A Pivotal Role of Cyclic AMP-Responsive Element Binding Protein in Tumor Progression

Abstract

Tumor microenvironment controls the selection of malignant cells capable of surviving in stressful and hypoxic conditions. The transcription factor, cyclic AMP-responsive element binding (CREB) protein, activated by multiple extracellular signals, modulates cellular response by regulating the expression of a multitude of genes. Previously, we have demonstrated that two cystein residues, at the DNA binding domain of CREB, mediate activation of CREB-dependent gene expression at normoxia and hypoxia. The construction of a dominant-positive CREB mutant, insensitive to hypoxia cue (substitution of two cystein residues at position 300 and 310 with serine in the DNA binding domain) and of a dominant negative CREB mutant (addition of a mutation in serine 133 ), enabled a direct assessment, in vitro and in vivo, of the role of CREB in tumor progression. In this work, we demonstrate both in vitro and in vivo that CREB controls hepatocellular carcinoma growth, supports angiogenesis, and renders resistance to apoptosis. Along with the identification, by DNA microarray, of the CREB-regulated genes in normoxia and hypoxia, this work demonstrates for the first time that in parallel to other hypoxia responsive mechanisms, CREB plays an important role in hepatocellular carcinoma tumor progression.

A Key Role for Cyclic AMP-Responsive Element Binding Protein in Hypoxia-Mediated Activation of the Angiogenesis Factor CCN1 (CYR61) in Tumor Cells

“…We have previously found by DNA microarray analysis that CCN1 is one of a group of angiogenesis genes that are up-regulated in response to hypoxia (10). Furthermore, CCN1 expression is increased in tumors (24).…”

“…2, 4-7). However, the transcriptional response to hypoxia is not limited to regulation by HIF-1 and other transcription factors such as the cyclic AMP (cAMP) responsive element-binding protein (CREB) may be involved in a tissue-specific manner, either independently or in cooperation with HIF-1 (8)(9)(10)(11). Recently, it was reported that CREB may regulate the HIF-1 -dependent expression of the angiogenic gene, vascular endothelial growth factor (VEGF; ref.…”

“…Moreover, we have shown that in response to hypoxia, wild-type CREB (wtCREB) increases the expression of CREB-mediated genes, whereas the CREB-mutated in cysteines at positions 300 and 310 (CREB300/310) already mimics at normoxia the increased activity of wtCREB at hypoxia (13). We have also shown that CREB plays a pivotal role in tumor growth and angiogenesis in hepatocellular carcinoma (HCC) in vivo (10).…”

“…The secretion of angiogenesis factors such as the fibroblast growth factor, VEGF, platelet-derived growth factor, interleukin-8, angiogenin, and CCN1, is enhanced in response to hypoxia (10,(14)(15)(16)(17)(18)(19)(20)(21)(22). The immediate early gene CCN1 (CYR61), a member of the CCN family, codes for a secreted heparin binding protein associated with the cell surface and extracellular matrix (23).…”

“…The mouse CCN1 promoter was found to be activated by serum components via a serum response element at position À2 kb and an Egr binding site at position À48 bp (32)(33)(34). DNA array analysis of mouse HCC cells suggested CCN1 as one of 77 candidate genes up-regulated in response to hypoxia cue (10).…”