2013 Seminars

Christmas Special CABD Seminar. December 18 @ 12:30pm
Transcriptional Precision in the Drosophila Embryo and Evolutionary Origins of Cranial Placodes in the Ciona Tadpole
Dr. Mike Levine
Professor of Genetics, Genomics and Development. University of Berkeley.
CABD Seminar December 12 @ 10am
Identification of cell polarity regulators through systematic genetic interaction screens
Dr. Josana Rodriguez
The Gurdon Institute and Department of Genetics, University of Cambridge, UK

Summary: Cell polarity and asymmetric cell divisions are essential for the generation of cellular diversity. The importance of these processes during development has been emphasized by recent findings, which show that polarity defects lead to the formation of tumors. Towards identifying new genes and functional interactions involved in cell polarity, we focused 17 large-scale genetic interaction screens on C. elegans mutants of key polarity events: actomyosin regulation, PAR protein signalling and spindle positioning/microtubule (MT) regulation. Using these data, we have generated the most comprehensive metazoan cell polarity network, containing 184 genes. The network contains most of the known and expected interactions validating our screening approach. Importantly, the majority of the interactions are new and 80% of the genes have a human homolog. Demonstrating the value of this network, we experimentally confirmed cell polarity roles for 12 genes, identifying genes involved in PAR protein localization & function, acto-myosin dynamics and in a novel functional redundancy required for cell polarity (Fievet & Rodriguez et al, Nat. Cell. Biol. 2013). In addition, the cell polarity network has identified 58 genes functionally linked to known MT regulators. Among these genes 19 are also linked to actomyosin regulators and/or PAR proteins, indicating their possible role in MT regulation and cell polarity. This set of genes contains MT binding proteins, motors and kinases & phosphatases involved in MT signalling; but the majority are unexpected. The functional characterization of these genes will significantly increase our understanding of how the cell polarity mechanisms and MT regulation are orchestrated.
CABD Seminar November 29 @ noon
Spindle Orientation: Can Epithelial Cells Get Along Without It?
Dr. Dan Bergstralh
The Gurdon Institute, Cambridge, UK

Summary: In most epithelia, cell division occurs along the plane of the tissue. So what happens if an epithelial cell divides in the wrong direction? Misoriented cell division has been suggested to promote tumorigenesis; an incorrectly positioned daughter cell, freed from constraints and cues provided by the tissue, could continue dividing in an uncontrolled manner. We are testing this model using follicle cells, which form a simple mono-layered epithelium that surrounds the developing Drosophila egg chamber. Having helped to confirm the mechanism whereby mitotic spindles are oriented in these cells, we are now investigating the consequence of its loss.
CABD Seminar November 22 @ noon
Long-distance regulatory interactions, ncRNA, cell biology and behavior: lessons from the Hox genes of the Bithorax complex in Drosophila.
Dr. François Karch
Dept. of Genetics and Evolution, Univ. of Geneva, Switzerland

Summary: Over the last three decades, tremendous progress has been achieved towards understanding the mechanisms governing the spatial and temporal control of gene expression. Although numerous regulatory elements (referred as to Cis-Regulatory-Modules (CRMs) have already been described, we still seem far from completely understanding the vast layers of complexity regulating gene expression in vivo. One of the most intriguing, new layers to be uncovered is the role of ncRNAs in gene regulation. Recently, multiple labs discovered that regulatory regions and CRMs are transcribed, giving rise to all sorts of ncRNAs (sometimes referred as to eRNAs). At present, it remains unclear if these ncRNAs play a role in the regulatory output or if they are simply the by-products of the regulatory activity. Another interesting layer of complexity exists in understanding of how the presence of CRMs influence the activity of other CRMs. While CRMs have the ability to work in isolation, they often display position effects, resulting in reporter gene expression only reminiscent of the true expression pattern of the gene. This clearly demonstrates that each element must be somehow influenced by its neighboring elements. Understanding how these elements cooperate/communicate to create a gene expression pattern is something of significant importance. To understand these complex questions, we study the regulation of the homeotic genes of the bithorax complex (BX-C) in Drosophila. Over the past two decades, we, and others, have identified a number of important regulatory elements controlling BX-C Hox gene expression, including: enhancers, chromatin domain boundary elements and maintenance elements (PREs/TREs). We started to address the question of CRM communication and showed that a set of elements, previously described as enhancers (called initiators), actually function as control centers to determine the activity state of nearby enhancers and silencers. Based on our current work, we now know that initiators cause the transcription of ncRNAs that emanate outward from the initiators. We are presently characterizing these non-coding transcripts and propose to test if these transcripts play a functional role in CRM coordination. In parallel to this work, we have recently found that a very long ncRNA (the iab-8ncRNA, 92kb-long) is important for the negative regulation of the abd-A hox gene through both a miRNA and a second, still unclear mechanism. Finally, if time allows it, I will describe our finding that Abd-B is expressed in the secondary cells of the male accessory gland where it regulates the post-mating response that is elicited in females after mating.
CABD Seminar November 15 @ noon
Growth control by the Hippo tumour suppressor pathway in Drosophila.
Dr. Nic Tapon
Cancer Research UK London Research Institute

