu cell mutants that lack type I receptors yet have normal levels of type II receptors (5) suggests that type I receptors may be needed to mediate these activities of TGF-P through the type II receptor. Our findings also indicate that the signaling through the type I receptor does not require a functional type II receptor to mediate autonomously its distinct set of TGF-P-induced activities. This is consistent with the fact that 293 cells, which lack detectable levels of type II receptors but have type I receptors (Fig. 1A), are not responsive to the antiproliferative activity of TGFyet display a TGF-P-induced synthesis of fibronectin (20). These data further support our conclusion that the type II receptor is required for and mediates the antiproliferative effect of TGF-1, whereas the induction of fibronectin synthesis is mediated by the type I receptor. Moreover, the wide variability and lack of correlation between the cell surface levels of the type I and type II receptors suggest that besides possible heterodimers, there may be type I and type II receptors that do not physically interact with each other and that perhaps could function as homodimers. Finally, the selective functional abolition of the type II receptors results in alterations in the complex response to TGF-P similar to the functional inactivation of pRB by viral transforming proteins such as the SV40 large T antigen, that is, a specific abrogation of the antiproliferative effect of TGF(1 1, 21) without affecting the induction of expression of several genes by TGF-P (11). This very similar phenotype thus indicates that the role of pRB in the response to TGF-P is specific for and restricted to the signaling pathway associated with the type II receptor. Accordingly, our transfected cells lacking functional type II receptors did not show TGF-,B-induced inhibition ofpRB phosphorylation, suggesting that type II receptors are required for the effect of TGF-,B on the phosphorylation state of pRB. Roberts, M. B. Spom, J. Biol. Chem. 267, 2588 (1992). 10. J. Wrana et al., Cell 71, 1003 (1992). 11. M. Laiho et al., Mol. Cell. Biol. 11, 972 (1991). 12. A. Kimchi, X.-F. Wang, R. A. Weinberg, S. Cheifetz, J. Massagu6, Science 240, 196 (1988). 13. R.-H. Chen, R. Ebner, R. Derynck, unpublished results. 14. I. Herskowitz, Nature 329, 219 (1987). 15. 0. Kashles, Y. Yarden, R. Fischer, A. Ullrich, J. Schlessinger, Mol. Cell. Biol. 11, 1454 (1991). 16. H. Ueno, H. Colbert, J. A. Escobedo, L. T. Williams, Science 252, 844 (1991). 17. E. Amaya, T. J. Musci, M. W. Kirschner, Cell 66, 257 (1991). 18. A. Hemmati-Brivanlou and D. A. Melton, Nature 359, 609 (1992). 19. M. Laiho, T. A. De Caprio, J. W. Ludlow, D. M. Livingston, J. Massague, Cell 62, 175 (1990). 20. B. A. Arrick et al., J. Cell Biol. 118, 715 (1992). 21. J. A. Pietenpol et al., Cell 61, 777 (1990). 22. T. P. Hopp et al., Biotechnology 6, 1204 (1988). 23. C. Gorman, R. Padmanabhan, B. H. Howard, Science 221, 551 (1983). 24. D. Gazit, R. Ebner, A. J. Kahn, R. Derynck, Mol. Endocrinol. 7,189 (1993). 25. To generate the truncated type 11 receptor, we made an Eco RI-Hpa fragment corresponding to nucleotides -7 through 573 of the type 11 receptor by polymerase chain reaction (PCR), followed by cleavage at the unique Hpa site located immediately 3' of the transmembrane domain sequence. A double-stranded oligonucleotide adaptor for the sequence encoding the epitope tag, FLAG (22), was ligated to the 3' end of the truncated type 11 cDNA, and the resulting fragment was inserted into the Eco RI and Xba sites of the expression vector pRK5, thus generating the expression plasmid for the truncated type 11 receptor. DNA was transfected into the 0T6 cells and 293 cells by calcium phosphate precipitation (23). Cross-linking analysis was performed as described (3, 24), and the cross-linked proteins were separated by denaturing and reducing polyacrylamide (7.5%) gel electrophoresis and subsequent autoradiography. 26. J. L. Graycar et al., Mol. Endocrinol. 3, 1977 (1989). 27. The expression vector for the complete human type 11 TGF-P receptor was made by insertion of an Eco RI fragment containing the entire cDNA coding sequence into the Eco RI opened pRK5, a human cytomegalovirus promoter-based expression plasmid (26). This Eco RI cDNA fragment was generated by the PCR with the human type 11 TGF-P receptor (8) as template and corresponded to nucleotides -7 to 1703 of the published sequence (8). 28. F. Ausubel et al., Current Protocols in Molecular Biology (Greene, New York, 1991). 29. B. Like and J. Massague, J. Biol. Chem. 261, 13426 (1986). 30. R. Ignotz and J. Massague, ibid., p. 4337. 31. We thank H. Lin and R. Weinberg for providing the type 11 TGF-P receptor cDNA and E. Filvaroff, Z. Werb, and K. Yamamoto for helpful discussions. Supported by grants from the American Cancer Society and the National Institutes of Health (to R.D.) and by a fellowship from the Deutsche Forschungsgemeinschaft (to R.E.).