Research Associate/Fellow position (3 years) to work on a BBSRC funded project investigating cell fate regulation during mammalian gastrulation in the laboratory of Dr. Ramiro Alberio (U. of Nottingham, UK), in collaboration with Prof. Jennifer Nichols (U. of Cambridge, UK) and Dr Matt Loose (U. of Nottingham).
Project description:
The project will investigate the molecular mechanisms of mammalian gastrulation. The project involves working with embryos and embryonic stem cells combined with next generation sequencing as a mean to understand cell fate decisions in early embryos. This post is suitable to candidates with experience in single cell RNA seq., embryology and micromanipulation. This post offers a unique opportunity to work in fast developing fields (stem cell biology, single cell genomics and gene editing) and to develop skills in state-of-the-art technologies.
Ideal applicant:
Highly motivated and self-driven, with a PhD (or near completion) in cell/developmental or related biological science with experience in some of the following areas: stem cell biology (preferably hESC), single cell RNA seq, gene editing, and bioinformatics. Experience in embryo dissection, generation of transgenic reporter cell lines, gene targeting and other genome editing techniques are also relevant for the project.
On the Sunday night of 2nd September of 2018, one month ago, most Brazilians were watching TV shows while a large part of our national story was burning out in the National Museum’s fire. In a few hours, a 200-year old institution and several biological, anthropological, and geological collections were consumed by the fire. Publications have been written in respectable journals and newspapers about the fact and its consequences for the whole of society (https://www.theguardian.com/world/2018/sep/03/fire-engulfs-brazil-national-museum-rio).
In this post, I would like to provide a brief personal view from a Brazilian Evolutionary Developmental Biology (Evo-Devo) researcher who also acts as the Director of one of UFRJ’s Institutes. Our Institute also hosts important scientific collections (http://www.macae.ufrj.br/nupem/index.php/colecao-de-peixes-npm), which could be or might be affected in the future if we do not improve our administrative practices.
First, as a Brazilian Evo-Devo researcher, the loss of holotype specimens from some of the most extensive invertebrate collections worldwide will not be recovered sooner or later. Some of the specimens collected by famous naturalists – such as one the Darwin´s greatest colleagues, the German Naturalist Fritz Muller – are of special interest for Evo-Devo researchers (https://onlinelibrary.wiley.com/doi/full/10.1002/jez.b.22687
), since it has been argued that many evolutionary secrets and different morphotypes might lie in the unexplored museums of the world. Unfortunately, in this situation we cannot come back and retrieve the samples and a part of Latin American Biodiversity is gone forever.
Although the reason for the fire is a case for the specialists, I would like to point out some issues which do not apply only for the National Museum of Brazil, but also for other public institutions. After the fire, some authorities and some of the public opinion of the country tried to put the responsibility onto our Rector, who has only been ahead UFRJ for three years and has been trying to obtain special funding to make the required changes in this historical building. This led to a strong response of our employees and students supporting our Rector and our democratic institution, our Federal University and the Museum, the first research institute in Brazil.
If you are a researcher from a developed country, you can´t imagine how much paperwork a Professor or a Director in a country like Brazil has to deal with to buy a simple equipment, or to develop a fire alarm system. The paperwork to hire a company or to develop such a project in Brazil impairs our scientific progress. This can be justified by the lack of qualified personal from the university, and our current low budget to hire a specialized company. In the past years, federal research money for Science has dropped over 60% and the Science and Technology Ministry has been largely neglected by the government.
To solve daily problems of infrastructure one must undergo a complex paperwork process which can take years and contain over a thousand pages and, at the end, the company might still not be hired due to lack of funding. Unfortunately, the corruption scandals in the past years led to a general impression that corruption is widespread all over the country. I can assure that most, if not all, professors and colleagues from my University are honest and, in many occasions, buy consumables for the university from their own salaries, to avoid undergoing this stressful and many times unsuccessful process of buying something using University money.
