Regulation of nucleocytoplasmic trafficking by cell adhesion receptors and the cytoskeleton

It has become widely accepted that adhesion receptors can either directly activate, or significantly modulate, many of the signaling cascades initiated by circulating growth factors. An interesting recent development is the realization that adhesion receptors and their cytoskeletal partners can regulate the trafficking of signaling proteins between the cytoplasm and nucleus. Cell adhesion molecule control of nucleocytoplasmic trafficking allows adhesion to influence many cell decisions, and highlights the diversity of nuclear import and export mechanisms

Regulation of nucleocytoplasmic trafficking by cell adhesion receptors and the cytoskeleton

It has become widely accepted that adhesion receptors can either directly activate, or significantly modulate, many of the signaling cascades initiated by circulating growth factors. An interesting recent development is the realization that adhesion receptors and their cytoskeletal partners can regulate the trafficking of signaling proteins between the cytoplasm and nucleus. Cell adhesion molecule control of nucleocytoplasmic trafficking allows adhesion to influence many cell decisions, and highlights the diversity of nuclear import and export mechanisms

Covalent conjugation of oligonucleotides with cell-targeting ligands

A continuing problem in the area of oligonucleotide-based therapeutics is the poor access of these molecules to their sites of action in the nucleus or cytosol. A number of approaches to this problem have emerged. One of the most interesting is the use of ligand-oligonucleotide conjugates to promote receptor mediated cell uptake and delivery. Here we provide an overview of recent developments regarding targeted conjugates, including use of peptides, carbohydrates and small molecules as ligands. Additionally we discuss our own experience with this approach and point out both advantages and limitations

Delivery of MDR1 Small Interfering RNA by Self-Complementary Recombinant Adeno-Associated Virus Vector

Small interfering RNAs (siRNAs) are potentially powerful tools for therapeutic gene regulation. DNA cassettes encoding RNA polymerase III promoter-driven hairpin siRNAs allow long-term expression of siRNA in targeted cells. A variety of viral vectors have been used to deliver such cassettes to cells. Here we report on the development and use of a self-complementary recombinant adeno-associated virus (scAAV) vector for siRNA delivery into mammalian cells. We demonstrate that this modified vector efficiently delivers siRNA into multidrug-resistant human breast and oral cancer cells and suppresses MDR1 gene expression. This results in rapid, profound, and durable reduction in the expression of the P-glycoprotein multidrug transporter and a substantial reversion of the drug-resistant phenotype. This research suggests that scAAV-based vectors can be very effective agents for efficient delivery of therapeutic siRNA

The role of protein tyrosine phosphorylation in integrin-mediated gene induction in monocytes

Integrin-mediated cell adhesion, or cross-linking of integrins using antibodies, often results in the enhanced tyrosine phosphorylation of certain intracellular proteins, suggesting that integrins may play a role in signal transduction processes. In fibroblasts, platelets, and carcinoma cells, a novel tyrosine kinase termed pp125FAK has been implicated in integrin-mediated tyrosine phosphorylation. In some cell types, integrin ligation or cell adhesion has also been shown to result in the increased expression of certain genes. Although it seems reasonable to hypothesize that integrin-mediated tyrosine phosphorylation and integrin-mediated gene induction are related, until now, there has been no direct evidence supporting this hypothesis. In the current report, we explore the relationship between integrin- mediated tyrosine phosphorylation and gene induction in human monocytes. We demonstrate that monocyte adherence to tissue culture dishes or to extracellular matrix proteins is followed by a rapid and profound increase in tyrosine phosphorylation, with the predominant phosphorylated component being a protein of 76 kD (pp76). Tyrosine phosphorylation of pp76 and other monocyte proteins can also be triggered by incubation of monocytes with antibodies to the integrin beta 1 subunit, or by F(ab')2 fragments of such antibodies, but not by F(ab) fragments. The ligation of beta 1 integrins with antibodies or F(ab')2 fragments also induces the expression of immediate-early (IE) genes such as IL-1 beta. When adhering monocytes are treated with the tyrosine kinase inhibitors genistein or herbimycin, both phosphorylation of pp76 and induction of IL-1 beta message are blocked in a dose-dependent fashion. Similarly, treatment with genistein or herbimycin can block tyrosine phosphorylation of pp76 and IL-1 beta message induction mediated by ligation of beta 1 integrin with antibodies. These observations suggest that protein tyrosine phosphorylation is an important aspect of integrin-mediated IE gene induction in monocytes. The cytoplasmic tyrosine kinase pp125FAK, although important in integrin signaling in other cell types, seems not to play a role in monocytes because this protein could not be detected in these cells