Structural Studies and Functional Regulation of KLF4

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DESCRIPTION (provided by applicant): My long term goal is to understand the role of KLF4, a gut enriched transcriptional factor, in cancer development in the gastrinintestinal tract (Gl tract). KLF4 has potential use in the development of therapeutic strategies to treat cancer, especially colon cancer. Colorectal cancer is one of the most common cancers in Western countries and is the third most common cause of cancer-related deaths in the US. Approximately 50% of individuals develop a colorectal neoplasm by age 70 years. At the molecular level, in vitro systems are an irreplaceable method for studying the mechanism of KLF4 regulates the transcription of genes important for cancer development. Therefore I propose to pursue in-depth studies (Aim1 and Aim 2) using different cancer cell lines and human umbilical vein endothelial cells as in vitro systems to determine how KLF4 regulates transcription. These studies will identify the functional domains in KLF4 that are responsible for its proper localization; its interaction with another protein, Tip60; and its transcriptional activities. In addition, mouse models have been widely used to study human diseases. In Aim3, we will use a mouse model to study the in vivo regulation of KLF4 by Notch signaling pathway, inhibition of which causes goblet cell metaplasia. In an animal study, KLF4 has been shown to control goblet cell differentiation in colon. On the other hand, goblet cell differentiation is known to be controlled by Notch signal pathway, raising the possibility that KLF4 is under the control of Notch signaling pathway. Specifically, we will use a KLF4-GFP construct derived from a bacterial artificial chromosome (BAC) clone as the transgene to generate a transgenic mouse model (KLF4-GFP). Since GFP is under the control of native KLF4 promoter, this model will minimize the likelihood of misexpression and the insertion-position effects that are seen in transgenic studies using a small construct as the transgene. We will study the in vivo regulation of KLF4 by Notch signaling pathway using a gamma-secretase inhibitor and a mouse model overexpressing human progastrin, since in this model an increase of goblet cell number has been observed and KLF4 is upregulated by progastrin from our preliminary data.