Regulation of transforming growth factor expression in a growth factor- independent cell line

Gillian M Howell, Lisa E. Humphrey, Barry L. Ziober, Rana Awwad, Basker Periyasamy, Alan Koterba, Wenhui Li, James K.V. Willson, Kevin Coleman, Joan Carboni, Mark Lynch, Michael G Brattain

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

Aberrant transcriptional regulation of transforming growth factor α (TGFα) appears to be an important contributor to the malignant phenotype and the growth factor independence with which malignancy is frequently associated. However, little is known about the molecular mechanisms responsible for dysregulation of TGFα expression in the malignant phenotype. In this paper, we report on TGFα promoter regulation in the highly malignant growth factor-independent cell line HCT116. The HCT116 cell line expresses TGFα and the epidermal growth factor receptor (EGFR) but is not growth inhibited by antibodies to EGFR or TGFα. However constitutive expression of TGFα antisense RNA in the HCT116 cell line resulted in the isolation of clones with markedly reduced TGFα mRNA and which were dependent on exogenous growth factors for proliferation. We hypothesized that if TGFα autocrine activation is the major stimulator of TGFα expression in this cell line, TGFα promoter activity should be reduced in the antisense TGFα clones in the absence of exogenous growth factor. This was the case. Moreover transcriptional activation of the TGFα promoter was restored in an antisense-TGFα-mRNA-expressing clone which had reverted to a growth factor- independent phenotype. Using this model system, we were able to identify a 25-bp element within the TGFα promoter which conferred TGFα autoregulation to the TGFα promoter in the HCT116 cell line. In the TGFα-antisense-RNA- expressing clones, this element was activated by exogenous EGF. This 25-bp sequence contained no consensus sequences of known transcription factors so that the TGFα or EGF regulatory element within this 25-bp sequence represents a unique element. Further characterization of this 25-bp DNA sequence by deletion analysis revealed that regulation of TGFα promoter activity by this sequence is complex as both repressors and activators brad in this region but the overall expression of the activators is pivotal in determining the level of response to EGF or TGFα stimulation. The specific nuclear proteins binding to this region are also regulated in an autocrine- TGFα-dependent fashion and by exogenous EGF in EGF-deprived TGFα antisense clone 33. This regulation is identical to that seen in the growth factor- dependent cell line FET, which requires exogenous EGF for optimal growth. Moreover, the time response of the stimulation of trans-acting factor binding by EGF suggests that the effect is directly due to growth factor and not mediated by changes in growth state. We conclude that this element appears to represent the major positive regulator of TGFα expression in the growth factor-independent HCT116 cell line and may represent the major site of transcriptional dysregulation of TGFα promoter activity in the growth factor-independent phenotype.

Original languageEnglish (US)
Pages (from-to)303-313
Number of pages11
JournalMolecular and cellular biology
Volume18
Issue number1
DOIs
StatePublished - Jan 1 1998

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Transforming Growth Factors
Intercellular Signaling Peptides and Proteins
Cell Line
Epidermal Growth Factor
HCT116 Cells
Clone Cells
Phenotype
Antisense RNA
Epidermal Growth Factor Receptor
Growth

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

Cite this

Howell, G. M., Humphrey, L. E., Ziober, B. L., Awwad, R., Periyasamy, B., Koterba, A., ... Brattain, M. G. (1998). Regulation of transforming growth factor expression in a growth factor- independent cell line. Molecular and cellular biology, 18(1), 303-313. https://doi.org/10.1128/MCB.18.1.303

Regulation of transforming growth factor expression in a growth factor- independent cell line. / Howell, Gillian M; Humphrey, Lisa E.; Ziober, Barry L.; Awwad, Rana; Periyasamy, Basker; Koterba, Alan; Li, Wenhui; Willson, James K.V.; Coleman, Kevin; Carboni, Joan; Lynch, Mark; Brattain, Michael G.

In: Molecular and cellular biology, Vol. 18, No. 1, 01.01.1998, p. 303-313.

Research output: Contribution to journalArticle

Howell, GM, Humphrey, LE, Ziober, BL, Awwad, R, Periyasamy, B, Koterba, A, Li, W, Willson, JKV, Coleman, K, Carboni, J, Lynch, M & Brattain, MG 1998, 'Regulation of transforming growth factor expression in a growth factor- independent cell line', Molecular and cellular biology, vol. 18, no. 1, pp. 303-313. https://doi.org/10.1128/MCB.18.1.303
Howell, Gillian M ; Humphrey, Lisa E. ; Ziober, Barry L. ; Awwad, Rana ; Periyasamy, Basker ; Koterba, Alan ; Li, Wenhui ; Willson, James K.V. ; Coleman, Kevin ; Carboni, Joan ; Lynch, Mark ; Brattain, Michael G. / Regulation of transforming growth factor expression in a growth factor- independent cell line. In: Molecular and cellular biology. 1998 ; Vol. 18, No. 1. pp. 303-313.
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