RNAi screening and the use of small silencing RNAs for specific gene
knockdown has revolutionized basic science and translational medicine, both through
the discovery of novel gene function and as a means to perturb disease-causing genes
for therapeutic intervention. Availability of genome-wide RNAi libraries has made it
possible to screen, in an unbiased manner, for all genes involved in any cellular process.
This promises a more comprehensive understanding of complex cellular response
networks, a fundamental goal in the emerging field of systems biology. Despite the
obvious potential of this technology, cells of the immune system pose certain challenges
for application of large scale RNAi screening, particularly in balancing the efficient
delivery of silencing RNA while avoiding non-specific immune responses to the
introduced nucleic acid. However, recent advancements in RNAi technology,
improvements in delivery methods and the development of robust screening assays have
made this technology more accessible to immunologists. Consequently, several
examples of successful application of RNAi screening at both genome and sub-genome
scales in immune cells are emerging, and are significantly advancing our knowledge of
immune cell function. In this chapter, we outline the major challenges of using large
scale RNAi screening in hematopoietic cells and describe different methodologies and
assays that have been adopted for screening, with an emphasis on how these published
studies have advanced our understanding of the immune system in health and disease.
We conclude with a discussion of future opportunities and screening approaches that
will realize the potential of RNAi screening in immune cells.
Keywords: Assay design, electroporation, Hematopoietic cells, immune cells, innate
immune response, nucleofection, RNAi screening, siRNA delivery, viral shRNA.