Summary: Our work is aimed at understanding how tissue size is specified during development, which remains one of the most challenging questions in biology. In order to achieve consistent organ and body size in individuals of the same species, cell growth and cell number must be tightly controlled. Since the genes that restrict organ size are often the targets of tumour-promoting mutations, the dissection of these size control mechanisms should lead to advances in cancer therapy. Genetic screens in Drosophila melanogaster have identified the Hippo (Hpo) pathway as one of the major signalling pathways required for tissue size control. The Hpo pathway controls tissue and organ size by both inhibiting cell proliferation and promoting apoptosis. Subsequent studies in mammals have shown that this growth control function is conserved and that Hpo signalling is dysregulated in many types of cancer. At the core of the Hippo pathway lies a kinase cascade comprising the Ste20-related kinase Hpo and the Dbf2-related kinase Warts (Wts). Upon Hpo activation, the downstream kinase Wts phosphorylates and inhibits the pro-growth transcriptional co-activator Yorkie (Yki). Hpo signalling has been proposed to sense various local cues relating to cell density (contact inhibition of growth and mechanical tissue properties) or patterning (morphogen gradients), and translate these cues into a growth arrest signal once an individual tissue has reached its appropriate size. In this talk, I will discuss our latest findings on the relationship between tissue growth and mechanics, as well as the control of the Hpo pathway by the cell surface polarity determinant Crumbs.
CABD Seminar October 31 @ noon
Centrosomes in asymmetric stem cell divisions.
Dr. Jordan Raff
Sir William Dunn School of Pathology University of Oxford South Parks Road OXFORD

Summary: Centrosomes are the major microtubule organising centres in many cell types. Recent evidence has shown that certain asymmetrically dividing stem cells use a centrosome size asymmetry mechanism to ensure faithful asymmetric division. Our work suggests that an unexpectedly simple model can explain how centrosomes assemble and how centrosome size asymmetry is established.
CABD Seminar October 25 @ noon
Olfactory evolution in Drosophila.
Dr. Richard Benton
Center for Integrative Genomics UNIL (Université de Lausanne) Genopode Building CH- Lausanne

Summary: The detection of odours in the environment is universally important for primal behaviours such as feeding, mating, kin interactions and escape responses. Moreover, animal olfactory systems display enormous evolutionary capacity, as species acquire and discard olfactory receptor genes, neurons and behaviours in an ever-changing landscape of external chemical stimuli. I will present our recent insights into olfactory system evolution yielded by analysis of a recently-discovered family of olfactory receptors, the Ionotropic Receptors (IRs), and their neuronal circuits, in Drosophila.
CABD Seminar October 21 @ noon
Geometric control of cell division in fission yeast.
Dr. Sophie Martin
Department of Fundamental Microbiology University of Lausanne Biophore Building CH- Lausanne Switzerland

Summary: Size is a fundamental property of cells. A geometric model of cell length sensing has been proposed in rod-shaped fission yeast cells. This relies on intracellular gradients of the DYRK-family kinase Pom1 emanating from cell poles. These negatively regulate the SAD-like kinase Cdr2, located at mid-cell, which controls timing of mitotic commitment via Wee1 inhibition. Pom1 also restricts Cdr2 localization to position division medially through Mid1/Anillin recruitment. I will present data showing that distinct thresholds of Pom1 activity define the timing and positioning of division. I will also describe quantitative analyses of cortical Pom1 and Cdr2 profiles, which reveal lower overlap than previously thought. I will discuss possible models in which Pom1 gradients both delineate a medial Cdr2 domain and control its activity.
CABD Seminar October 18 @ noon
Dr. Leonor Saude
Instituto de Medicina Molecular. Universidade de Lisboa
CABD Seminar October 15 @ noon
Cell plasticity and differentiation in the Drosophila nervous system.
Dr. Angela Giangrande
IGBMC, Strasbourg, France

