A global analysis of genetic interactions in Caenorhabditis elegans

Abstract Background Understanding gene function and genetic relationships is fundamental to our efforts to better understand biological systems. Previous studies systematically describing genetic interactions on a global scale have either focused on core biological processes in protozoans or surveyed catastrophic interactions in metazoans. Here, we describe a reliable high-throughput approach capable of revealing both weak and strong genetic interactions in the nematode Caenorhabditis elegans. Results We investigated interactions between 11 'query' mutants in conserved signal transduction pathways and hundreds of 'target' genes compromised by RNA interference (RNAi). Mutant-RNAi combinations that grew more slowly than controls were identified, and genetic interactions inferred through an unbiased global analysis of the interaction matrix. A network of 1,246 interactions was uncovered, establishing the largest metazoan genetic-interaction network to date. We refer to this approach as systematic genetic interaction analysis (SGI). To investigate how genetic interactions connect genes on a global scale, we superimposed the SGI network on existing networks of physical, genetic, phenotypic and coexpression interactions. We identified 56 putative functional modules within the superimposed network, one of which regulates fat accumulation and is coordinated by interactions with bar-1(ga80), which encodes a homolog of β-catenin. We also discovered that SGI interactions link distinct subnetworks on a global scale. Finally, we showed that the properties of genetic networks are conserved between C. elegans and Saccharomyces cerevisiae, but that the connectivity of interactions within the current networks is not. Conclusions Synthetic genetic interactions may reveal redundancy among functional modules on a global scale, which is a previously unappreciated level of organization within metazoan systems. Although the buffering between functional modules may differ between species, studying these differences may provide insight into the evolution of divergent form and function

Role of fibroblast growth factor receptor in regulation of membrane traffic

Several studies show that membrane transport mechanisms are regulated by signalling molecules. Recently, genome-wide screen analyses in C.elegans have enabled scientists to identify novel regulators in membrane trafficking and also signalling molecules which are found to couple with this machinery. Fibroblast growth factor (FGF) via binding to fibroblast growth factor receptor (FGFR) mediate signals which are essential in the development of an organism, patterning, cell migration and tissue homeostasis. Impaired FGFR-mediated signalling has been associated with various developmental, neoplastic, metabolic and neurological diseases and cancer. In this study, the potential role of FGFR-mediated signalling pathway as a regulator of membrane trafficking was investigated. The GFP-tagged yolk protein YP170-GFP trafficking was analysed in worms where 1) FGFR signalling cascade components were depleted by RNAi and 2) in mutant animals. From these results, it was found that the disruption of the genes egl-15 (FGFR), egl-17(FGF), let-756(FGF), sem-5, let-60, lin-45, mek-2, mpk-1 and plc-3 lead to abnormal localization of YP170-GFP, suggesting that signalling downstream of FGFR via activation of MAPK and PLC-γ pathway is regulating membrane transport. The route of trafficking was further investigated, to pinpoint which membrane step is regulated by worm FGFR, by analysing a number of GFP-tagged intracellular membrane markers in the intestine of Wild Type (WT) and FGFR mutant worms. FGFR mutant worms showed a significant difference in the localisation of several endosomal membrane markers, suggesting its regulatory role in early and recycling steps of endocytosis. Finally, the trafficking of transferrin in a mammalian NIH/3T3 cell line was investigated to identify the conservation of these membrane trafficking regulatory mechanisms between organisms. Results showed no significant changes in transferrin trafficking upon FGFR stimulation or inhibition

Fibroblast growth factors as tissue repair and regeneration therapeutics

Cell communication is central to the integration of cell function required for the development and homeostasis of multicellular animals. Proteins are an important currency of cell communication, acting locally (auto-, juxta-, or paracrine) or systemically (endocrine). The fibroblast growth factor (FGF) family contributes to the regulation of virtually all aspects of development and organogenesis, and after birth to tissue maintenance, as well as particular aspects of organism physiology. In the West, oncology has been the focus of translation of FGF research, whereas in China and to an extent Japan a major focus has been to use FGFs in repair and regeneration settings. These differences have their roots in research history and aims. The Chinese drive into biotechnology and the delivery of engineered clinical grade FGFs by a major Chinese research group were important enablers in this respect. The Chinese language clinical literature is not widely accessible. To put this into context, we provide the essential molecular and functional background to the FGF communication system covering FGF ligands, the heparan sulfate and Klotho co-receptors and FGF receptor (FGFR) tyrosine kinases. We then summarise a selection of clinical reports that demonstrate the efficacy of engineered recombinant FGF ligands in treating a wide range of conditions that require tissue repair/regeneration. Alongside, the functional reasons why application of exogenous FGF ligands does not lead to cancers are described. Together, this highlights that the FGF ligands represent a major opportunity for clinical translation that has been largely overlooked in the West

