Analysis of Nucleocytoplasmic Shuttling of NFB Proteins in Human Leukocytes

Controlled nucleocytoplasmic localization regulates activity of NFκB as well as other transcription factors. Analysis of the nucleocytoplasmic protein shuttling has been greatly facilitated by the use of leptomycin B (LMB), an inhibitor of CRM1-dependent nuclear export. The authors have previously shown that LMB inhibits NFκB activity in human neutrophils by increasing the nuclear accumulation of NFκB inhibitor, IκBα. In this chapter, the authors describe a protocol that uses LMB to study the nucleocytoplasmic shuttling of IκBα in human macrophage-like U937 cells, thus inhibiting NFκB activity. This protocol should be readily adaptable to analyze the nucleocytoplasmic shuttling of other proteins in human leukocytes.