RNA interference (RNAi) has become a powerful technique for reverse genetics and drug discovery and, in both of these areas, large-scale high-throughput RNAi screens are commonly performed. The statistical techniques used to analyze these screens are frequently borrowed directly from smallmolecule screening; however small-molecule and RNAi data characteristics differ in meaningful ways. We examine the similarities and differences between RNAi and small-molecule screens, highlighting particular characteristics of RNAi screen data that must be addressed during analysis. Additionally, we provide guidance on selection of analysis techniques in the context of a sample workflow.

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Crawling T cell locomotion in which activated lymphocyte function-associated antigen 1 (LFA-1) integrins participate is associated with translocation of the protein kinase C-beta (PKC-beta) isoenzyme to the microtubule cytoskeleton. In normal T cells and T lymphoma cell lines, this type of motility is accompanied by PKC-beta-sensitive cytoskeletal rearrangements and the formation of trailing cell extensions, which are supported by microtubules. Expression of PKC-beta(I) and enhanced green fluorescent protein (EGFP) in nonmotile PKC-beta-deficient T cells restored their locomotory behavior in response to a triggering stimulus delivered via LFA-1 and correlated directly with the degree of cell polarization. We have also shown that PKC-beta(I) is a component of the tubulin-enriched LFA-1-cytoskeletal complex assembled upon LFA-1 cross-linking. These observations may have physiological equivalents at advanced (post-integrin activation) stages of lymphocyte extravasation.

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