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Viruses are intracellular parasites that commandeer cellular processes, such as RNA processing or protein synthesis, to perform virus-specific functions. For this purpose, many viral proteins shuttle between the nuclear and cytoplasmic compartments, even when the viral genome is replicated in the cytoplasm. This shutding process is usually regulated by classical nuclear import and export signals (NLSs and NESs, respeaively), which are also found in many cellular proteins and are described elsewhere in this book. In this Chapter, we will focus on viruses that replicate in the nucleus, and in particular on the mechanisms by which they transport their genomes into and out of the nucleus.

Although the nuclear import of DNA may not be a normal event in the cell, mechanisms do exist for its transport. Some of these have evolved over a billion years, as viruses and other pathogens have perfected ways to invade the host, while others appear to be fortuitous piracy, as in the case of the SV40 enhancer which binds to proteins on their way to the nucleus. Regardless, the mechanism is the same: NLS-containing proteins, either provided by the host or pathogen, bind to the DNA and target it to the nucleus. The goal of all gene therapy approaches is to target enough DNA to the nuclei of cells to obtain sufficient expression for a therapeutic effect. As is well accepted, one of the major barriers to this goal is the nuclear envelope and our relative inability to target substantial amounts of DNA to the nucleus. By characterizing and understanding the mechanisms of DNA nuclear import we can begin to exploit these pathways to increase the nuclear targeting of genes for transfection, transgenic plant production, and ultimately, gene therapy.

It is impossible to detail all of the techniques used in the nuclear transport field within the allotted space. Since some methods (EM, cell and molecular biology, etc.) are dealt with in other Chapters, here I will focus on the approaches that are less familiar to the cell and molecular biologist (e.g., patch-clamp and atomic force microscopy). I believe that some of the observations I make are significant when interpreting experimental measurements.