A second possibility, the donation of private money, rarely occurs. In general, wealthy people from Brazil do not donate to Universities or research institutes, as it is typical for US Universities. This situation is already changing with some great initiatives such as the Serrapilheira Institute (https://serrapilheira.org/en/), although the whole donation system is still in its infancy. Current Brazilian laws limit the use of the money that the University obtains from rents, museum tickets, donations, and any other source of funding. Money allocated in one year does not stay for the next year: it goes back to the Federal Government. Any money that enters in the University must undergo the same complex process that avoid any reasonable speed and progress that science needs. Thus, laws must change for science and technology improvement in Brazil.
Thus, although the museum tragedy cannot be solely attributed to lack of public funding and extensive inefficacy of paperwork and unreasonable laws to spend public resources by the University, these issues have contributed for the tragedy. Lastly, I believe that researchers from public universities have undermined the role and the importance of museums and collections for the Universities. I often see comments that museums are just repository of specimens, but particularly in the case of the National Museum a large part of Zoological, Anthropological, Paleontological research of our country was being carried out in this historical building. Importantly, immediately after the fire, the community of National´s Museum has risen together, and the collections are being restarted by the researchers and students.
The National Museum is ALIVE and I hope from now on, the Museum acquires the status it should never have lost, the home of our history and our knowledge, which fortunately lies in our public universities.
Living Systems Institute, University of Exeter, UK
The importance of Wnt signalling in developmental processes, wound healing and stem cell control has long been established. Historically, scientists attributed the transport of Wnt proteins from the source to the receiver cell to simple diffusion, however, this explanation did not seem sufficient to support the precise delivery of Wnt proteins required to satisfy healthy development. In 2015, our group discovered that finger-like membrane protrusions termed cytonemes delivered the Wnt proteins to responsive cells, but the governance of Wnt-positive cytonemes was yet to be elucidated. A new paper published by our group indicates that Wnt retains sovereignty over its own delivery by binding to the kinase Ror2, which acts upstream of cytoskeletal regulators to manage the formation of Wnt-positive cytonemes.
Wnt signalling is absolutely fundamental to embryogenesis. Present in all metazoans, Wnt proteins have been closely examined in many model organisms such as the fruit fly, mouse, frog, and zebrafish. Since its discovery in mice by Roel Nusse and Harold Varmus, and in parallel by Christiane Nüsslein-Volhard and Eric Wieshaus in Drosophila, it made its début as the viral insertion site int-1 and segment polarity gene Wingless combined as Wnt-1 in the eighties. In the three decades since it was unearthed, Wnt has gained traction as a family of proteins with famously important roles in embryonic development, cell survival, proliferation and stem cell regulation, which act by the formation of concentration gradients over responsive cell types.
Notwithstanding the accumulation of knowledge concerning the action of Wnt proteins, exactly how Wnt protein gradients are secreted in a controlled manner is a source of debate; several theories persist, including simple diffusion and exocytosis. Despite the appealing adherence to the principles of Occam’s Razor, it seemed that a more complex mechanism was required to explain the precision of Wnt delivery. Indeed, in 2015, our group discovered such a mechanism in the zebrafish, Danio rerio. Wnt proteins were in fact delivered from the sender cell by finger-like membrane protrusions carrying Wnt proteins at their tips, allowing full end-end control of Wnt morphogenetic signalling by the sender cell. These membrane protrusions, termed cytonemes, had earlier been suggested to carry the Dpp and Hedgehog signalling molecules in Drosophila development. Now, the function of cytonemes in the facilitation of developmental processes has been expanded to the delivery of Wnt signals in vertebrates.
Cytonemes mobilize Wnt protein
Membrane protrusions are not unusual and are commonly referred to as filopodia. The distinction between cytonemes and filopodia is the cargo transported – the signalling molecules. Both, however, are actin-dependant and, we discovered, contingent on Cdc42, a small Rho family GTPase integral to dynamic F-actin assembly when we were based at the Karlsruhe Institute of Technology (KIT) still. Manipulating the length and number of Wnt-positive cytonemes led to malformed embryos because of aberrant tissue patterning, and therefore we hypothesised that the regulation of formation, timing, number and length of Wnt-positive cytonemes must be tightly controlled in order to produce the fine balance of Wnt gradients that produce wild type tissue patterning.
