Autocrine TGFα expression in the regulation of initiation of human colon carcinoma growth

Previously, we reported that unaggressive, growth factor-dependent FET human colon carcinoma cells downregulated their transforming growth factor alpha (TGFα) expression in a quiescent state (G₀/G₁) induced by growth factor and nutrient deprivation (Mulder, 1991, Cancer Res., 51:2256-2262). In contrast, highly aggressive, growth factor-independent HCT116 human colon carcinoma cells aberrantly upregulated this autocrine activity in the quiescent state (Mulder, 1991, Cancer Res., 51:2256-2262; Howell et al., 1998, Mol. Cell. Biol., 18:303-313). In this report, the role of autocrine TGFα and the mechanism of its regulation of expression during reentry into the cell cycle from a noncycling growth state were determined in FET cells. Optimal induction of DNA synthesis from a quiescent state in FET cells is dependent upon autocrine TGFα as well as exogenous transferrin and insulin. Reentry into the cell cycle resulting from treatment with exogenous transferrin and insulin resulted in ~3-fold induction of TGFα expression within 1 hr. TGFα induction was controlled at the transcription level, and the cis-controlling element was localized to the region between bp -370- - 201 relative to the translation start codon within the TGFα promoter. Thus neutralization of autocrine TGFα protein revealed that the induced TGFα autocrine activity was necessary for DNA synthesis and acted only in the early G₁ phase of the cell cycle. Blockade of autocrine TGFα expression early in the cell cycle resulted in the reduction of DNA synthesis, whereas treatment with neutralization antibody at later times had no effect. This suggested that autocrine TGFα functions to initiate cell growth from noncycling states. This was further confirmed by the dependence of FET cells upon autocrine TGFα for colony formation in experiments where the plating density was sufficiently low to generate a lag phase in tissue culture. In contrast, TGFα autocrine activity was not required for exponential phase cells, as evidenced by the failure of TGFα neutralizing antibody to inhibit proliferation in this growth state. Taken together, these results suggest that autocrine TGFα acts primarily in the process of growth initiation by moving cells from a noncycling state back into the cell cycle, rather than supporting cell growth already initiated.