Insulin-like growth factor (IGF)-binding protein 1 (IGFBP-1) contains an Arg-Gly-Asp (RGD) integrin recognition sequence. In vitro mutagenesis was used to alter this RGD sequence to Trp-Gly-Asp (WGD). Migration of Chinese hamster ovary (CHO) cells expressing the wild-type protein was more than 3-fold greater in 48 hr compared with ceUs expressing the WGD mutant form of IGFBP-1. Similarly, wild-type IGFBP-1 added to the media of control CHO cells stimulated migration 2-fold compared with the WGD protein.A synthetic RGD-containing peptide, when added to the medium with wild-type IGFBP-1, blocked the effect of IGFBP-1 on cell migration. The addition of IGF-I to the culture medium had no effect on the migration of cells expressing IGFBP-1 or vector alone. Affmity chromatography of 12I-labeled CHO cel membrane proteins, using IGFBP-1 coupled to agarose, iden- These studies demonstrate that IGFBP-1 stimulates CHO cell nmgration and binds to the a4s31 integrin receptor, both by an RGD-dependent mechanism. The effect of IGFBP-1 on migration is independent of IGF-I and is probably mediated through the a43.B integrin.The insulin-like growth factor (IGF)-binding proteins (IGFBPs) are a family of homologous but distinct proteins that specifically bind IGF-I and IGF-II with high affinity and are ubiquitous in biologic fluids, tissues, and the extracellular matrix of most cell types (1). Currently six IGFBPs have been purified, cloned, and sequenced (2,3). The roles of these proteins in intercellular transport, extracellular localization, and the modulation of the actions of IGF-I and IGF-II are areas of active study. The major focus of studies published to date has been to examine the effects of the IGFBPs on modifying IGF physiology. However, few direct non-IGFmediated effects of a purified form of IGFBP on cellular functions have been reported. IGFBP-1 is a phosphorylated protein of approximately 25 kDa that is expressed in greatest amounts during fetal development. It has been reported to potentiate (4) or inhibit (5) IGF actions, depending upon the experimental conditions and degree of IGFBP-1 phosphorylation (6). IGFBP-1 contains an Arg-Gly-Asp (RGD) integrin recognition sequence (7,8) and has been shown to bind to cell surfaces (9). The specific mechanism by which IGFBP-1 or any of the other IGFBPs bind to cell surfaces has not been previously described.We undertook these studies after observing that transfected Chinese hamster ovary (CHO) cells expressing humanThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.IGFBP-1 appeared to spread and migrate on tissue culture plates more rapidly than CHO cells not expressing IGFBP-1. Cell migration is known in many cell types to involve cellular integrin receptor binding to RGD sequences in extracellular matrix proteins (10). Since IGFBP-1 binds to cell surfaces and contains an RGD sequence, we sought to determine ...

Loss of p53 function by mutation is common in cancer. However, most natural p53 mutations occur at a late stage in tumor development, and many clinically detectable cancers have reduced p53 expression but no p53 mutations. It remains to be fully determined what mechanisms disable p53 during malignant initiation and in cancers without mutations that directly affect p53. We show here that oncogenic signaling pathways inhibit the p53 gene transcription rate through a mechanism involving Stat3, which binds to the p53 promoter in vitro and in vivo. Site-specific mutation of a Stat3 DNA-binding site in the p53 promoter partially abrogates Stat3-induced inhibition. Stat3 activity also influences p53 response genes and affects UV-induced cell growth arrest in normal cells. Furthermore, blocking Stat3 in cancer cells up-regulates expression of p53, leading to p53-mediated tumor cell apoptosis. As a point of convergence for many oncogenic signaling pathways, Stat3 is constitutively activated at high frequency in a wide diversity of cancers and is a promising molecular target for cancer therapy. Thus, repression of p53 expression by Stat3 is likely to have an important role in development of tumors, and targeting Stat3 represents a novel therapeutic approach for p53 reactivation in many cancers lacking p53 mutations.The p53 protein is a potent inhibitor of cell growth, arresting cell cycle progression at several points and inducing apoptosis of cells undergoing uncontrolled growth (23,24). It has been well documented that the Ras and Myc oncogenes activate p53 by inhibiting degradation of p53 protein and that transformation by these oncogenes requires mutation of p53 itself or silencing of ARF expression in cultured cells and animal models (19,22). The critical role of p53 as an important tumor suppressor is further underscored by the fact that p53 is the most commonly altered gene in cancer. However, p53 mutation is often a late event in malignant progression (2), and many clinically detectable cancers without p53 mutations exhibit reduced p53 expression (33,36). In breast cancer, for example, 80% of the tumors do not have p53 mutations and a 5-to 10-fold reduction of the p53 mRNA level is found in tumor relative to normal breast cells and tissues (36). These observations indicate the importance of mechanisms to either block p53 activity or silence p53 expression during malignant initiation and progression. Indeed, it has been shown that the oncogenic potential of simian virus 40 (SV40) large T antigen depends on its ability to negatively regulate p53 activity, providing a mechanism by which oncoproteins inhibit p53 function in the absence of p53 mutations (3,16,34,38). Moreover, lack of HOX5A, a p53 transcription activator, has been shown to contribute to the inhibition of p53 expression in breast cancer (36).Several recent studies have reported that the c-Src tyrosine kinase opposes p53 activity during platelet-derived growth factor (PDGF)-induced mitogenesis (7,18). Because the requirement for c-Src in PDGF receptor (P...

Our recent study demonstrated miR-15a/16-1 downregulation in mantle cell lymphoma (MCL). Here, we investigated mechanisms of miR-15a/16-1 transcriptional repression and its epigenetic regulation by c-Myc and histone deacetylase (HDAC) in MCL. c-Myc expression was detected in MCL cell lines and in the primary MCL samples, and pri-miR-15a/16-1 mRNAs were significantly upregulated in Mino and Jeko-1 cells with c-Myc knockdown by small interfering RNAs (siRNAs). Our co-immunoprecipitation analysis showed that c-Myc interacted with HDAC3. Moreover, using chromatin immunoprecipitation, we demonstrated that both c-Myc and HDAC3 co-localized to the two promoters of the miR-15a/16-1 cluster gene, DLEU2, and inhibition of HDAC3 increased histone acetylation of the DLEU2 promoters. Luciferase reporter assay confirmed the dependence of Myc-mediated DLEU2 transcriptional repression on HDAC3. Treatment with the pan-HDAC inhibitor, suberoylanilide hydroxamic acid and HDAC3 siRNA resulted in increased miR-15a/16-1 expression. The regulatory mechanism of miR-15a/16-1 was further demonstrated in Burkitt lymphoma and Myc overexpressing cell lines. These findings highlight the role of HDAC3 in Myc-induced miR-15a/16-1 changes and reveal novel mechanisms for c-Myc-driven microRNA suppression and malignant transformation in aggressive B-cell malignancies.