General Purpose: Function as a Lab Manager overseeing a Molecular Biology Lab using a zebrafish model system. Research in the Hoffman Lab at Yale focuses on using zebrafish as a model system for the functional analysis of autism risk genes (www.hoffmanlab.net). Work on an independent research project, master generation of zebrafish mutants using CRISPR/Cas9 technique, maintain wild-type and mutant fish lines, characterize new mutant fish lines by genotypic and phenotypic analysis, supervise undergraduates and post-graduate associates in the laboratory and fish facility.
Required Education and Experience: Master’s Degree in a scientific discipline and one year experience or an equivalent combination of education and experience.
Qualifications:
Demonstrated knowledge and ability with zebrafish husbandry and maintenance.
Demonstrated proficiency in molecular biology, including cloning, PCR, in vitro transcription, in situ hybridization, western blotting, and immunohistochemistry.
Demonstrated ability in zebrafish analysis and mRNA injection.
Demonstrated excellent interpersonal skills.
Demonstrated strong work ethic.
Preferred Education, Experience and Skills: Master’s Degree in the Biological Sciences, Neuroscience, Genetics or a related discipline, and one year experience or an equivalent combination of education and experience. 5 or more years of experience. Ability to lead and provide oversight in a lab setting.
Application: For more information and immediate consideration, please apply online at http://bit.ly/2txRpIY. Please be sure to reference this website when applying for this position.
Here at Development we are very sad to be saying goodbye to two of our editors: Ottoline Leyser and Geraldine Seydoux. Both Ottoline and Geraldine have been valued members of the editorial team since 2011, and we are hugely grateful for the time and effort they have put in to handling research papers, helping to shape the journal’s future plans and, in Ottoline’s case, coordinating our 2016 Special Issue on Plant Development. They will both be greatly missed, but will maintain connections with and continue to provide input on the journal as members of our Editorial Advisory Board.
Taking their places, we are delighted to announce two new editors: Yka Helariutta and Susan Strome. Yka is taking on Ottoline’s role as Development’s plant editor. After a PhD in plant genetics at the University of Helsinki, Finland, Yka undertook postdoctoral research at New York University and New York Botanical Garden with Philip Benfey before moving back to Helsinki in 1998 to establish his own lab. From 2003 to 2014 he was also affiliated with the Umeå Plant Science Centre in Sweden, and was elected as an EMBO Member in 2008. In 2014, Yka became Professor of Plant Developmental Biology at the University of Cambridge, UK, and moved to the Sainsbury Laboratory, although he still runs an active group in Finland. Yka’s work focusses on vascular development in plants, from patterning to cell differentiation. He primarily uses Arabidopsis as a research model, but is also interested in how the basic molecular mechanisms operate in species with an extensive vascular domain, such as trees.
Susan Strome began her career as a biochemist working on bacteriophages at the University of Washington, before moving to the University of Colorado for a postdoc with William Wood. There, she started working on the C. elegans germline – an interest that has continued throughout her career. Susan gained an independent position at Indiana University in 1984, staying there until 2007, when she moved to the Department of Molecular, Cell and Developmental Biology at the University of California, Santa Cruz, where she currently holds the position of Distinguished Professor. A Member of the American Academy of Arts and Sciences, Susan’s current research focusses on how cells are instructed to develop as germ or soma, and particularly on how chromatin regulators act to promote and maintain germline fate.
One other significant piece of news is that I [Olivier] will be stepping down as Development’s Editor in Chief in September 2018, after 8 years in the role. During this time, I have tried to adapt the journal to the new challenges faced by developmental biology. Notably, I have tried to engage the journal more actively with the stem cell field and, more recently, with human developmental biology. I have also sought to promote emerging fields such as evo-devo and quantitative biology. We have added new sections to the journal: the ʻTechniques and Resources’ and ʻStem Cells and Regeneration’ sections, which have proved to be very popular. We have also tried to listen to the community and implement a number of changes to the submission and review processes. My time as Editor in Chief has been a fantastic experience and I am pleased I had such a great opportunity to serve our community. However, I think turnover is important to maintain journal dynamism and I have chosen to end my tenure at the 2018 Development/The Company of Biologists meeting ʻFrom Stem Cells to Human Development’, which I am co-organising (for details, please see http://www. biologists.com/meetings/from-stem-cells-to-human-developmentseptember-2018/).
Both the journal, which has undergone significant changes during the past 8 years, and the field as a whole are, right now, in an exciting place. We are of course aware of the challenges that researchers face in today’s funding and publishing environment, but we continue to believe that developmental biology has a bright future. Not only can we now use stem cells, genomic and other techniques to analyse human development as never before, but traditional and new model systems are also being exploited in evermore innovative ways to understand the molecular, cellular and physical bases of development across evolution in unprecedented spatiotemporal detail.
We (The Company of Biologists and Development) are enthusiastic about the prospects for developmental biology, and part of our mission is to support you, the research community. We are seeking a new Editor in Chief with a strong vision for Development at the heart of the field. This search will be led by members of The Company of Biologists’ Board of Directors: Sarah Bray, James Briscoe and Kate Storey. As a key part of the process, we want to consult a broad cross-section of the developmental biology community about the strengths of Development, where we can improve, into which (new or existing) areas the journal should expand and what more we can do to support our field. We therefore encourage you to get in touch with any feedback that you may have, particularly where you think it might be helpful in directing our search for a new Editor in Chief. Please contact Sarah, James and Kate – along with Development’s Executive Editor Katherine Brown and the Company’s Publisher Claire Moulton – via dev.feedback@biologists.com, or get in touch with any of us individually. With over a year to go before Olivier steps down, we are happy to have the time to gather and digest community input to help us make the right choice for the next Editor in Chief and ensure that Development continues to hold an important place in the community long into the future.