PAR-5 is a PARty hub in the germline

As our understanding of how molecular machineries work expands, an increasing number of proteins that appear as regulators of different processes have been identified. These proteins are hubs within and among functional networks. The 14-3-3 protein family is involved in multiple cellular pathways and, therefore, influences signaling in several disease processes, from neurobiological disorders to cancer. As a consequence, 14-3-3 proteins are currently being investigated as therapeutic targets. Moreover, 14-3-3 protein levels have been associated with resistance to chemotherapies. There are seven 14-3-3 genes in humans, while Caenorhabditis elegans only possesses two, namely par-5 and ftt-2. Among the C. elegans scientific community, par-5 is mainly recognized as one of the par genes that is essential for the asymmetric first cell division in the embryo. However, a recent study from our laboratory describes roles of par-5 in germ cell proliferation and in the cellular response to DNA damage induced by genotoxic agents. In this review, we explore the broad functionality of 14-3-3 proteins in C. elegans and comment on the potential use of worms for launching a drugs/modifiers discovery platform for the therapeutic regulation of 14-3-3 function in cancer

CALCIUM/CALMODULIN DEPENDENT PROTEIN KINASE II SIGNALING AND IGFR SIGNALING ARE COUNTER-BALANCED TO REGULATE PROTEIN DEGRADATION IN C. ELEGANS MUSCLE

The signal transduction network controlling protein degradation in the striated body-wall muscle of C. elegans has been shown to be composed of stimulatory signaling from the FGF receptor and inhibitory signaling from the IGF receptor engaged in cross talk at the level of the Raf-MEK-MAPK cascade. Calcium/calmodulin-dependent protein kinase II (CaMKII) is an additional, pro-degradation input into the network at the level of Raf. Splice forms of CaMKII may play a role in effective CaMKII signaling. The IGFR/FGFR/CaMKII network regulates protein degradation via autophagy. Degradation caused by increase of the pro-degradation signal from CaMKII can be suppressed by increase of opposing IGFR signaling, and degradation caused by decrease in the IGFR signal can be suppressed by decreased CaMKII signaling. This implies that regulation of protein degradation is based on integration of multiple signals by Raf and not on absolute amounts of individual signals

The Egf-Ras-Erk Pathway and the Nkx-5/hmx Homeodomain Protein Mls-2 Promote Tube Development in the C.elegans Excretory System

Tubular epithelial cells are one of the most abundant cell types in multicellular organisms. Tubular cells transport gases and liquids, and funnel harmful excretory waste from our bodies. It is clear that Receptor Tyrosine Kinase (RTK) signaling is essential for the formation of many tubular organs such as our kidneys and blood vessels. However, which steps in tube development require RTK signaling is less well understood. The C.elegans excretory system is a primitive renal system with just three essential cells (duct, pore, and canal cells), providing a simple yet dynamic system to study tube specification and morphogenesis. In the C.elegans excretory system, we demonstrated that the EGF-Ras-Erk signaling pathway specified the excretory duct tube versus the pore tube fate. In addition, EGF-Ras-Erk signaling influenced the positions that the duct and pore cells adopted within the tubular network. And finally, after position and fate determination, EGF-Ras-Erk signaling had a continued role in maintaining organ architecture of the duct tube. Goals for future research will be to determine how EGF-Ras-ERK signaling controls these genetically distinct steps during tube development. In a separate project, I studied the Nkx5 homeodomain transcription factor, MLS-2, which was identified in a mutagenesis screen by a former graduate student in the lab. I discovered a role for MLS-2 in promoting proper cell shape of the duct and pore. mls-2 cooperated with the EGF-Ras-Erk pathway to turn on lin-48/Ovo during duct morphogenesis. I speculate that MLS-2 and other Nkx5 family members have conserved functions in promoting shape acquisition in cells that adopt complex morphologies similar to the duct and pore