Summary: Multipotency and lineage specification are the forces that control the generation of specific, stable, fates at the correct number, place and time. Fate determinants trigger specific transcriptional programs thereby ensuring proper organogenesis, however the progression from multipotent, plastic precursors to stably differentiated cells remains poorly understood. Using the Drosophila nervous system we demonstrate that interlocked transcriptional/posttranscriptional circuits control timely and threshold activity of the glial determinant and transcription factor Glide/Gcm. This ensures gliogenesis at proper time, place and amount. Initially, the CBP lysine acetyltransferase stabilizes Glide/Gcm, which triggers the expression of the homeodomain protein Repo. Glide/Gcm and Repo positively auto/crossregulate transcriptionally, however, as Repo accumulates, it destabilizes Glide/Gcm and this is facilitated by CBP. Thus, Glide/Gcm acts as a ‘time-bomb’ triggering the gliogenic program and its own extinction. Defective homeostasis between the negative and positive loops alters the neuron:glia ratio and freezes cells in an intermediate glial/neuronal phenotype that is present in metastatic cells.
CABD Seminar September 27 @ noon
Regulation and function of the JAK/STAT signaling center in the Drosophila ovary.
Dr. Anne-Marie Pret
CNRS - Centre de Génétique Moléculaire. Gif-sur-Yvette. France
CABD Seminar July 18 @ noon
Understanding melanoma progression using the zebrafish Danio rerio.
Dr. Craig Ceol
University of Massachusetts Medical School, USA

Summary: Our laboratory focuses on major questions related to cancer initiation and maintenance through the lens of melanoma. First, we are interested in understanding how cancer arises – which genetic pathways are defective and cellular activities subverted in melanoma initiation? Second, how do solid tumors persist – which cells in a tumor and what properties of these cells allow melanomas to be maintained either in the absence of therapeutic intervention or following chemotherapy? New sequencing technologies have yielded a wealth of data describing the genomes of melanomas and other cancers. These studies are retrospective. They offer a catalog of defects present in individual cancers but do not discriminate between tumor-promoting genomic changes and those changes that arose incidentally during tumor progression. Functional studies are necessary to draw this distinction, and in many cancers the list of candidate genomic changes can be quite large. To parse through such changes we have utilized a zebrafish melanoma model. Using this model, genetic and epigenetic abnormalities that contribute to melanoma progression have been defined. In addition, this model has been used to explore the relationship between melanocyte stem cells and cells driving melanoma maintenance. Adult melanocyte stem cells have been identified, and pathways governing their activity and that of nascent melanomas uncovered. Together these studies have revealed novel regulators and mechanisms of melanoma progression.
CABD Seminar Jun 5 @ 11:00 am
How multi-tasking macrophages deal with death, damage and development.
Dr. Will Wood
Wellcome Trust Senior Research Fellow and Professor, University of Bath, UK

Summary: Immune cells live in a complex, three-dimensional environment and are exposed to multiple guidance cues that are often pulling them in opposite directions. In order to respond efficiently immune cells must prioritize competing cues and our current understanding of this impressive act of signal integration is very limited. The Wood lab uses Drosophila to investigate, in vivo, how macrophages sense and respond to the developmental guidance cues that direct their dispersal through the embryo as well as the inflammatory cues that stimulate their chemotaxis towards epithelial wounds and the ‘eat me’ cues produced by apoptotic corpses. Using the powerful genetics in Drosophila together with live imaging we are starting to unpick the molecular mechanisms that underlie this impressive multi-tasking in vivo.
CABD Seminar May 31 @ 12:30 pm
Mechanisms underlying pigmentation pattern evolution in flies: from gene network tinkering to selective forces
Dr Benjamin Prud'Homme
Institut de Biologie du Développement de Marseille-Luminy (IBDML)

Summary: The typical pattern of morphological evolution associated with the radiation of a group of related species is the emergence of a novel trait and its subsequent diversification. From butterfly eyespots and their various colorful rings to the diversity of shapes assumed by vertebrate teeth, seashells or horn beetles, this pattern of emergence-diversification holds for countless characters across most animal groups. Yet the genetic mechanisms associated with these two evolutionary steps are poorly characterized. We're studying the evolution of wing pigmentation patterns in flies to address from a gene regulatory perspective how morphological novelties (rarely) emerge and how they (often) diversify. We're also studying the function of wing pigmentation patterns in mating behavior in order to identify the possible selective mechanisms underlying the evolution of this morphological trait.
CABD Seminar May 24 @ noon
Phosphorylation cascade, loop and code in Hedgehog signalling: a smoothened situation
Dr. Anne Plessis
Institut Jacques Monod, Paris.

