Studies were performed to identify membrane receptors for transferrin in cultured normal rat kidney (NRK) cells. Cells were surface iodinated or metabolically labeled with radioactive glycoprotein precursors. Membrane receptors for transferrin were solubilized with the nonionic detergent Triton X-100. The soluble transferrin receptor has been purified approximately 1500-fold by affinity chromatography using transferrin coupled to Sepharose. Experiments demonstrated that the receptor can be adsorbed to a transferrin-Sepharose gel and be eluted specifically with transferrin. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the receptor preparations obtained by one cycle of affinity chromatography display, in addition to components of Mr lower than 20 000, a major glycoprotein component of approximately 170 000. Solubilized receptor preparations subjected to two cycles of affinity chromatography revealed a single polypeptide of approximately 20 000 daltons. Further studies indicated that the 20 000-dalton polypeptide is a degradation product of the 170 000 glycoprotein. Immunological studies showed that antitransferrin antibodies specifically precipitate a transferrin-170 000 complex and that a specific antibody against 170 000 glycoprotein precipitates the same complex. These results suggest that the 170 000 glycoprotein associates with transferrin in specific fashion and that this protein may be a subunit of the transferrin receptor of NRK cells.
We have investigated the actions of transforming growth factor (TGF) type alpha on epidermal growth factor (EGF) receptor mRNA expression in MDA-468 human mammary carcinoma cells in serum-free media. We found that exposure of MDA-468 cells to TGF alpha results in elevated levels of EGF receptor mRNA. This increase in mRNA accumulation showed time and dose dependence. Addition of TGF beta 1 enhanced the accumulation of EGF receptor mRNA induced by TGF alpha in a time- and dose-dependent manner. We also found that triiodothyronine at physiological concentrations exerts synergistic control on the action of TGF alpha alone, or in association with TGF beta 1, on EGF receptor mRNA expression. Similarly, retinoic acid treatment also enhanced in a time- and dose-dependent manner the TGF alpha-dependent response of EGF receptor mRNA and acted synergistically with TGF beta 1. The results described here suggest that optimum regulation of EGF receptor gene expression by TGF alpha is a complex process involving synergistic interactions with heterologous growth factors and hormones.
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