Does nucleolin bind the NF kappa B DNA binding motif?
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Amino Acid Motifs↗
Biomedical subjects
Lori R Bernstein
Publications and source records attributed to Lori R Bernstein.
YB-1 represses AP1-dependent gene transactivation and interacts with an AP-1 DNA sequence.
Involvement of the AP-1 (activator protein-1) transcription factor has been demonstrated previously in the regulation of cell proliferation and cell-cycle progression, in the control of cell migration, invasion and metastasis, and in signal transduction, stress responsiveness, DNA replication and DNA repair. YB-1 (Y-box-binding protein-1) has also been implicated in many of these processes. However, the mechanism by which YB-1 mediates these processes is poorly understood. In the present study, we report that overexpression of a transfected gene encoding YB-1 in human HeLa cervical carcinoma cells significantly represses the transactivation of a minimal AP-1 reporter construct in response to the tumour promoter PMA. YB-1 also represses mRNA expression and PMA-induced promoter transactivation of the endogenous AP-1 target gene encoding matrix metalloproteinase-12 (metalloelastase). YB-1 transrepression of both the minimal and matrix metalloproteinase-12 promoter reporter constructs is dependent on the AP-1 sequence. To identify new nuclear proteins that bind specifically to the AP-1 DNA-binding site, we devised a DNA-affinity-chromatography-based assay termed NAPSTER (nucleotide-affinity preincubation specificity test of recognition) and discovered a 49 kDa protein from human cancer cells that binds in a sequence-specific manner to the AP-1 DNA sequence. By tandem MS fragmentation sequencing analyses we determined that p49 is a YB-1. Immunoblotting of the NAPSTER-purified p49 protein using anti-YB-1 antibodies confirmed YB-1 binding to the AP-1 DNA sequence, as did gel mobility-supershift assays using YB-1 antibodies. This is the first report of YB-1 transrepression and interaction at the AP-1 DNA-binding site.
Base Sequence↗
Adhesion, migration, transcriptional, interferon-inducible, and other signaling molecules newly implicated in cancer susceptibility and resistance of JB6 cells by cDNA microarray analyses.
Relative expression levels of 9500 genes were determined by cDNA microarray analyses in mouse skin JB6 cells susceptible (P+) and resistant (P-) to 12-O-tetradecanoyl phorbol-13 acetate (TPA)-induced neoplastic transformation. Seventy-four genes in 6 functional classes were differentially expressed: (I) extracellular matrix (ECM) and basement membrane (BM) proteins (20 genes). P+ cells express higher levels than P- cells of several collagens and proteases, and lower levels of protease inhibitors. Multiple genes encoding adhesion molecules are expressed preferentially in P- cells, including six genes implicated in axon guidance and adhesion. (II) Cytoskeletal proteins (13 genes). These include actin isoforms and regulatory proteins, almost all preferentially expressed in P- cells. (III) Signal transduction proteins (12 genes). Among these are Ras-GTPase activating protein (Ras-GAP), the deleted in oral cancer-1 and SLIT2 tumor suppressors, and connexin 43 (Cx43) gap junctional protein, all expressed preferentially in P- cells. (IV) Interferon-inducible proteins (3 genes). These include interferon-inducible protein (IFI)-16, an Sp1 transcriptional regulator expressed preferentially in P- cells. (V) Other transcription factors (4 genes). Paired related homeobox gene 2 (Prx2)/S8 homeobox, and retinoic acid (RA)-regulated nur77 and cellular retinoic acid-binding protein II (CRABPII) transcription factors are expressed preferentially in P- cells. The RIN-ZF Sp-transcriptional suppressor exhibits preferential P+ expression. (VI) Genes of unknown functions (22 sequences). Numerous mesenchymal markers are expressed in both cell types. Data for multiple genes were confirmed by real-time PCR. Overall, 26 genes were newly implicated in cancer. Detailed analyses of the functions of the genes and their interrelationships provided converging evidence for their possible roles in implementing genetic programs mediating cancer susceptibility and resistance. These results, in conjunction with cell wounding and phalloidin staining data, indicated that concerted genetic programs were implemented that were conducive to cell adhesion and tumor suppression in P- cells and that favored matrix turnover, cell motility, and abrogation of tumor suppression in P+ cells. Such genetic programs may in part be orchestrated by Sp-, RA-, and Hox-transcriptional regulatory pathways implicated in this study.
Animals↗
Interaction of retroviral Tax oncoproteins with tristetraprolin and regulation of tumor necrosis factor-alpha expression.
BACKGROUND: The Tax oncoproteins are transcriptional regulators of viral expression involved in pathogenesis induced by complex leukemogenic retroviruses (or delta-retroviruses, i.e., primate T-cell leukemia viruses and bovine leukemia virus). To better understand the molecular pathways leading to cell transformation, we aimed to identify cellular proteins interacting with Tax. METHODS: We used a yeast two-hybrid system to identify interacting cellular proteins. Interactions between Tax and candidate interacting cellular proteins were confirmed by glutathione S-transferase (GST) pulldown assays, co-immunoprecipitation, and confocal microscopy. Functional interactions between Tax and one interacting protein, tristetraprolin (TTP), were assessed by analyzing the expression of tumor necrosis factor-alpha (TNF-alpha), which is regulated by TTP, in mammalian cells (HeLa, D17, HEK 293, and RAW 264.7) transiently transfected with combinations of intact and mutant Tax and TTP. RESULTS: We obtained seven interacting cellular proteins, of which one, TTP, was further characterized. Tax and TTP were found to interact specifically through their respective carboxyl-terminal domains. The proteins colocalized in the cytoplasm in a region surrounding the nucleus of HeLa cells. Furthermore, coexpression of Tax was associated with nuclear accumulation of TTP. TTP is an immediate-early protein that inhibits expression of TNF-alpha at the post-transcriptional level. Expression of Tax reverted this inhibition, both in transient transfection experiments and in stably transfected macrophage cell lines. CONCLUSION: Tax, through its interactions with the TTP repressor, indirectly increases TNF-alpha expression. This observation is of importance for the cell transformation process induced by leukemogenic retroviruses, because TNF-alpha overexpression plays a central role in pathogenesis.
Animals↗