Evan Brooks is a rising senior at North Carolina State University. For the past two years, he has worked in the lab of Nanette Nascone-Yoder studying the developmental mechanisms of cardiac left-right asymmetry using Xenopus as his model system. Evan was awarded a Choose Development! Fellowship from the Society of Developmental Biology to conduct two summers of mentored research in developmental biology. He was selected to spend one week with us at the Embryology Course this year. I had the pleasure of working next to Evan for the zebrafish and Xenopus module. I thought I would take this opportunity to check in with Evan and ask him a few questions!
Were you told any stories of Woods Hole or did you know anything about the course before you arrived?
Before getting my invitation to join the course, I had only heard of Woods Hole and the Embryology course in passing in the lab. I had never heard of anyone’s stories or experiences with Woods Hole or the Embryology course.
When I got my invitation in May from SDB to join the course for a week, Nanette talked to me about her experience in the course when she was a graduate student. She not only told me how much she enjoyed the course, but also how it allowed her to think more about the evolutionary similarity between model organisms. She also told me how much fun she had in Woods Hole when she wasn’t in the lab, whether it was looking at bioluminescence in Eel Pond or staying up late with her classmates at Captain Kidd. With the knowledge about her experience and its impact on her, I was super excited to join the course for a week.
Did your time in the Embryology course match what you had anticipated?
I will say that I was expecting to be immersed in science while I was there. I looked at the schedule before arriving and was excited to see so many prominent developmental biologists, like Richard Harland and Ray Keller, participating in the course in some way. I was totally expecting to be thinking about possible new experiments all the time.
However, I was not expecting to dive headfirst into the course on my first day in the lab and stay up super late conducting experiments. I was up past 1am every night and woke up every morning around 7am to get back into the lab and check my experiments. On the day I left Woods Hole to return home, I got up at 5am to finish an in situ hybridization before catching a 7:20am bus ride to Boston Logan airport. I was committed to performing so many experiments and seeing them all pan out during my one week and I do not regret it.
What was the most memorable part of your week?
I’d probably say the camaraderie within the program and how integrated I felt in the course is what I will remember most from my week. Everyone was so welcoming and I was frequently included in microscope demos down in the basement of Loeb Laboratory, on coffee runs before lectures, and during meals in Swope Hall. Also, people would encourage me to stay up into the wee hours of the morning with them to try a new experiment. Everyone that I talked to was interested in my research at NC State and what I was doing in the lab during the course. As the week progressed, I was told by more and more people to talk to the course directors, Rich Schneider and Dave Sherwood, to see if there was any way to extend my stay. I really wish that I could’ve extended my stay as I was just getting comfortable with working in the lab and getting to know everyone when it was time for me to leave. I’m still connected to everyone in the course through WhatsApp and I know that I’ve made at least twenty-four new friends.
What did your learn during the week?
Scientifically, I learned so many new things in the course. I learned so much about the current research going on in Xenopus and zebrafish labs around the world, like work on liver regeneration and development in Elke Ober’s lab and the roles of planar cell polarity in development in John Wallingford’s lab. The one lecture that I was most intrigued by was Andrew Gillis’ lecture about gill arch development in skates and its comparative development to limb buds. The ideas, techniques, and approaches presented in the lectures that week showed me how broad the field of developmental biology really is.
From the course, I’ve also learned to take more risks not only in the lab, but in life in general. Taking a week off from my research at NC State to come to Woods Hole was somewhat of a risk as I had to get more data and prepare for my presentation at the SDB Annual Meeting a few weeks after my return. The risk ended up being well worth it in the end as I learned a lot more about myself and developmental biology in that one week.
How did this course change how you approach your current research at NCSU? Did it impact your future plans?
Evan working in lab in Loeb
In terms of working with my hands in the lab, I can say that I have a greater confidence with microinjecting Xenopus embryos. Before the course, all of my embryos would die after microinjections. I can’t really say what caused my embryos to die, but I moved away from microinjection experiments for a bit in the lab. During my week in the course, John Young, one of the Xenopus TAs, showed me how he calibrated his needles and microinjected his embryos. After employing his method, I had quite a few successful morpholino microinjection experiments that I used as the basis for my show-n-tell at the end of the week. I even had the chance to microinject zebrafish embryos and those survived injections too! With my new confidence in microinjecting, I’m looking to incorporate that technique back into my lab repertoire.
Additionally, one thing that I realized that I did not take advantage of during my time in the Embryology course was the cutting-edge microscopy. Before the course, I had only used light microscopes in my lab. During the show-n-tell at the end of my stay, there were so many amazing time-lapse movies and images of cells, tissues, and embryos that I was in awe! I can say that this alone has changed the next steps of my project as I’ve talked to Nanette about designing experiments that will allow me to use other types of microscopes available. I know that there is an expansive microscopy core at NC State, so I may ask for a guided tour in there soon. In terms of my next steps in the Nascone-Yoder lab, I’m planning on performing some antibody stains and confocal microscopy for a few proteins to understand their left-right distributions during developing heart tube stages. I hope to get some pretty images that may be used in a future publication from the Nascone-Yoder lab!
This opportunity is given yearly by SDB, is there any advice you would give to next year’s Choose Development! Fellow who will come to the Embryology Course?
Do not be afraid to jump right in by planning experiments and making friends! The course directors, the instructors and TAs, and fellow course participants are all there to help and get you acclimated to the course and Woods Hole. I remember staying up until 2am with Ray Keller one night because I wanted to learn how to create a two-headed Xenopus embryo. Even though I failed miserably (probably due to exhaustion), he was super patient with me and made sure that I gave it my best effort.
Do you have anything else you would like to include?
