Editorial from Guest Editors [ High Throughput Screening Using Eukaryotic Platform Organisms Guest Editors: Hildegard Buning and Christian J. Buchholz ]

High throughput screening (HTS) has become an indispensable tool in modern drug development and has been extended from the screening of chemical libraries to the screening of biological molecules or even whole organisms. Libraries of proteins and peptides can be screened for diverse applications including enzyme optimisation, functional genomics, or therapeutic purposes like drug development or targeted delivery. Mainly for technical difficulties in generating libraries covering large repertoires of polypeptide variants platform organisms were previously restricted to the prokaryotic kingdom. The scope of this edition is to review recent developments in display technologies beyond the well known and often used phage display technology. During the past years more and more library and screening systems have been described that are based on viruses and eukaryotic cells as platforms. This way, many sophisticated screening systems have been established. In this issue of Combinatorial Chemistry & High Throughput Screening, we have assembled a collection of articles describing the state of the art for different eukaryotic display platforms and for specific screening strategies performed in eukaryotic expression systems. The first two reviews focus on the use of two enveloped viruses as screening platform, baculovirus and retrovirus (reviews one and two). Both are used in many laboratories as vectors to produce large amounts of recombinant protein in a eukaryotic cell system or to genetically modify mammalian cells. Several unique properties of these viruses make them an ideal platform also for protein display and screening. Small peptides and also large polypeptides can be displayed on the viral surface or, using the virus as vector, on the cell membrane. In each case full processing and glycosylation of the displayed protein is possible. Screening strategies using retroviruses and baculoviruses include, but are not limited to, the identification of single-chain antibodies, protein interaction partners, antigenic epitopes or protease substrate peptides. Besides screening, display of polypeptides can also be applied for other purposes such as evoking an efficient immune response, identifying ligand receptor interaction partners, or modifying the tropism of viral vectors. HTS is also being used to improve viral gene transfer vectors. Libraries of vial variants of retroviruses and adeno-associated viruses are being generated by random peptide insertions, error prone PCR, DNA shuffling or transposon-based approaches (reviews three and four). Vectors based on the natural occurring virions have been used in many clinical gene therapy trials worldwide. However, to become more efficient in gene delivery in patients, properties like particle stability, nuclear entry into quiescent cells, escape from an inactivating immune response, or receptor choice and thus biodistribution are being improved by library screening. Yeast cells have become increasingly popular as display platform during the past years. Large repertoires can be covered and screening can be facilitated through fluorescent cell sorting techniques. Yeast based display libraries have been screened for different purpose including epitope mapping, the identification of interacting protein partners, or single-chain antibodies recognising specific molecules on the surface of human cells and tissue (review five). In peptide aptamers and Znfinger domains, which are in the focus of review six and seven, amino acid residues available for random diversification and library generation are well defined. Libraries of peptide aptamers are often screened in eukaryotic cell systems to identify peptides that interfere with certain cellular pathways by binding to a target protein whereas Zn-finger domains can be engineered to bind selectively to specific sequences of DNA. By combining them with effector domains of transcription factors, designer transcription factors can be generated. Effective “designer” transcription factors can be obtained through library screening for optimal binding and activation of a defined DNA sequence or a gene of interest. We conclude this issue with a review describing a novel approach that allows the screening for gene functions based on reversely transfected cell arrays. Effects on the cell phenotype resulting from the transfection of cDNA or siRNA can be screened in multi-well dishes. Applications of this technique in functional genomics are described and a laboratory protocol is provided. In summary, we have collected several articles that discuss the most important display platforms and screening strategies in eukaryotic expression systems. Only they enable the screening for properties, that exclusively manifest in mammalian cells or that require processing of the displayed polypeptide in a eukaryotic environment. Moreover, translational problems as they occur when molecules selected in a prokaryotic expression system are transferred to mammalian or human cells are avoided. It is therefore well conceivable that especially for therapeutic purposes HTS based on eukaryotic expression systems will become more and more popular in near future.