Fig. 1 Cytoneme delivering Wnt8a (red) to a neighbouring cell, causing clustering of the receptor Lrp6 (green) at the membrane of the target cell.
The method and importance of Wnt gradient formation thus elucidated, we set out to determine the upstream control of the genesis of Wnt-positive cytonemes. Benjamin Mattes, PhD student in the lab, decided to examine whether a kinase could in fact be a candidate due to their multifaceted roles in cellular processes in 2016. Preliminary findings indicated that his hunch was correct, and multiple kinases induced changes in the number of membrane protrusions. To narrow down the selection of kinases, we collaborated with computer scientists at KIT who used modelling techniques to select the tyrosine kinase receptor Ror2. Unlike the other kinases identified by the screen, Ror2 is involved in both Wnt signal transduction of the non-canonical Wnt signalling pathway and in regulation of the F-actin cytoskeleton, thus uniting two key molecular functions in the building of cytonemes.
Ror2 controls Wnt cytonemes in zebrafish
To truly hammer home the importance of Ror2, Benjamin decided to try some studies in both zebrafish embryos and in vitro in zebrafish PAC2 cell cultures. The result of Ror2 overexpression was increased number of Wnt-positive cytonemes, inferring that Ror2 was involved in the nucleation of cytonemes, and also that it worked upstream of small Rho GTPase Cdc42, which had earlier been implicated in the regulation of the actin cytoskeleton used to build the protrusions via the non-canonical Wnt signalling pathway. Surprisingly, filopodia which did not carry Wnt8a-GFP did not change in number. Consistently, when Ror2 was knocked down, there was a significant decrease in the number of cytonemes and again filopodia number was unaffected. Regulators of cytonemes presented so far do interfere also with filopodia. However, our data suggests that Ror2 is a cytoneme specific regulator – the first one at least to our knowledge.
Fig. 2: Cytonemes transport Wnt protein (red) between gastric cancer cells. A halo of Wnt8a protein can be observed around these cells and its diameter correlates with the average length of their cytonemes (green).
Subsequently, Benjamin set out to investigate how exactly Ror2 was able to stimulate cytoneme formation by using image-based approaches, which showed that Wnt family member Wnt8a – which has an established role in zebrafish embryonic development – and Ror2 associate and move together in a complex. Still, he could not entirely prove that Ror2 and Wnt8a bind each other – until he collaborated with the physicist Uli Nienhaus at the KIT who works on developing novel super-resolution techniques. Over one nail-biting summer, they worked together to finally prove that Ror2 and Wnt8a bind together in the living zebrafish embryo – in membrane associations with presumably other mystery players.
At the end of this summer, the lab moved from the KIT in Germany to the newly established Living Systems Institute (LSI) in Exeter, UK. Benjamin continued with the experiments in the world-class institute immediately. In endless imaging sessions using the newly purchased confocal microscope he found that these Wnt8a-Ror2 membrane associations activate PCP signalling in ligand-producing cells, which induces the formation of cytonemes ready to deliver their cargo – Wnt proteins. Therefore, it appears that Wnt proteins retain sovereignty over their own delivery by controlling the nucleation of cytonemes. The complex gradients of Wnt proteins that dictate tissue patterning are governed by Wnt itself through binding with Ror2, activation of PCP signalling, and subsequent cytoneme nucleation.
Fig. 3: The Living Systems Institute, Exeter, UK – a world class hub of interdisciplinary research.
Wnt cytonemes in cancer tissue
The dependence of cytoneme nucleation on Ror2 signalling raises some interesting questions – for example, does Ror2 regulate Wnt signalling in other paracrine processes, such as cancer cell proliferation? Using a co-culture of AGS human gastric cancer cells, Benjamin found that Ror2 enhanced Wnt response in adjacent cells by greater nucleation of cytonemes and therefore increased cancer growth.
For some cells, paracrine Wnt signalling is required throughout their whole lifespan. To investigate whether Ror2 has a role in regulating cytoneme formation in these cells, we started a collaboration with mouse geneticist David Virshup of the Duke-NUS Medical School in Singapore. The intestinal crypt requires a constant supply of Wnt signalling, and David’s team demonstrated recently that the myofibroblasts that surround the crypt have an abundance of cytonemes. When we silenced Ror2 in myofibroblasts, they produced less cytonemes and the intestinal crypt cells that they surround die.