I thought I was super excited about embryology and developmental biology before the course. My level of excitement for the discipline has grown so much to where I feel that putting my excitement into words will not accurately describe it. I’m glad that I had the opportunity to experience one week of the course. I am totally looking forward getting into a Ph.D. program and reaching candidacy so that I can be eligible to return to Woods Hole to experience the full six-week course!
Lastly, I honestly cannot thank SDB, Dave Sherwood, and Rich Schneider enough for allowing me to experience one week of the Embryology course this year and for past course directors, Richard Behringer and Alejandro Sánchez Alvarado, for spearheading the idea of inviting Choose Development! Fellows to the course for a week. Past Fellows that I’ve met that have participated in the course absolutely love the experience and I am glad we can partake in this course. The course has helped me affirm my desire to pursue a career in developmental biology!
To check out all the cool things we did during the course follow us on Twitter #embryo2017 or instagram #embryology2017
The Drosophila tracheal system is a powerful model for understanding the genetic and cell biological control of tubulogenesis. In their new PLoS Genetics paper, Ivette Olivares-Castiñeira and her PI Marta Llimargas of the Molecular Biology Institute of Barcelona connect EGFR signalling to intracellular cell trafficking during tracheal morphogenesis. We caught up with Marta and Ivette to hear the story behind the paper.
Marta and Ivette in the lab
Marta, can you give us your scientific biography and the main questions your lab is trying to answer?
MA I studied Biology at the University of Barcelona where I took a course on developmental biology and embryology, given by Dr. Jaume Baguña that made me enthusiastic about development. Then I was very lucky because Dr. Jordi Casanova, a well-known Drosophila developmental biologist, just got a position in Barcelona and he accepted me as a PhD student in his lab. By chance we started to work on tracheal development, and I became fascinated by the development of such a beautiful and interesting tissue. The analysis of tracheal development was just an emerging field by then and this allowed me to follow and contribute to the field during all these years.
In 1997 I moved for a postdoctoral stay in Dr. Peter Lawrence’s lab, at the LMB-MRC in Cambridge. There I had an incredible scientific and personal experience, and I was encouraged by Peter to develop my own projects. Because I was really thrilled by tracheal development and Peter allowed me the freedom, I continued working on the subject while there. In 2002 I moved back to Barcelona where I started my lab and later, in 2007, I got a permanent position at the Molecular Biology Institute of Barcelona. My lab has always been, and still is, interested in tracheal formation as a model to understand the morphogenesis of branched tubular organs. We are interested in the genetic mechanisms that drive tracheal formation and in the molecular mechanisms used by these genetic networks to instruct the cellular changes that underlie organ formation.
With the recent announcement of a new EMBL site and a number of universities and research institutes, Barcelona seems to be an exciting place for life sciences at the moment?
MA Yes, Barcelona has become a real hub for science in the past few years and at the moment it is an excellent place for researchers. Barcelona is attracting many senior and young researchers, from many different areas of research. This provides an exciting and encouraging atmosphere for interdisciplinary research. Moreover, many pharmaceutical and biotech companies are set in Barcelona, which also encourages translational research and transfer of knowledge. Obviously the lifestyle, the climate and being a medium sized city also positively contribute to the success of Barcelona as a scientific hotspot.
Barcelona has a number of very good institutions and the production of high quality science in the region is high. Some institutions are very good and very visible, but there are other good institutions and talented people that are less known and overall people struggle with lack of consistent funding to properly develop their research. The general funding situation for science is still below what would be required to fully take advantage of the people and resources.
And Ivette, how did you come to join the Llimargas lab?
IO-C I joined Marta’s lab in 2013, when I had just finished my master’s degree and I realised that I would like to do a PhD. I was looking for an opportunity and Marta gave me the chance to join her lab. After the first year, I became interested in analysing how EGFR was controlling the development of the tracheal system. Being in a developmental biology lab gave me the opportunity to investigate the mechanisms of organ and tissue formation. I find Drosophila tracheal development particularly interesting and ideal to analyse the genetic control of morphogenesis and the cellular mechanisms at play. Doing my PhD with Marta is a great experience for my future career and allows me to get insights into the morphogenesis of epithelial tissues.
Lateral views of stage 16 embryos expressing EGFR constructs, from Fig. 1, Olivares-Castiñeira & Llimargas 2017.
What was known about the role of EGFR signalling in tracheal development before your paper?
MA & IO-C We already knew a few things. Our lab had published work on the role of EGFR during embryonic tracheal development, in which we showed that it is required for invagination of tracheal cells and to maintain epithelial integrity. In addition, other labs, like Dr. Casanova’s and Dr. Hayashi’s, had also investigated the role of EGFR in tracheal invagination and identified an EGFR-dependent regulation of Myosin-II.
However, a role for EGFR in tube growth was not previously reported. It was actually, back in 2006, during our analysis of EGFR signalling on epithelial integrity that we noticed the effect of EGFR on tube length. However, this observation coincided with the end of the PhD of the student working on this project and it was not followed at the time. When Ivette started her PhD we thought that investigating EGFR requirement on tracheal elongation was a good project to explore, also because there was much more information about tracheal tube growth at the moment and we felt it was the right time to approach the issue. In addition we were also interested in analysing in more detail the possible different molecular mechanisms of EGFR underlying the different tracheal requirements such as invagination, integrity and tube length mainly at the cell biology level.
After analysing EGFR activity in our and other labs, we now know that, as it happens in many other organs and tissues in Drosophila and in other organisms, EGFR is used reiteratively during tracheal formation, but the exact molecular mechanisms behind each of these requirements still need to be fully clarified.
Apical cell shape in tracheal metameres, from Fig. 1, Olivares-Castiñeira & Llimargas 2017.
Can you give us key results of the paper in a paragraph?