Summary: The transduction of the morphogen and oncogene Hedgehog requires Smoothened, a GPCR-related protein. Our goal is to understand how Smoothened activity is regulated in response to different doses of Hedgehog. Using the fly wing imaginal disc as a model, we identified and analyzed a positive feedback loop between Smoothened and the protein kinase Fused. Our data support the notion of a Smoothened phosphorylation code which allows the transduction of different doses of HH into differential signaling activity.
CABD Seminar May 17 @ noon
Hox Control of Drosophila Feeding Behaviour
Dr. Ingrid Lohmann
BIOQUANT-Zentrum, University of Heidelberg

Summary: Food uptake is a crucial behaviour of all animals, which depends on the rhythmic activity of feeding-associated muscles triggered by brain motoneurons. Despite its vital importance, critical determinants instructing the development, wiring and connectivity of neuromuscular feeding networks are largely unknown. Here we identify the Hox transcription factor Deformed (Dfd) to be expressed and functional in specific motoneurons and muscles, which are essential for food ingestion in Drosophila. Using genetic, molecular, genomic and behavioural approaches we demonstrate that Dfd is required at subsequent phases of motor network formation by directly controlling neuronal specification, axon outgrowth, synapse formation and neurotransmission. The synchronous regulation of cell adhesion molecules in feeding-associated motoneurons and muscles furthermore uncovers Dfd as a decisive factor of synaptic target recognition. Finally, we demonstrate that the Dfd homolog Hoxb4 is expressed in neurons projecting towards head muscles in the vertebrate model medaka, indicating a general and conserved role of homology group 4 Hox genes in neuromuscular network formation.
CABD Seminar May 13 @ noon
How cell know how big they are
Prof. Fred Chang
Columbia University, NY
CABD Seminar May 10 @ noon
Transcription and R-loops in genome instability
Dr. Andrés Aguillera
Universidad de Sevilla-CABIMER

Summary: Genome instability is a cell pathology characterized by high levels of mutation, recombination or chromosome loss that is commonly associated with cancer and a number of genetic diseases. One key cellular process compromising genome integrity is transcription, which can contribute to genome instability via the formation of R-loops or by collisions with the replication machinery. Deciphering the factors and mechanisms responsible for transcription-associated instability is essential to understand genome dynamics and the molecular basis by which different gene expression steps, from transcription to RNA processing and export, can threaten genome integrity.
CABD Seminar May 3 @ noon
Reflections on the Future
Dr. Peter Rigby
The Institute of Cancer Research, London; Deputy Chairman to the Wellcome Trust.

Summary: I have been asked to think about the future of our field. When I first became interested in developmental biology, almost exactly thirty years ago, there were three big questions. The first was what is the molecular basis of Lewis Wolpert’s French Flag, which roughly translates as “what is a morphogen”? The second was how are the body axes determined? And the third was how do you make a specialised cell from an uncommitted progenitor? Do we have satisfactory answers to those questions? Yes but it all depends on who you are and how much granularity you want. I think that most biologists are of the view that we know enough. On the other hand, I could easily make you a long list of things that I still want to know about how you make a skeletal muscle, which has been my obsession for twenty years. There may well be technological advances that will take the field to a new level. Ever improving microscopes, ways of getting cells to talk to us, improved proteomics,and ways of imaging molecular interactions in real time in vivo. But if we want to continue to explore fundamental developmental problems, we have to persuade both our scientific colleagues and our political masters to give us money, and that, with the notable exception of the brain,may not be easy. We will have to find ways of justifying what we want to do other than simply saying “it is interesting and important”. Applied developmental biology, stem cell biology and regenerative medicine, will be fine. But if we want to continue to do pure developmental biology we will have to focus on issues which have general applicability and are thus of interestto the wider community of biologists. I do not wish to be gloomy because I think that there is much we can do but we will ignore the issues at our peril.
CABD Seminar April 26 @ noon
Kinase crosstalk: mitotic kinases as anti-cancer targets
Dr. Mar Carmena
University of Edinburgh

