SATB1 collaborates with loss of p16 in cellular transformation.

Tumor progression is associated with invasiveness and metastatic potential. The special AT-rich binding protein 1 (SATB1) has been identified as a key factor in the progression of breast cancer cells to a malignant phenotype and is associated with progression of human tumors. In normal development, SATB1 coordinates gene expression of progenitor cells by functioning as a genome organizer. In contrast to progenitor and tumor cells, SATB1 expression in nontransformed cells is not compatible with proliferation. Here we show that SATB1 expression in mouse embryonic fibroblasts induces cell cycle arrest and senescence that is associated with elevated p16 protein levels. Deletion of p16 overcomes the SATB1-induced senescence. We further provide evidence for an interaction of SATB1 with the retinoblastoma (RB)/E2F pathway downstream of p16. A combined deletion of the RB proteins, RB, p107 and p130 (triple-mutant; TM), prevents SATB1-induced G1 arrest, which is restored upon the reintroduction of RB into SATB1-expressing TM fibroblasts. SATB1 interacts with the E2F/RB complex and regulates the cyclin E promoter in an E2F-dependent manner. These findings demonstrate that p16 and the RB/E2F pathway are critical for SATB1-induced cell cycle arrest. In the absence of p16, SATB1 causes anchorage-independent growth and invasive phenotype in fibroblasts. Our data illustrate that p16 mutations collaborate with the oncogenic activity of SATB1. Consistent with our finding, a literature survey shows that deletion of p16 is generally associated with SATB1 expressing human cell lines and tumors.

Tumor progression induced by the loss of E-cadherin independent of beta-catenin/Tcf-mediated Wnt signaling.

E-cadherin-mediated cell-cell adhesion is frequently lost during the development of malignant epithelial cancers. Employing a transgenic mouse model of beta-cell carcinogenesis (Rip1Tag2) we have previously shown that the loss of E-cadherin is a rate-limiting step in the progression from adenoma to carcinoma. However, the mere loss of cell adhesion may not be sufficient and additional signals are required to cause tumor cells to permeate the basal membrane and to invade surrounding tissue. Besides being an important component of the E-cadherin cell-adhesion complex, beta-catenin plays a critical role in canonical Wnt signaling. We report here that beta-catenin-mediated Wnt signaling does not contribute to tumor progression in Rip1Tag2 mice. E-cadherin downregulates beta-catenin/Tcf-mediated transcriptional activity by sequestrating beta-catenin into E-cadherin cell-adhesion complexes even in the presence of activated Wnt signaling. Upon loss of E-cadherin expression, beta-catenin is degraded and Tcf/beta-catenin-mediated transcriptional activity is not induced. Moreover, forced expression of constitutive-active beta-catenin or genetic ablation of Tcf/beta-catenin transcriptional activity in tumor cells of Rip1Tag2 transgenic mice does not affect tumor progression. Together, the data indicate that signals other than beta-catenin/Tcf-mediated Wnt signaling are induced by the loss of E-cadherin during tumor progression in Rip1Tag2 transgenic mice.

Can molecular mechanisms of biological processes be extracted from expression profiles? Case study: endothelial contribution to tumor-induced angiogenesis.

Whereas the genome contains all potential developmental programs, expression profiles permit the determination of genes that are actively transcribed under defined physiological conditions. In this article, the idea of extracting biological mechanisms from expression data is tested. Molecular processes of the endothelial contribution to angiogenesis are derived from recently published expression profiles. The analysis reveals the sensitivity limits of experimental detection of transcriptional changes and how sequence-analytic techniques can help to identify the function of genes in question. We conclude that the transcripts (http://mendel.imp.univie.ac.at/SEQUENCES/TEMS/) found to be up-regulated in angiogenesis are involved in extracellular matrix remodeling, cellular migration, adhesion, cell-cell communication rather than in angiogenesis initiation or integrative control. Comparison with tissue-specific patterns of EST occurrence shows that, indeed, the presumptive tumor-specific endothelial markers are more generally expressed by cell types involved in migration and matrix remodeling processes. This exemplary study demonstrates how bioinformatics approaches can be helpful in deriving mechanistic information from diverse sources of experimental data.

Ubiquitylation in plants: a post-genomic look at a post-translational modification.

In this article, we summarize Arabidopsis genes encoding ubiquitin, ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzymes (E2s) and an additional selected set of proteins related to ubiquitylation. We emphasize comparisons to components from Saccharomyces cerevisiae, with occasional reference to animals. Among the E1 and E2s, Arabidopsis usually has two to four probable orthologs to one yeast gene. Also, Arabidopsis has genes with no likely ortholog in yeast, although they often have potential orthologs in animals. The large number of components with known function in ubiquitylation indicates that this process plays a complex role in cellular physiology.

Molecular cloning and characterization of EndoGlyx-1, an EMILIN-like multisubunit glycoprotein of vascular endothelium.

EndoGlyx-1, the antigen identified with the monoclonal antibody H572, is a pan-endothelial human cell surface glycoprotein complex composed of four different disulfide-bonded protein species with an apparent molecular mass of approximately 500 kDa. Here, we report the purification and peptide analysis of two EndoGlyx-1 subunits, p125 and p140, and the identification of a common, full-length cDNA with an open reading frame of 2847 base pairs. The EndoGlyx-1 cDNA encodes a protein of 949 amino acids with a predicted molecular mass of 105 kDa, found as an entry for an unnamed protein with unknown function in public data bases. A short sequence tag matching the cDNA of this gene was independently discovered by serial analysis of gene expression profiling as a pan-endothelial marker, PEM87. Bioinformatic evaluation classifies EndoGlyx-1 as an EMILIN-like protein composed of a signal sequence, an N-terminal EMI domain, and a C-terminal C1q-like domain, separated from each other by a central coiled-coil-rich region. Biochemical and carbohydrate analysis revealed that p125, p140, and the two additional EndoGlyx-1 subunits, p110 and p200, are exposed on the cell surface. The three smaller subunits show a similar pattern of N-linked and O-linked carbohydrates, as shown by enzyme digestion. Because the two globular domains of EndoGlyx-1 p125/p140 show structural features shared by EMILIN-1 and Multimerin, two oligomerizing glycoproteins implicated in cell-matrix adhesion and hemostasis, it will be of interest to explore similar functions for EndoGlyx-1 in human vascular endothelium.

An unexpected role for p53 in augmenting SV40 large T antigen-mediated tumorigenesis.

Simian virus 40 large T antigen transforms cells by sequestration and inactivation of the tumor suppressor proteins p53, retinoblastoma gene product (pRb), and the pRb-related proteins p107 and p130. Thus, the absence of functional p53 is expected to promote T antigen-mediated tumorigenesis. However, in a transgenic mouse model of T antigen-mediated beta cell carcinogenesis (Rip1Tag2), tumor volumes are significantly diminished when these mice are intercrossed with p53-deficient mice. Whereas the incidence of beta tumor cell apoptosis is unaffected, their proliferation rate is reduced in p53-deficient beta cell tumors in vivo and in cell lines established from these tumors in vitro. Biochemical analyses reveal higher levels of T antigen in wild-type tumor cells as compared to p53-deficient tumor cells. The data indicate that p53 stabilizes SV40 large T antigen, thereby augmenting its oncogenic potential as manifested by increased proliferation rates in wild-type beta tumor cells as compared to p53-deficient beta tumor cells.