Here, we come to almost present day. All that remains to be said is that we are grateful that last summer, Christiane Nüsslein-Volhard visited the Living Systems Institute at Exeter as key note speaker for its official opening and remarked that she was very pleased to see that Wingless/Wnt trafficking is on its way to being finally solved, and that zebrafish are playing such a large part in that. This said, there is undoubtedly more to learn; cancer cell proliferation and the survival of the mouse intestinal crypt are just two examples where our insights into cytoneme nucleation and the Wnt signalling generated are proving to be instrumental. It seems that where Wnt signalling is concerned, it is in sickness and in health, to death us do part.
Figure 4 Scholpp lab outing 2018 – paddling on the Exe
The lab of Rohit Bose MD PhD at UCSF is hiring postdocs.The principal investigator is predominantly a lab-based assistant professor and also a practicing genitourinary medical oncologist at the UCSF Cancer Center.Both his clinical and lab focus is prostate biology, although open to related areas of study. Specifically, the lab focuses on themes of hypermutation, drug sensitization and transcription factor networks (Bose et al, Nature, 2017).
A postdoctoral fellow is sought to participate in a study to better understand the role of genetic changes on the development and dysfunction of cerebral vasculature. The successful candidate will work with Drs. Sarah Childs and Kristina Rinker, their teams and collaborators to advance our understanding of vascular dysfunction enabling future diagnostics and therapeutic strategies. The project entails the use of imaging to capture vascular characteristics and further evaluation with computational tools for quantification of the effect of genetic changes on the architecture, wall properties and blood flow in models of genetic disease. The Childs labs has extensive experience in creation of genetic models in zebrafish. We seek a creative, energetic, and self-directed postdoc for a two year term as part of the University of Calgary Eyes High Postdoctoral Fellowship program. A PhD degree in biomedical engineering or similar field within the past 3 years is desired. Experience in computational fluid dynamics required. Experience with cardiovascular systems, analysis of images, and machine learning is beneficial. Knowledge of developmental or vascular biology would be an asset.
We are located at the University of Calgary, in newly renovated labs within the Alberta Children’s Hospital Research Institute and in the Schulich School of Engineering. State of the art imaging and molecular biology facilities are available.
Interested candidates should send their CV, and a cover letter outlining their interests to Sarah Childs, schilds@ucalgary.ca. Review of applications will begin October 30th.
This is the latest dispatch from a recipient of a Development Travelling Fellowship, funded by our publisher The Company of Biologists.
Learn more about the scheme, including how to apply, here, and read more stories from the Fellows here.
Barbara Swierczek
I am a PhD student at the University of Warsaw in Poland. In my home lab, headed by Prof. Maria Anna Ciemerych-Litwinienko, I study the signalling pathways affecting pluripotency and differentiation in mouse embryonic stem cells, focusing on Wnt proteins. Wnts were first described more than 30 years ago, and since then became an intensively studied yet challenging field of cell biology. Why? First of all, there are 19 Wnt proteins found in vertebrates that activate a wide range of signalling pathways. Additionally, they often act antagonistically, creating a complicated and complex network of interactions. Next, Wnts play a key role during development, when their activity is orchestrated in a very precise manner: a defined Wnt has to be expressed at a right moment in a defined place.
It has long been appreciated that Wnts, particularly those dependent on β-catenin transduction (so called ‘canonical’ Wnts), are crucial for gastrulation, the process by which the three germ layers are formed in the embryo. However, despite well-documented function of Wnts in this process, it is not fully understood exactly how these proteins are regulated during gastrulation. Expression of Wnts can be regulated by inhibitors, other Wnts acting antagonistically or miRNAs. The latter are short (approximately 20 nucleotide long) non-coding RNAs, which block the expression of target genes. While studying Wnts and their regulation during mouse embryonic stem cell differentiation I came across some evidence that Wnt signalling can be regulated by miRNAs targeting either Wnts themselves or Wnt receptors or transducers. I found it very interesting and decided to pursue this link further. Also, I wanted to learn how to study this link in a developing embryo.