Summary: Cell division is regulated by highly conserved families of mitotic kinases, among them Polo-like and Aurora kinases. Our research focuses in the study of how these kinases interact and coordinate their activities to ensure accurate cell cycle progression. In recent years the use of anti-mitotic agents in anticancer chemotherapy has proven a highly successful approach. The quest is now on to find new anti-mitotic drugs equally efficient but with less undesirable side effects. Aurora and Polo kinases have been identified as targets for the development of new anti mitotic drugs. I will describe our new screens for anti-mitotic agents using both human cells and Drosophila as model systems.
CABD Seminar April 12 @ noon

Dr Katherine Brown
Development, COB
CABD Seminar April 5 @ noon
Enhancing hexosamine pathway flux improves C. elegans protein homeostasis and extends life
Dr. Adam Antebi
Max Planck Institute for Biology of Ageing, Cologne

Summary: Organismal ageing entails the progressive loss of cellular homeostasis - including protein homeostasis, which comprises all processes maintaining a functional proteome. The endoplasmic reticulum (ER) is an important site of protein synthesis and quality control, and a particularly vulnerable point in disease. To identify mutants with improved ER quality control we isolated mutants resistant to tunicamycin, a drug which interferes with ER protein folding by inhibiting N-glycosylation. We identified gain-of-function mutations in glucosamine-fructose 6-phosphate aminotransferase (gfat-1), which is the rate limiting enzyme in the hexosamine pathway producing UDP-GlcNac for O- and N-linked glycosylation. Notably, such mutants are long-lived and improve the management of proteotoxic species. Mutants have elevated levels of UDP-GlcNac, and feeding this metabolite to wild type animals is sufficient to extend life span and ameliorate various proteinopathies. These studies reveal that endogenous metabolites can enhance health and extend life through protein quality control mechanisms.
CABD Seminar March 15 @ noon
Multiple functions of Fibroblast Growth Factor signaling in Drosophila gastrulation
Dr. Arno Muller
College of Life Sciences, Univ. of Dundee

Summary: The Muller laboratory is interested in cell signaling networks that control epithelial mesenchymal morphogenesis using Drosophila melanogaster as a model. During gastrulation of the fly embryo, and in fact many other organisms, mesoderm cells are derived from epithelial precursors and undergo a transition to mesenchymal cells, which collectively move to form a cell layer that becomes separated from the other germ layers. We used genetic screens to identify genes involved in mesoderm layer formation and identified components of an FGF signaling pathway to be central players in this process. Our studies show that FGF signaling operates at several distinct steps during mesoderm morphogenesis including epithelial mesenchymal transition, cell attachment, directional migration and cell differentiation. The screens also revealed molecular players in the FGF signaling pathway elucidating how the FGF receptor translates the extracellular signal into changes in cell behavior. The seminar will summarise the cellular roles of FGF signaling in mesoderm layer formation and discuss requirements of small GTPases of the Rho family and amino-sugar nucleotide metabolism for this morphogenetic event.
CABD Seminar March 1 @ noon
Heart Development and Regeneration
Dr. Eldad Tzahor
Weizmann Institute of Science. Department of Biological Regulation

Summary: Over the past few years we have focused on the developmental programs of the heart and skeletal muscles of the head. In contrast to our understanding of how skeletal muscle is formed in the trunk, much less is known about the tissues and molecules that induce the formation of the head musculature. Our studies over the last decade have addressed the origins, signaling, and genetics of distinct head muscles. Considerable cellular and genetic variations exist among the different craniofacial muscle groups, and their associated satellite cells. Cellular and molecular parallels are drawn between cardiac and pharyngeal muscle developmental programs, and argue for the tissues’ common evolutionary origins. During embryogenesis, parts of the heart and craniofacial muscles arise from common origins within the pharyngeal mesoderm (PM). I will present unpublished data revealing a hierarchical gene regulatory network of transcription factors expressed in PM progenitors that initiates heart and craniofacial organogenesis. Genetic ablation experiments that perturbed this network in mice resulted in severe heart and craniofacial muscle defects. We identified Lhx2, a LIM domain transcription factor, as a novel player during cardiac and pharyngeal muscle development. Lhx2 and the bHLH transcription factor Tcf21 genetically interact with Tbx1; furthermore knockout of these genes recapitulates specific features of DiGeorge/velo-cardio-facial/22q11.2 deletion syndrome. We suggest that PM-derived cardiogenesis and myogenesis are network properties, rather than properties specific to individual PM members. These findings shed new light on the developmental underpinnings of congenital heart and craniofacial defects.
CABD Seminar February 15 @ 10 am
From Networks to Pattern Formation - Computational Models of Development
Dr. Dagmar Iber
ETHZ, Zurich

