Lomberk, Gwen L

SP/KLF (Specificity protein/Krüppel-like factor) transcription factors comprise an emerging group of proteins that may behave as tumour suppressors. Incidentally, many cancers that display alterations in certain KLF proteins are also associated with a high incidence of KRAS (V-Ki-ras2 Kirsten rat sarcoma viral oncogene homologue) mutations. Therefore in the present paper we investigate whether SP/KLF proteins suppress KRAS-mediated cell growth, and more importantly, the potential mechanisms underlying these effects. Using a comprehensive family-wide screening of the 24 SP/KLF members, we discovered that SP5, SP8, KLF2, KLF3, KLF4, KLF11, KLF13, KLF14, KLF15 and KLF16 inhibit cellular growth and suppress transformation mediated by oncogenic KRAS. Each protein in this subset of SP/KLF members individually inhibits BrdU (5-bromo-2-deoxyuridine) incorporation in KRAS oncogenic-mutant cancer cells. SP5, KLF3, KLF11, KLF13, KLF14 and KLF16 also increase apoptosis in these cells. Using KLF11 as a representative model for mechanistic studies, we demonstrate that this protein inhibits the ability of cancer cells to form both colonies in soft agar and tumour growth in vivo. Molecular studies demonstrate that these effects of KLF11 are mediated, at least in part, through silencing cyclin A via binding to its promoter and leading to cell-cycle arrest in S-phase. Interestingly, similar to KLF11, KLF14 and KLF16 mechanistically share the ability to modulate the expression of cyclin A. Collectively, the present study stringently defines a distinct subset of SP/KLF proteins that impairs KRAS-mediated cell growth, and that mechanistically some members of this subset accomplish this, at least in part, through regulation of the cyclin A promoter.

Abstract Reflux-induced injury promotes esophageal adenocarcinoma, one of the most rapidly increasing, highly lethal cancers in Western countries. Here, we investigate the efficacy of a combinatorial chemoprevention strategy for esophageal adenocarcinoma and characterize the underlying molecular mechanisms. Specifically, our approach involves the use of ursodeoxycholic acid (Urso) due to its ability to decrease injury-inducing bile salts in combination with Aspirin to mitigate the consequences of injury. We find that Urso-Aspirin combination reduces the risk of adenocarcinoma in vivo in animals with reflux, decreases the proliferation of esophageal adenocarcinoma cells, and downregulates a key cell cycle regulator, CDK2. Mechanistically, using cell growth, luciferase reporter, expression, and chromatin immunoprecipitation assays, we identify GLI1, a Hedgehog-regulated transcription factor, as a novel target of Urso-Aspirin combination. We show that GLI1 is upregulated during esophageal carcinogenesis, and GLI1 can bind to the CDK2 promoter and activate its expression. Although the Urso-Aspirin combination downregulates GLI1, the GLI1 overexpression not only abrogates the effect of this combination on proliferation but it also restores CDK-2 expression. These findings support that the chemopreventive effect of the Urso-Aspirin combination occurs, at least in part, through a novel GLI1-CDK2–dependent mechanism. To further understand the regulation of CDK2 by GLI1, both pharmacologic and RNAi-mediated approaches show that GLI1 is a transcriptional activator of CDK2, and this regulation occurs independent of Smoothened, the central transducer of the Hedgehog canonical pathway. Collectively, these results identify a novel GLI1-to-CDK2 pathway in esophageal carcinogenesis, which is a bona fide target for effective combinatorial chemoprevention with Urso and Aspirin. Cancer Res; 70(17); 6787–96. ©2010 AACR.

Sp1 is one of the best characterized transcriptional activators. The biological importance of Sp1 is underscored by the fact that several hundreds of genes are thought to be regulated by this protein. However, during the last 5 years, a more extended family of Sp1-like transcription factors has been identified and characterized by the presence of a conserved DNA-binding domain comprising three Krüppel-like zinc fingers. Each distinct family member differs in its ability to regulate transcription, and, as a consequence, to influence cellular processes. Specific activation and repression domains located within the N-terminal regions of these proteins are responsible for these differences by facilitating interactions with various co-activators and co-repressors. The present review primarily focuses on discussing the structural, biochemical and biological functions of the repressor members of this family of transcription factors. The existence of these transcriptional repressors provides a tightly regulated mechanism for silencing a large number of genes that are already known to be activated by Sp1.