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Volume 271, Number 47, Issue of November 22, 1996 pp. 30290-30296
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

Post-transcriptional Control Regulates Transforming Growth Factor alpha  in the Human Carcinoma KB Cell Line

(Received for publication, February 6, 1996, and in revised form, August 26, 1996)

Gabriella Nicolini Dagger , Mariarosaria Miloso Dagger , Maria Cristina Moroni Dagger , Laura Beguinot Dagger and Luigi Scotto Dagger

From the Dagger  Laboratory of Molecular Oncology, DIBIT, and  Istituto di Neuroscience e Bioimmagini del CNR, H. S. Raffaele, Via Olgettina 60 20132 Milano, Italy

Expression of epidermal growth factor receptor (EGF-R) antisense RNA results in a drastic reduction of EGF-R levels in the human carcinoma KB cell line and induces a reversion of their transformed phenotype (Moroni, M. C., Willingham, M. C., and Beguinot, L. (1992) J. Biol. Chem. 267, 2714-2722). We used parental and EGF-R antisense KB clones as a genetic system to study, in the same cell line, the role of transforming growth factor alpha  (TGF-alpha ) in the establishment and maintenance of the transformed phenotype. KB cells produce TGF-alpha mRNA, and their conditioned medium is able to sustain growth of antisense cells, mimicking the effect of exogenous EGF or TGF-alpha . In antisense cells there is a marked reduction of TGF-alpha mRNA steady-state levels. In addition, the decrease in TGF-alpha parallels the levels of residual EGF-R in the various antisense clones, indicating a direct correlation between receptors and growth factor levels. The addition of exogenous TGF-alpha (10 ng/ml) to antisense clones induces TGF-alpha levels. The half-life of TGF-alpha mRNA is 40-60 min in antisense cells and more than 8 h in parental KB cells, as determined by actinomycin D decay curves. This result indicates a predominant regulation of TGF-alpha mRNA at the post-transcriptional level. Nuclear run-on experiments show that there is only a marginal effect at the transcriptional level. We conclude that the autocrine loop responsible for the transformed phenotype of the human carcinoma KB cell line is dependent on both elevated levels of EGF-R and the presence of TGF-alpha . In addition, TGF-alpha is able to induce its own mRNA via a signal due to activation of the EGF-R acting predominantly at the post-transcriptional level.


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