Akirins are highly conserved nuclear proteins required for NF-kappaB-dependent gene expression in drosophila and mice.

During a genome-wide screen with RNA-mediated interference, we isolated CG8580 as a gene involved in the innate immune response of Drosophila melanogaster. CG8580, which we called Akirin, encoded a protein that acted in parallel with the NF-kappaB transcription factor downstream of the Imd pathway and was required for defense against Gram-negative bacteria. Akirin is highly conserved, and the human genome contains two homologs, one of which was able to rescue the loss-of-function phenotype in drosophila cells. Akirins were strictly localized to the nucleus. Knockout of both Akirin homologs in mice showed that one had an essential function downstream of the Toll-like receptor, tumor necrosis factor and interleukin (IL)-1beta signaling pathways leading to the production of IL-6. Thus, Akirin is a conserved nuclear factor required for innate immune responses.

Identification of JAK/STAT signalling components by genome-wide RNA interference.

Signalling pathways mediating the transduction of information between cells are essential for development, cellular differentiation and homeostasis. Their dysregulation is also frequently associated with human malignancies. The Janus tyrosine kinase/signal transducer and activator of transcription (JAK/STAT) pathway represents one such signalling cascade whose evolutionarily conserved roles include cell proliferation and haematopoiesis. Here we describe a systematic genome-wide survey for genes required for JAK/STAT pathway activity. Analysis of 20,026 RNA interference (RNAi)-induced phenotypes in cultured Drosophila melanogaster haemocyte-like cells identified interacting genes encoding 4 known and 86 previously uncharacterized proteins. Subsequently, cell-based epistasis experiments were used to classify these proteins on the basis of their interaction with known components of the signalling cascade. In addition to multiple human disease gene homologues, we have found the tyrosine phosphatase Ptp61F and the Drosophila homologue of BRWD3, a bromo-domain-containing protein disrupted in leukaemia. Moreover, in vivo analysis demonstrates that disrupted dBRWD3 and overexpressed Ptp61F function as suppressors of leukaemia-like blood cell tumours. This screen represents a comprehensive identification of novel loci required for JAK/STAT signalling and provides molecular insights into an important pathway relevant for human cancer. Human homologues of identified pathway modifiers may constitute targets for therapeutic interventions.

When to file for a patent? The scientist's perspective.

Changes in academic systems with respect to intellectual property (IP) as well as the increasing demand for external funding for high-tech research have led to a more and more prominent desire in the scientific environment to pursue both publishing and patenting. This article looks at the current state of the disclosure requirements in the context of patenting of life sciences inventions. This is done with the aim of providing some practical guidelines for researchers as to when an invention has been made and at what point in time it may be worth/reasonable to start filing a patent application, i.e. when there is sufficient data and information to allow a reasonable expectation of success.

A large-scale RNAi screen identifies Deaf1 as a regulator of innate immune responses in Drosophila.

Innate immune signalling pathways are evolutionarily conserved between invertebrates and vertebrates. The analysis of NF-kappaB signalling in Drosophila has contributed important insights into how organisms respond to infection. Nevertheless, significant gaps remain in our understanding of how the activation of intracellular signalling elicits specific transcriptional programs. Here we report a genome-wide RNA interference survey for transcription factors that are required for Toll-dependent immune responses. In addition to the NF-kappaB homologs Dif, Dorsal and factors of the general transcription machinery, we identified Deformed Epidermal Autoregulatory Factor 1 (Deaf1) to be required for the expression of the Toll target gene Drosomycin in cultured cells and in Drosophila in vivo. We show that Deaf1 is required for the survival of flies after fungal, but not E. coli, infection. We determine that Deaf1 acts downstream of the NF-kappaB factors Dorsal and Dif. These results indicate that Deaf1 is an important contributor to innate immune responses in vivo.

An RNA interference screen identifies Inhibitor of Apoptosis Protein 2 as a regulator of innate immune signalling in Drosophila.

Innate immunity in vertebrates and invertebrates is of central importance as a biological programme for host defence against pathogenic challenges. To find novel components of the Drosophila immune deficiency (IMD) pathway in cultured haemocyte-like cells, we screened an RNA interference library for modifiers of a pathway-specific reporter. Selected modifiers were further characterized using an independent reporter assay and placed into the pathway in relation to known pathway components. Interestingly, the screen identified the Inhibitor of Apoptosis Protein 2 (IAP 2) as being required for IMD signalling. Whereas loss of DIAP 1, the other member of the IAP protein family in Drosophila, leads to apoptosis, we show that IAP 2 is dispensable for cell viability in haemocyte-like cells. Cell-based epistasis experiments show that IAP 2 acts at the level of Tak 1 (transforming growth factor-beta-activated kinase 1). Our results indicate that IAP gene family members may have acquired other functions, such as the regulation of the tumour necrosis factor-like IMD pathway during innate immune responses.

Genome-wide RNAi as a route to gene function in Drosophila.

With the sequencing of the human genome and the genomes of most major model organisms completed, the systematic characterisation of gene functions remains a key challenge. During the past few years, RNA interference (RNAi) has become a powerful tool to silence the expression of genes and analyse their loss-of-function phenotype when mutant alleles are not available. Genome-wide RNAi screens against all predicted genes have been successfully used to dissect a variety of biological processes in Caenorhabditis elegans. Recently, a genome-wide library of double-stranded RNAs, that target every gene in the Drosophila genome and that is suitable for high throughput cell-based assays, was published. In this paper, recent advances will be summarised. Screening strategies and applications as a route to comprehensively characterising gene function will be discussed.