Summary: Mouse organogenesis has been studied for decades. While much is known about the genes that affect organ formation, general regulatory paradigms are largely lacking to explain the emergence of functional organization in biology. A number of mathematical theories have been proposed to explain biological pattern formation, but these have rarely been tested experimentally. With the emergence of the field of systems biology the two approaches are becoming increasingly interlinked and experimentally validated computational models of mouse organogenesis that integrate the detailed experimental knowledge are being generated. I will present our recent models of branching morphogenesis and limb development. Our data-based models not only help to detect inconsistencies in experimental data, but also allow to define the underlying regulatory mechanisms and to distinguish core and accessory regulatory interactions. These models also present a useful tool for in silico genetics, i.e. the computational simulations of the expected phenotypes of mutant states that are difficult or impossible to generate by mouse molecular genetics.
CABD Seminar February 1 @ noon
Role of skeletal muscle in the epigenetic shaping of organs and cell fate choices
Prof. Boris Kablar
Dalhousie University School of Medicine, Halifax, Canada

Summary: In this presentation, I will review a few concepts in Developmental Biology, according to which, cells, tissues and organs are each other’s environment. Various cues from that environment, such as molecular (e.g., paracrine), mechanical and other cues, are thought to influence embryonic development in a reciprocal (mutual) way, creating complex networking systems that connect the genotype and the phenotype. In the first part, I will explain how lung development depends on mechanical stimuli from the respiratory musculature. I will introduce novel players in the mechanochemical signal transduction pathway that operate to connect the mechanical stimuli and the alveolar epithelial cell differentiation. The human disease that is modeled in this example is Pulmonary Hypoplasia. In the second part, I will explain the developmental relationship between skeletal muscle, motor neurons and giant pyramidal cells in the spinal cord and brain, respectively. I will introduce novel molecular players in the process of motor neuron number regulation. The human disease that is modeled in this example is Amyotrophic Lateral Sclerosis (ALS) or Lou Gehrig's Disease. In the last part, I will explain developmental relationships between skeletal muscle and the skeleton. In particular, I will concentrate on the events of fusion and secondary cartilage development. I will introduce novel molecular players involved in skeletal development, and examine new concepts explaining the human disease that is modeled here, known as Palatoschisis or Cleft Palate.
CABD Seminar January 25 @ 11 am
Sequence-based discovery of human and Drosophila regulons
Dr Stein Aerts
Laboratory of Computational Biology Center for Human Genetics University of Leuven

Summary: The identification and characterization of functional cis-regulatory elements in eukaryotic genomes remains a key challenge in genome biology. We present a computational framework to analyse a human, mouse, or fly gene network or gene signature and to confidently identify cis-regulatory modules and transcription factor (TF) binding sites. The framework uses Hidden Markov Models to identify motif clusters, combined with comparative genomics cues, rank statistics to identify enriched motifs, and a “motif2TF” step to prioritize candidate transcription factors (TF) for each enriched motif. The Drosophila version of our method (called i-cisTarget) utilizes large collections of motifs (>6000 position weight matrices) and of 'regulatory tracks' (> 500 data sets) as cues, including the entire modEncode and BDTNP data sets. The human version of our method (called iRegulon) works as a Cytoscape plugin and thereby integrates cis-regulatory sequence analysis with network biology. To illustrate our methods, we show two case studies, one in Drosophila retinal determination and the second in human cancer. For the Drosophila case study we have performed cross-species transcriptomics by next-generation sequencing across three Drosophila species and obtained a highly conserved core set of eye-specific genes. Motif and CRM discovery unveiled a regulatory network downstream of the transcription factor Glass, which we validated by RNA-Seq in glass mutant eyes and by in vivo enhancer-reporter assays. For the human case study we have performed RNA-Seq and ChIP-Seq analysis for TP53 in a breast cancer cell line and show how iRegulon successfully identifies known and novel TP53 binding sites and target genes. Encouraged by these results, we applied iRegulon to more than twenty thousand cancer gene signatures obtained both from signature databases and from bi-clustering 91 large cancer gene expression datasets, and defined for each TF a context-free “meta-targetome”. In conclusion, i-cisTarget and iRegulon are next-generation motif discovery methods that confidently identify master regulators and bona fide direct targets from sets of co-regulated genes.