MA & IO-C We have found that EGFR is required to prevent an excessive growth of the tracheal tubes. EGFR is required for the proper accumulation and localisation of two previously known regulators of tube length, the apical determinant Crumbs and the apical extracellular matrix (aECM) regulator Serpentine. This positions EGFR as a hub coordinating cell intrinsic properties (Crumbs-mediated apical expansion) and cell-extrinsic mechanisms (Serpentine-mediated modification of the aECM) regulating tube length. Interestingly we found that these two proteins accumulate in common sorting endosomes, and that EGFR is required for the proper organisation of these common endosomes, likely regulating the correct delivery of both cargoes to their final destination. We observed that the two cargoes are partitioned into different discrete domains within this common sorting endosome, consistent with the hypothesis that they use different retrieval pathways to recycle. We also observed that, during tracheal development, Crumbs undergoes a complex pattern of recycling, which involves internalisation and different sorting pathways. Our results illustrate a role for EGFR in endocytic trafficking, a molecular mechanism that could potentially underlie different developmental and pathogenic EGFR activities.
What do you think the EGFR receptor is doing in the endosomes?
MA / IO-C That is what we would like to unravel! Many reports in the literature describe the internalisation and trafficking of EGFR receptor, the many factors involved in the process and the consequences for EGFR activity. So obviously EGFR is a cargo of endocytic trafficking, and this trafficking regulates its signal termination and the recycling of the receptor to the membrane. Much less is known about a possible role of the receptor itself in controlling its own or other protein’s trafficking. Our results point to a role of EGFR in organising the endosome, because when EGFR is downregulated endosomes are bigger, cargoes are mis-sorted and WASH accumulation, which is recruited by the Retromer complex, is also affected. Whether this is a direct or an indirect effect of EGFR downregulation is still unclear. However, the fact that we find EGFR itself in Crumbs/Serp sorting endosomes and the fact that endosomes are affected leads us to speculate that EGFR regulates targets in the endosome. In fact, EGFR cytoplasmic targets and endosomal targets have been identified, suggesting that intracellular organelles can act as EGFR signalling platforms. To get further insights into the molecular mechanism of EGFR in tube elongation we are working with the hypothesis that EGFR (or EGFR downstream effectors) signals from the endosome to organise this organelle.
Lateral views of embryos at the indicated stages carrying AbdGal4-UASGFP stained with Serp antibody, from Fig. 2, Olivares-Castiñeira & Llimargas 2017.
Do you have any clues as to how the apical ECM instructs the underlying tracheal epithelium during tube elongation?
MA / IO-C This is an outstanding question in the field. It is clear that the aECM and the underlying tracheal cells cross-talk and that this cross-talk is required for the correct tube growth in length and diameter. But the exact molecular mechanism is still unclear. The aECM can potentially instruct tracheal cells by exerting a mechanical force that is received by mechanosensors in the tracheal cells. It could also trap or expose chemical signal/s that could be received by tracheal cells. Or it could both act as a mechanical and a signalling platform. Bo Dong and Shigeo Hayashi have put forward a model where the apical membrane expansion force in tracheal cells that drives tube elongation is balanced with the resistance of an elastic aECM that restricts overelongation. The aECM mechanical tension and the apical expansion force are coupled through proteins attaching or coordinating the apical membrane to the aECM, such as Zona Pellucida proteins (as proposed by Hayashi lab) or Src42A (as proposed by Sofia Araújo and Jordi Casanova). Hopefully the dedicated work of several labs in the subject will soon shed light into this issue and we will better understand not only the growth of tubular structures but also how the cross-talk of extracellular matrices and organ/tissue/cells instructs morphogenesis.
Model for EGFR’s role in intracellular trafficking, from Fig. 7, Olivares-Castiñeira & Llimargas 2017.
When doing the research, did you have any particular result or eureka moment that has stuck with you?
IO-C From the beginning of the work, our main goal was to identify the target/s of EGFR that control tracheal tube length. We found that Crumbs and Serpentine were affected in the tracheal tubes upon modulation of the EGFR activity. My eureka moment was when one afternoon, we realized that these two protein seemed to be more related than expected and we observed that they were loaded in common vesicles (we did not know that these were endosomes at the moment). It was a moment of euphoria as this was shedding light into a complicated puzzle of many pieces, although this also raised many other questions.
And what about the flipside: any moments of frustration or despair?
IO-C Science always has moments of frustration. It was particularly frustrating to repeat many times some of the stainings before we got good samples. In general, I find that technical details can often be really time-consuming and despairing. It was also frustrating to use several RNAi or dominant negative lines that did not produce phenotype, although I knew that the RNAi technique does not always work well for embryonic development. And finally, I found it very frustrating when we had different results but we were not able to understand what was happening. It is not always easy to put the pieces of the puzzle in the right place to get the final picture.
What are your career plans following this work?
IO-C First, I need to finish my PhD, which I plan to do this year. In the lab I am trying to get more details on the molecular mechanisms of EGFR and on the role of Crumbs in tube size. I am particularly interested in the connection of these proteins with intracellular trafficking pathways. After I finish my PhD, I will probably continue in the research world, as I find it fascinating.
Crb expression in different conditions, from Fig. S4, Olivares-Castiñeira & Llimargas 2017.
And what next for the Llimargas lab?
MA Well, we are continuing different aspects derived from this work. For instance, we are now very interested in understanding Crumbs localisation in different apical subcellular domains (in trachea and other tissues) and whether this correlates with different Crumbs activities. Obviously this aspect is not only relevant for tracheal development, but may help to better understand the complexities of Crumbs. We are also interested in understanding how Crumbs, Serpentine, Vermiform, Src, EGFR and Dumpy, all of them known regulators of tube length, interact and regulate each other. Furthermore, we are really curious to understand how the retromer complex selects the different cargoes, Crumbs and Serpentine in this case, and sorts them into different recycling pathways. For this particular question we have identified a putative regulator of retromer trafficking, a nexin, that could provide cargo and itinerary specificity for the specific sorting of Serpentine and Crumbs. We are now investigating this aspect, generating mutants for this nexin and characterising its role. But obviously, the main question we are trying to answer is how EGFR regulates endocytic trafficking.
