Expression of Smad4 in the FaDu cell line partially restores TGF-beta growth inhibition but is not sufficient to regulate fibronectin expression or suppress tumorigenicity

✍ Scribed by B. Timothy Hummer; Christopher Bartlett; Erin Henry; Bernard E. Weissman

• Publisher: John Wiley and Sons
• Year: 2003
• Tongue: English
• Weight: 354 KB
• Volume: 194
• Category: Article
• ISSN: 0021-9541
• DOI: 10.1002/jcp.10202

✦ Synopsis

Abstract

Mutations of the Smad4 gene, a member of a group of TGF‐β signal transduction components, occur in several types of cancer suggesting that its inactivation significantly affects TGF‐β responsiveness in these tumors. To further investigate the role of Smad4 with respect to TGF‐β signaling and carcinogenesis, we re‐expressed the Smad4 gene in the Smad4‐deficient cancer cell line FaDu by microcell‐mediated chromosome transfer (MMCT) and retroviral infection to closely approximate physiological protein levels. The Smad4‐expressing FaDu clones were then evaluated for TGF‐β responsiveness to assess the role of Smad4 in TGF‐β‐induced growth inhibition and target gene regulation. We found that the re‐expression of the Smad4 gene by either method partially restored TGF‐β responsiveness in FaDu cells with respect to both growth inhibition and expression of p21^WAF1/CIP1^ and p15^INK4B^. However, only the microcell hybrids showed growth retardation in organotypic raft culture and an enhanced ability to upregulate fibronectin. In contrast, the re‐expression of Smad4 by either method failed to suppress tumorigenicity. These results suggest that in addition to a homozygous deletion of Smad4, FaDu cells contain additional defects within the TGF‐β signaling pathway, thereby limiting the extent of TGF‐β responsiveness upon Smad4 re‐expression and perhaps accounting for the inability to induce p15^INK4B^ to a high level. They also demonstrate the advantages of providing a physiological extracellular environment, when assessing TGFβ responsiveness. © 2003 Wiley‐Liss, Inc.