Around this time, I heard that the The Company of Biologists funds Travelling Fellowships for a short research visit. I decided to give it a go, and applied for three-month long fellowship in the Professor Andrea Münsterberg’s lab at the University of East Anglia in Norwich, Great Britain. Professor Münsterberg is a leading specialist in miRNA functions during development, especially their functions in modulating signalling pathways in skeletal muscle and the heart. During my stay in Norwich I had an opportunity to work on a model organism which was different for me: a chick embryo. Chick is a potent model: in contrast to mammalian development which occurs mostly within the female’s body, chick development can be studied and manipulated easily in the egg, even during later stages of development. Working with a new model was quite a challenge at first, but also an exciting perspective in the same time.
Me isolating embryos
UEA in Spring
UE covered in snow
Goodbye party
My project concerned the role of miRNAs in regulating Wnt pathway in the gastrulating chick embryo. I focused on Wnt5a, which is a double-faced Wnt: it can bind to different receptors and activate both canonical and noncanonical signalling pathways. I studied the relationship between this Wnt and an miRNA which I selected as a putative regulator as a result of sequence analyses. During my fellowship I learned how to isolate chick embryos and how to manipulate them. Since the group of Professor Münsterberg studies different aspects of chick development, I became familiar with various stages of this process, from gastrulation to more advanced stages. I was able to learn a lot from people in the group: real specialists in working on chick development. Also, I learned a lot about miRNAs and the methods of miRNA analysis which will be beneficial for my future research.
Thanks to my short-term fellowship at UEA, I had a great opportunity to experience an international scientific environment. I enjoyed my stay in Norwich which is not a big city – there are about 100,000 people living there – and for this reason a peaceful and quiet one. Thanks to the Development Travelling Fellowship I was able to learn a lot about the techniques I had not used before, something that encouraged me to seek new challenges for my future research. But what I think is the most important aspect of my visit was making new connections and meeting people working in the field. I wanted to thank once again everyone in the Münsterberg group for helping me – I hope to work with you again!
From the MRC Weatherall Institute of Molecular Medicine blog.
Stem cell turnover and tissue maintenance is a stochastic process. This means that a randomly occurring mutation has an unknown chance of becoming fixed and spreading within a tissue. Clonal mutations have been observed in apparently healthy tissue, increase in frequency with age and – in some cases – have been described as a pre-malignant state (e.g. clonal haematopoiesis). In certain tissues, such as the colonic epithelium, the contribution of mutations in stem cells to neoplastic transformation remains unclear.
This process is a major interest of Ed Morrissey, who joined the MRC WIMM Centre for Computational Biology in late 2016. His group has recently published a mathematical model that aims to address how functional mutations can contribute to altered stem cell dynamics, with the hope of understanding precisely how these rare mutations accumulate in the lead up to cancer.
The Bloomekatz laboratory in the Department of Biology at the University of Mississippi in Oxford, MS is seeking a research associateto assist in our investigations of the fundamental mechanisms underlying cardiac morphogenesis and disease using zebrafish. Please see our website thebloomekatzlaboratory.org for further details on our research. The successful candidate will have an opportunity to be involved in all aspects of our dynamic innovative research program; from experimental design to data analysis and publication. Duties may involve – conducting developmental/cell biological experiments, analyzing imaging and next-generation sequencing data, zebrafish husbandry, and mentoring undergraduate laboratory members.The candidate will work closely with and be trained by Dr. Bloomekatz. Interested in joining our group, apply online: https://careers.olemiss.edu, keyword cardiac.Salary dependent on experience. This position is eligible for benefits. The University of Mississippi is an EOE/AA/Minorities/Females/Vet/Disability/Sexual Orientation/Gender Identity/Title VI/Title VII/Title IX/ADA/ADEA employer.