Besides this project, in the lab we are also interested in other aspects of tracheal formation, related with chitin deposition and with adhesion and polarity maintenance and remodelling.
Finally, what do you like to do when you are not in the lab?
MA I spend as much time as I can with my family. I try to get some time to read, to go to the theatre, to dance and to go out with friends. I love cooking for friends and family and I love eating, particularly new and surprising foods. I also exercise, particularly gym and running, in the nature when possible.
IO-C I try to switch off my mind from the lab issues and I like to go for run and do sport, which is good to clear my mind. Also, I like to read or to be with my friends. If I had more time I would like to travel more.
The Paridaen lab at ERIBA is currently looking for a highly motivated PhD student and post-doctoral researcher.
ERIBA (The European Research Institute for the Biology of Ageing) is a newly established institute at the University Medical Centre Groningen (UMCG). The mission of ERIBA is to better understand the molecular mechanisms that result in age-related decline and age-related diseases. We aim to develop novel strategies to prevent or combat age-related diseases and to provide evidence-based recommendations for healthy ageing. The research teams are working with different model systems and technology platforms and share their knowledge and expertise to accelerate discoveries. The ERIBA is housed in a new building and is part of the central UMCG campus in the vibrant University City Groningen in the north of the Netherlands. Groningen is a small (200,000 inhabitants), but vibrant city hosting over 25,000 students.
Stem cells act in developing and adult organisms to produce the proper number of specialized cells in the body. It is important that stem cells select an appropriate division mode to ensure proper development, maintenance and repair of tissues. In the Paridaen lab, we aim to understand the mechanisms underlying stem cell decisions in developing and ageing tissues. We employ single cell analyses techniques to study stem cell behaviour in the developing brain, using zebrafish and mouse as our model systems. In particular, we are interested in the role of fate-determining factors such as signalling pathways and subcellular structures in symmetric and asymmetric stem cell divisions.
For more information and application (before 3 September 2017), please see: http://ow.ly/NDeh30dKAxq
A new set of DMDD embryo and placenta data has been released, taking our total dataset to 9.5 million images of around 1300 embryos.
DMDD is a primary screen of embryonic lethal knockout mice, and all data can be freely accessed at dmdd.org.uk. Detailed phenotypes are available for embryos from 73 different knockout lines, and we have phenotyped the placentas from 124 lines. We have also added data on the sex of each embryo.
Visitors to our website can now compare HREM embryo images with the closest-matching, annotated histological section from the Kaufman Atlas of Mouse Development. This follows a major project by the eMouseAtlas team at the University of Edinburgh to digitise the Kaufman Atlas at high resolution. The annotated Kaufman sections can be viewed alongside DMDD embryo images to help users who are unfamiliar with the detailed morphological features of a mouse embryo as it develops.
Severe brain phenotypes
Phenotyping of Hmgxb3 knockout embryos revealed severe brain defects, with half of the embryos displaying exencephaly. Embryos from this line also had a range of phenotypes including edema, abnormalities of the optic cup, and defects of the venous system including an abnormal ductus venosus valve and blood in the lymph vessels.
An Hmgxb3 homozygous knockout embryo displays exencephaly.
Gene knockout can lead to a huge number of phenotypes
Fifty-five different phenotypes were identified in Cnot1 knockout embryos. These included an absent hypoglossal nerve (which is needed for tongue movement and suckling), abnormalities of the inner ear, testis and thyroid gland, abnormal cell masses that have been classified as embryo tumours, and various heart defects including overriding aortic valve and ventricular septal defects.
The images below show an embryo tumour, and the absence of the mandibular nerve (click the image to view a larger version).
A Cnot1 knockout embryo displays an embryonic tumour (left) and absent mandibular nerve (right) amongst many other phenotypes.
Potential models of human disease
A number of genes studied by DMDD have already been associated with human diseases. For example, Prmt7 mutations have been associated with Short Stature Brachydactyly Obesity Global Developmental Delay Syndrome, an autosomal recessive disease characterised by developmental delay, learning disabilities, mild mental retardation, delayed speech, and skeletal abnormalities. Strikingly, in the Prmt7 knockout embryos studied, the most common phenotypes included neuroma of the motoric part of the trigeminal nerve (a tumour within the skull, affecting the nerve controlling the jaw movements needed for speaking and chewing) and abnormalities of the hypoglossal nerve (which controls movement of the tongue) and the ribs.
Many of the genes studied by DMDD do not currently appear to be associated with any disease, for example Hmgxb3 or Cbx6. There is potential that careful analysis of the phenotypes from lines such as these could contribute to the identification of new disease models, and our data is freely available in order to encourage this.
A detailed description of normal mouse embryo development
The Atlas of Mouse Development by Professor Matthew Kaufman describes normal mouse embryo anatomy using a series of hundreds of annotated histological sections. Even today, twenty three years after its publication, it is still considered to be the gold standard for describing mouse embryo development. As part of a project to update the book in 2012, the original sections were digitised by the Edinburgh Mouse Atlas Group and made freely available on their eHistology resource.
The images have now been integrated into the DMDD database, and users can directly compare any HREM embryo image with the closest-matching annotated Kaufman section.
Each HREM embryo image can now be viewed alongside the closest-matching section from the Kaufman Atlas of Mouse Development.
This new feature is intended to help users who are not fully confident of the details of mouse developmental anatomy. It means that mutant mouse data can now be explored alongside a fully-annotated wild-type reference point.
Development depends on cells coming together in the right place to form functioning tissues. Our latest paper in this series was published recently in Current Biology and asks what happens to the structure and function of the nervous system when motor neuron migration – and hence final position – is disrupted. We caught up with postdoc Kimberly McArthur and her PI Joseph Fetcho, Professor at the Department of Neurobiology and Behaviour at Cornell University.
Joe, can you give us your scientific biography and the main questions your lab is trying to answer?
JF I was always curious about different types of animals, but I was particularly interested in snakes and how they moved. As a graduate student, I studied the locations of spinal motor pools across multiple species, and found that anamniotes (like snakes and fish) – unlike amniotes (like mammals) – lack a clear topographic relationship between motor neuron location in the spinal cord and muscle target location in the periphery. As a postdoc, I started working more with physiology and circuit structure in goldfish, studying the hindbrain Mauthner cell circuit that generates fast escapes, because I thought I could use it to figure out a circuit for a vertebrate behavior from sensory input to motor output before I died. Once I started my own lab, I switched to studying neural circuits in larval zebrafish – which had been an important developmental model for many years, but which also had incredible potential as a vertebrate circuit and behavioral model because of its relative simplicity, optical transparency, and genetic accessibility. Since then, my lab has continued to use zebrafish to study the principles by which neural circuits generate behavior, and how these principles inform and are informed by our understanding of circuit development and evolution.
Imaging live synapses in the fish (image sourced from the Fetcho lab website)
And Kim, how did you come to join the Fetcho lab?
KM In my graduate work, I studied state-dependent changes in hindbrain sensorimotor circuits in pigeons – seeing how vestibular reflexes were modified by the transition into flight, and looking for the neural correlates. I was puzzled by the (apparent) lack of topography in the vestibular nuclei, how neighboring neurons can have completely different response properties – and I was a bit frustrated that I didn’t have the tools to easily nail down the identity of the neurons with a specific set of physiological properties. I came across a pair of papers from Joe’s lab (Kinkhabwala et al. 2011 and Koyama et al. 2011, both in PNAS) demonstrating the link between early neuronal topography and circuit recruitment for interneurons in the hindbrain, using larval zebrafish. I was impressed both by the scientific finding and by the way in which the model facilitates a direct line of investigation from genetic profile to cell morphology to physiology to behavior – all in an intact vertebrate.
How did you come to be interested in the relationship between cell positioning and function in the nervous system? Had this been experimentally tested before your work?
JF My predoctoral research dealt with the role of neuronal positioning in spinal motor circuits, and some of our current work demonstrated the correlation between hindbrain interneuron positioning and circuit recruitment. Previous studies have certainly addressed the importance of neuronal positioning; indeed, there is a great deal of evidence that positioning can be critical for proper circuit development. And it’s sensible for the developing nervous system to lean heavily on positioning as a strategy for targeting synaptic connections, as most neurons arise in specific locations and end up in stereotyped locations within a given species. We were interested in the importance of migration and hoped to get a better understanding of exactly what changes in a case where neuronal position is altered by a genetic block of migration in a population of cells that also changes position in evolution. We figured it would help us understand why migration and position seem to matter so much. We did not expect to find so much unaffected by a large position change.
KM Other labs were using zebrafish facial motor neurons to study cellular mechanisms of neuronal migration, because these neurons execute an early caudal migration through the developing hindbrain. Those researchers had identified several mutations that spared the gross organization of the hindbrain but blocked the caudal migration of the facial motor neurons. We saw an opportunity to investigate the impact of this dramatic mis-positioning on the facial motor neurons themselves and the developing cranial motor circuits. We expected that there would be clear functional deficits in mis-positioned neurons, and we thought we’d be able to work out the details of how those deficits occur.
Hindbrains of Tg(Islet1:GFP) larvae at 5 dpf, from Figure 1, McArthur and Fetcho, 2017
Can you give us the key results of the paper in a paragraph?
KM Contrary to our expectations, we found several lines of evidence indicating that facial motor neurons were robust to an abnormal shift in their segmental position. Even though their cell bodies and dendritic arbors were in entirely the wrong part of hindbrain, they still established intra-population topography (according to muscle target and relative age) and respiratory activity patterns (observed using intracellular electrophysiology and calcium imaging) that were very similar to wild type neurons. Indeed, wild type and mutant larvae exhibited similar respiratory behavior, which involves significant contributions from facial-innervated structures. Taken together, these results indicate that hindbrain motor networks can be surprisingly robust to a change in neuronal positioning – though there may be underlying differences in the motor circuits that we have yet to identify (which we’re pursuing now).
Backfills from cranial muscles reveal the location of facial motor pools, from Figure 1, McArthur and Fetcho, 2017
Is the resilience to change in position you’ve identified in motor neurons likely to be specific to these cells or a general feature of the nervous system?
KM I don’t believe that resilience to abnormal positioning is specific to facial motor neurons, but we also already know that it isn’t a general feature of every neuronal population – given previous work demonstrating that scrambling cell position can interfere with synaptic targeting in spinal cord, for example. Instead, I think that the developing nervous system deploys different synaptic targeting strategies in different circuits, and that some of these strategies are less sensitive to position than others.
Further, in the case of the facial motor neurons, I’m not convinced that positioning isn’t important. Facial motor neurons are born in roughly the same segmental location across vertebrates, but they end up in different places – suggesting that one location might be more adaptive than another in a particular species, depending on the dominant inputs to the facial motor neurons. It could be something to do with the metabolic cost of maintaining neural processes (i.e. wiring minimization), but I suspect that shifting segmental location might also affect the probability of synaptic input from a specific source (rather than specifying it in a deterministic fashion).