The entire genome of many species has now been sequenced, but the function of the majority of genes still remains unknown. This is where the International Mouse Phenotyping Consortium (IMPC) comes in, with the goal of characterising all 20,000 or so protein-coding mouse genes. To achieve this, genes are systematically inactivated then mice are put through a standardised phenotyping platform, with tests undertaken across a broad range of biological systems.
The consortium is comprised of 19 research institutions, 5 national funders and 11 countries. Each centre focuses on particular genes, applies standardised tests and then records the resulting data. After this, phenotype analysis is conducted and the resulting data and statistics made freely available to the research community. As well as completing large scale comparative studies, the overall aim of the project is to create a platform for this data where researchers and clinicians can search for genes, phenotypes or diseases of interest to help them understand human biology, health and disease.
Professor Steve Brown, the IMPC chair says “The IMPC is rising to the challenge of generating a complete functional catalogue of the mouse genome. Since its inception in 2011, it has made great strides with a third of the genome already analysed. Moreover, many startling and hitherto undiscovered features of the mammalian genome landscape have been revealed.”
There are now over 6,000 genes with mouse mutant data on an isogenic genetic background (C57BL/6N) on the IMPC website, all of which can be viewed and downloaded for free. In its initial stages the knock out lines used for IMPC were all made in ES cells by homologous recombination, all containing a lacZ reporter and many of them generating conditional mouse lines. However, as for many areas of developmental biology, new gene editing technologies, in particular CRISPR/Cas9, have condensed the process of generating knockout mouse lines. Advancements such as this have improved production and will allow all 20,000 genes to be characterised in the next few years.
Around a third of knockout genes are embryonic lethal and consequently developmental biology is an integral part of the IMPC project. In particular, there is an extensive embryo phenotyping pipeline that includes systematic harvest of embryos at set stages, capture of morphology by 3D imaging (OPT or microCT, depending on embryonic stages) and evaluations of morphological abnormalities in mutant embryos. These procedures can allow direct insight into the window of lethality for each mouse line, but they also provide valuable information on gene function. For example, accurate measurements of organ size and shape can be collected using microCT scan data, or macroscopic observations undertaken by a trained researcher. Importantly, all 3D data sets are available to download from the website for further in-depth analysis by specialist researchers.
In the last few years the IMPC has made major discoveries about parts of the genome that were up to now unexplored, with novel genes discovered relating to areas such as embryonic development, deafness, diabetes, and rare diseases. Recent high profile publications have included research focusing on inferring mammalian gene function, studies on specific human conditions, sexual dimorphism in mouse research, and even using IMPC data to help in wildlife conservation. New methods and analysis tools have also been developed under the umbrella of the IMPC, such as PhenStat, an extensive library of functions that analyse the phenotypical data. Another example is illustrated by a recent article that highlights a new bioimage informatics platform for high-throughput embryo phenotyping. Although this platform was built for the IMPC, the software tools that facilitate the analysis and dissemination of 3D images can be used by other researchers, and is available under an open-source licence. Indeed, sharing resources across the research community is a crucial aspect of the IMPC, and mutant mouse lines can be obtained from the website.
The IMPC is continuing to deliver data and mouse models for the developmental biology field and ultimately will be part of the effort to understanding and treating genetic conditions in humans. More information on the latest research of the IMPC can be found on our blog, and you can search the IMPC database for free at https://www.mousephenotype.org/.
A Technician position in Molecular Biology is available starting November 2018 in the group of Thomas Lecuit at the Developmental Biology Institute of Marseille (IBDM, CNRS UMR7288), Marseille, France. Funding is provided by an ERC grant. The initial appointment will be made for 1 year, with a possible extension to up to 5 years.
We are seeking a highly-motivated candidate with some practical experience in molecular biology with a specialized training as a laboratory technician recognized by a diploma (Bachelor +2 : BTS, DUT,…). Since the working language in the laboratory is English, a rather good understanding in English would be appreciated. The Technician will perform Genetic Engineering in Drosophila using recent molecular biology cloning and CRISPR mediated editing techniques. This position would suit a recent graduate with some practical experience in molecular biology and seeking training in recent Genetic Engineering techniques.
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from a recipient of a Development Travelling Fellowship, funded by our publisher The Company of Biologists.