Finally, it’s interesting to note that facial motor neurons did establish the same cross-sectional topography in wild type and mutant larvae. Perhaps relative dorsoventral or mediolateral positioning is more critical than segmental (rostrocaudal) positioning for synaptic targeting in this specific population. This could be because many tracts in the hindbrain run longitudinally, so moving along that axis might have less impact on the inputs than moving in another direction.
Tracking facial motor neuron development and organisation with photoconversion, from Figure 3, McArthur and Fetcho, 2017
And what might be the evolutionary implications of your findings?
KM I think of the developing nervous system as (implicitly) giving each neuron a set of instructions for locating potentially useful synaptic partners. Those instructions can be written using different strategies. For any given synapse, multiple strategies might work equally well – but if there is some perturbation (due to, for example, a genetic mutation or an abrupt environmental change), different strategies will fail in different ways. This will be reflected in how specific circuits respond to developmental perturbations (i.e. which perturbations actually cause dysfunction and disease), and it will shape the landscape of inter-individual variation that is available as a substrate for natural selection. Facial motor neurons, for example, have shifted their segmental location across phylogeny. Perhaps that’s possible because they are targeted for synaptic contact in such a way that core behaviors (like respiration) are robust to a shift in segmental position. This might give the facial nucleus a chance to sample (via mutation) novel, potentially adaptive synaptic connectivity without losing access to its original network.
When doing the research, did you have any particular result or eureka moment that has stuck with you?
KM I was initially a bit disappointed when I couldn’t find a clear functional difference between wild type and mutant facial motor neurons. I’d been framing my research as a case study in which to explore how abnormal positioning does impact function, in a particularly helpful animal model for working through the circuit-level implications. However, as I did more thinking and more reading, I realized how interesting and important this result might be. As useful as it can be to look for factors that (when disrupted) break the system, I think it’s also important to understand how much can go wrong without breaking the system. Indeed, I think this may be part of the reason that many human neurological disorders are so genetically complex: because breaking a single factor (or even a set of factors) involved in circuit formation and function doesn’t necessarily cause observable disease in every individual.
Whole-cell patch-clamp recordings from mutant neurons, from Figure 5, McArthur and Fetcho, 2017
And what about the flipside: any moments of frustration or despair?
KM The biggest technical challenge for me was learning how to do whole-cell recordings in the larval zebrafish brain. When I was a graduate student, I learned how to do extracellular in vivo electrophysiology – which isn’t trivial, but you can start getting data relatively early if you get lucky with your electrode placement. When I started my postdoc, I spent several months learning how to handle the tools, do the dissection, work the scope, and finally record from cells. And I spent almost two months straight focused entirely on recording without actually getting any data. So, it was challenging to spend the first part of my postdoc mostly failing all day – but I also kinda like those moments, because it’s rewarding to overcome a challenge. And it worked out well in the end.
What are your career plans following this work?
KM I’m on the academic job market this year, applying for tenure-track faculty positions that combine research and teaching. I’d like to join a department where I can continue studying the development and function of neural circuits using larval zebrafish.
Tracking motor neuron innervation, from Figure S1, McArthur and Fetcho, 2017
And what’s next for the Fetcho lab?
JF Recently, my lab has started to move into collaborations to advance technologies for neural circuit research, especially for looking with high resolution into brain and spinal cord of intact living adult zebrafish. The goal is to be able to study circuits and behavior in a vertebrate (even in one individual!) from embryo to adult.
Finally, what do you two like to do in NY state when you are not in the lab?
JF Not much. Lab has been my passion and my job. I mostly work, read, and think. I sometimes hang out in or near the woods behind our house looking for snakes and other animals including bears, pileated woodpeckers, turkeys, foxes and bobcats. Animals are my thing. People, not so much.
KM I think I’m your quintessential nerd! I get excited about going to the public library and checking out a stack of books. (No, seriously.) I try to balance my literary diet: nonfiction books about current events, plus lots of horror novels.
We came across this video from EuroStemCelldescribing a project that aimed to increase public awareness of stem cell biology, as well as encourage scientists to get involved in public engagement and interact with other professionals along the way. In their introduction to the video, EuroStemCell give us the background:
“Creative public engagement initiatives are often strengthened by collaborations but how do they come about? What are the benefits and challenges? This short animated film reveals the story, from beginning to end, of a collaborative public engagement initiative involving stem cell scientists, public engagement professionals, artists and social anthropologists. We hope that by sharing what we have learned, it will encourage others to get creative and collaborative!”
Here are the highlights from the current issue of Development:
Computing branching pattern complexity
Bile – a fluid that aids digestion – is transported from the liver to the intestine through the bile duct. Bile reaches the bile duct itself via a complex, highly branched structure called the intrahepatic biliary network. This network spreads throughout the liver but how it is patterned is unclear. On p. 2595, Takuya Sakaguchi and colleagues report a novel computational approach to analyse the 3D structure of this network in developing zebrafish. They use a computational algorithm that renders confocal scans of labelled livers into compact representations of the intrahepatic biliary network, which recapitulate endogenous branching patterns and simplify the branched networks into segments amenable to further analysis. Using this computational approach, the authors identify a small molecule inhibitor of Cdk5 that reduces the density of the biliary network, leaving liver size and biliary epithelial cell numbers unchanged. They also experimentally manipulate the downstream Cdk5-Pak1-LimK-Cofilin cascade to increase branching density, and demonstrate a role for this cascade in regulating actin dynamics in biliary epithelial cells. These findings demonstrate the utility of this computational approach to studying branched tissues and highlight the Cdk5-Pak1-LimK-Cofilin cascade as a potential therapeutic target for liver disorders.
No lung development without Sin(3a)
Sin3a is a co-repressor that modulates the transcription of numerous genes by complexing with chromatin remodelling enzymes that modify histones. Here, Barry Stripp and co-workers reveal a key role for Sin3a during lung development in mice (p. 2618). They report that the foregut endoderm-specific deletion of Sin3a leads to failed lung development and the death of neonatal pups due to respiratory failure. Although lung buds form in early mutant embryos, subsequent branching morphogenesis and development fails, with loss of both the lung endoderm and lung epithelium. Loss of Sin3a also disrupts lung mesoderm differentiation, possibly due to aberrant epithelial-mesenchymal interactions. Interestingly, histone H3 acetylation levels show no significant change in Sin3a-deficient epithelial cells, indicating that loss of histone deacetylation activity is unlikely to contribute to the lung phenotype. Instead, lung epithelial progenitor cells in mutant embryos enter a senescence-like state and arrest in G1. This cell cycle arrest is partially mediated by upregulation of the cell cycle inhibitors Cdkn1a and Cdkn2c. Together, these findings reveal that Sin3a plays a crucial role in regulating early lung endoderm progenitor cell fate, via the transcriptional repression of cell cycle inhibitors, to prevent the induction of a senescence-like state. Whether Sin3a plays a similar role in the postnatal lung awaits further investigation.
Set(d1b)ing oocyte gene expression
Mouse primordial germ cells (PGCs) – the precursors of oocytes and sperm – undergo extensive DNA demethylation as they migrate to the genital ridge. In primary oocytes, DNA undergoes re-methylation, and lysine residues on the tails of histone H3 become methylated. This important epigenetic mark is created in mammals by H3K4 methyltransferases, including Setd1a and Setd1b. Setd1a plays no role in oogenesis, but now Andrea Kranz and colleagues report that the loss of Setd1b in mouse oocytes causes female sterility (p. 2606). Although metaphase II-stage oocytes develop in female Setd1bconditional mutants, both their zona pellucida and meiotic spindle are abnormal. Upon fertilisation, extra sperm enters the perivitelline space of the mutant oocytes and the zygotes become stuck at the pronuclear stage. RNA profiling reveals that Setd1b is required for the expression of key oocyte transcription factors and that its inactivation causes twice as many mRNAs to be upregulated as downregulated. Thus, Setd1b likely promotes the expression of transcriptional repressors, possibly Zfp-KRAB factors, which maintain the oocyte-specific expression programme in late development by reducing earlier, unwanted gene expression. These findings reveal a novel role for Setd1b in regulating the oocyte-to-embryo transition, possibly by regulating the late-oocyte gene expression programme.
PLUS…
The status of the human embryo in various religions
Research into human development involves the use of human embryos and their derivative cells and tissues. How religions view the human embryo depends on beliefs about ensoulment and the inception of personhood, and science can neither prove nor refute the teaching of those religions that consider the zygote to be a human person with an immortal soul. In his Spotlight article, William Neaves discusses some of the dominant themes that have emerged with regard to how different religions view the human embryo, with a focus on the Christian faith as well as Buddhist, Hindu, Jewish and Islamic perspectives.
Interspecies chimeras for human stem cell research
Interspecies chimeric assays are a valuable tool for investigating the potential of human stem and progenitor cells, as well as their differentiated progeny. In their Spotlight article, Hideki Masaki andHiromitsu Nakauchi discuss the different factors that affect interspecies chimera generation, such as evolutionary distance, developmental timing, and apoptosis of the transplanted cells, and suggests some possible strategies to address them.
A framework for understanding the roles of miRNAs in animal development
MicroRNAs (miRNAs) contribute to the progressive changes in gene expression that occur during development. In their Review, Chiara Alberti andLuisa Cochella present a view of miRNAs as a hierarchical and canalized series of gene regulatory networks. In this scheme, only a fraction of embryonic miRNAs act at the top of this hierarchy, with their loss resulting in broad developmental defects, whereas most other miRNAs are expressed with high cellular specificity and play roles at the periphery of development, affecting the features of specialized cells.
Two fully funded PhD Positions in cell, developmental and systems biology
at the Universitätsklinikum Freiburg, Germany
The Walentek lab studies the molecular mechanisms of mucociliary development, regeneration and disease. Mucociliary epithelia line the embryonic epidermis as well as the respiratory tract of many animal species, and provide an important first line of defense against pathogens for the organism. We are particularly interested to elucidate the interactions between cell signaling, transcriptional and post-transcriptional regulation of gene regulatory networks, and the morphogenetic processes at the cellular and tissue-wide levels, which facilitate complex tissue formation and function. Our work aims to provide crucial insights into the logic of self-organization in biological systems as well as into the molecular mechanisms underlying chronic lung diseases.
Two PhD positions will be available starting October 2017 (or later). The group is supported through the Emmy-Noether-Program by the DFG, which provides fully funded PhD positions (65%). We offer a great environment to perform cutting-edge science at the interface between basic biology and translational medical studies. The Walentek lab is affiliated with the excellent Freiburg Medical Center (Universitätsklinikum Freiburg) and situated in the multidisciplinary research building of the ZBSA (Center for Biological Systems Analysis). This setting provides access to state-of-the-art core facilities and collaborations, including advanced light and electron microscopy, proteomics, computational biology, mathematical modeling and genetics/ genomics. PhD students will have the option to participate in the Spemann Graduate School of Biology and Medicine and benefit from advanced training and mentoring opportunities.
Prior experience in cell/developmental biology, genetics/ genomics, or (bio)informatics is highly desired. Interested students should apply with a cover letter stating their motivation, a CV (incl. list of publications if applicable), and contact information for two scientific references to applications_walenteklab@gmx.de .
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Research into human development involves the use of human embryos and their derivative cells and tissues. How religions view the human embryo depends on beliefs about ensoulment and the inception of personhood, and science can neither prove nor refute the teaching of those religions that consider the zygote to be a human person with an immortal soul. In his 
MicroRNAs (miRNAs) contribute to the progressive changes in gene expression that occur during development. In their 