The research group headed by Werend Boesmans and Veerle Melotte is looking for a highly motivated and talented PhD student to investigate the role of the intrinsic innervation of the gut in cancer. The enteric nervous system, also known as the second brain, is a mostly ignored member of the tumor microenvironment but is linked to the development and progression of colorectal cancer. In the current project, the involvement of specific neural cell types in colorectal tumorigenesis will be investigated in both in vivo and in vitro models, and combined with advanced optical microscopy, genetic lineage tracing, marker gene expression analysis and single cell transcriptomics.
This PhD project takes place within the School for Life Sciences (SLS) of the Transnational University of Limburg (tUL), a unique collaboration between Hasselt University (UHasselt, Belgium) and Maastricht University (UM, The Netherlands). The student will perform research at the Biomedical Research Institute (BIOMED, https://www.uhasselt.be/biomed) and the School for Oncology and Developmental Biology (GROW, https://www.maastrichtuniversity.nl/research/school-oncology-and-developmental-biology) embedded in the Department of Pathology of the Maastricht University Medical Center (MUMC).
You can only apply online.
The selection procedure consists of a preselection based on the application file and an interview.
Please provide the contact details of two referents in your application.
Story behind our recent paper in Current Biology “Cadherin-Mediated Cell Coupling Coordinates Chemokine Sensing across Collectively Migrating Cells” (Tugba Colak-Champollion, Ling Lan, Alisha R. Jadhav, Naoya Yamaguchi, Gayatri Venkiteswaran, Heta Patel, Michael Cammer, Martin Meier-Schellersheim, Holger Knaut)
Guided cell migration is a crucial event in many biological and mechanical processes. During development, orchestrated movements of cells pattern tissues and organs. Wounds in our bodies close by the migration of cell sheets. Cell migration is one of the weapons of the immune system, which sends leukocytes to the site of infection to fight against pathogens. But cell migration also contributes to several pathological conditions, such as the dissemination of cancer cells to the sites of metastasis in the body. Thus, detailed studies of mechanisms of cell migration are likely to improve our understanding of animal development, homeostasis, and disease progression.
Cell migration occurs in two major modes: single cell migration and collective cell migration. Single cell migration can be defined as a cell migrating on its own upon extracting directional information and polarizing and moving towards the target site. While collectively migrating cells also extract directional information and polarize toward the target site, they also have the additional task of coordinating with each other to move in the same direction and at the same speed.
How do cells move as a coherent collective following guidance cues? In the latest study from our lab [1], my coauthors and I investigated how directional sensing in collectively migrating cells is organized. We used the Zebrafish lateral line primordium (primordium) as a model for our study. The primordium is a collective of tightly adhering ~100 cells, which originates behind the ear of the fish at around 18 hours post fertilization (hpf) and migrates to the tip of the tail by 42 hpf.
The primordium migration is guided by chemokine signaling. For proper migration, the primordium requires the chemokine Cxcl12a and its receptors Cxcr4b and Cxcr7b. Importantly, the signaling receptor Cxcr4b is expressed in every primordium cell, while the Cxcr7b expression is restricted to the rear of the collective, where it acts as a sink receptor and generates a Cxcl12a gradient across the tissue [2-5].
Schematic of primordium migration and chemokine signaling system in primordium
Primordium migration in wild-type embryo
In collective cell migration, a prevailing model suggests that there is a division of labor between two groups of cells: leaders and followers. These two groups of cells have both topological and functional attributes. For example, leaders are located in front of the collective, whereas followers are located in the rear. In terms of functionality, the model suggests that leader cells explore their surroundings extracting directional information and relaying this information to the follower cells.
It was proposed that the primordium migration also follows a leader-follower model. An elegant study by Haas and Gilmour generated supporting evidence for this model using chimeric analysis [6]. They placed wild-type cells, which can see the chemokine signal into cxcr4b mutant primordia whose migration stalls prematurely. When at least a few wild-type cells ended up in the front region, the migration was restored – albeit at a slower speed than the speed of wild-type primordia.
However, when I started my Ph.D. project in the Knaut lab, two studies that recently were published made my advisor Holger Knaut and me rethink the leader-follower model [4, 5]. These two studies demonstrated that there is a linear chemokine signaling gradient across the tissue that is essential for the directionality of the primordium. This gradient is available to both leader and follower cells, not just to the leader cells.
Meanwhile, Gayatri Venkiteswaran, a post-doc in our lab, made an interesting observation when she was scoring the primordium migration phenotype in chemokine receptor mutants. Previously, it was known that loss of Cxcr4b leads to primordium stalling. However, loss of the other Cxcr4 receptor, Cxcr4a, that is also expressed in the primordium cells [7] does not result in a primordium migration defect. Gayatri found that taking away one copy of cxcr4a from cxcr4b-/-primordia makes the cxcr4b-/- phenotype worse and taking away both copies of both receptors is much worse than the cxcr4b-/- or cxcr4a+/-; cxcr4b-/-phenotypes. This observation suggested that Cxcr4a also contributes to migration and a primordium that is cxcr4b-/- can still see a little bit of the chemokine signal. This meant the earlier study which suggested that the existence of a few chemokine-sensitive cells could restore the migration phenotype of chemokine-blind (cxcr4b-/-) cells was conducted using primordia that could still see some of the directional signal because they retained functional Cxcr4a.
Quantification of primordium migration in 48 hpf embryos of indicated genotypes
Primordium migration in cxcr4b-/- embryo
Primordium migration in cxcr4a-/-;cxcr4b-/- embryo
In the light of these two pieces of new information—existence of a linear gradient across tissue and Cxcr4a’s contribution to directional migration—we decided to take a closer look at the leader-follower model and the contribution of follower cells to directionality and speed of the primordium.
Our first question was whether a few chemokine-sensitive (wild-type) cells could restore the migration phenotype of completely blind (cxcr4a-/-; cxcr4b-/-) primordia. To answer this question, we used a classical developmental biology technique named chimeric analysis. The idea was putting cells from wild-type donor embryos into cxcr4a-/-; cxcr4b-/- host embryos at the blastula stage. The donor primordium cells are labeled with a red transgenic marker and the host primordium cells are labeled with a green transgenic marker. After transplantation, the embryos are grown until 30 hpf and the transplanted host embryos are screened for the presence of red donor cells in the primordium. Although chimeric analysis provides very clean and reliable data, it is an inefficient technique. Unfortunately, not every single embryo transplanted gets the donor cells in the primordium. Additionally, it is a difficult technique and takes a while to perfect it. Even when you become very good at it, you can still have some days when the host mortality is high for unknown reasons.
Schematic of blastomere transplantation
Despite the difficulty and inefficiency of chimeric analysis, we still chose to do it because it was the best way to answer our questions. There was an additional layer of difficulty which turned out to be a good thing later: to generate cxcr4a-/-; cxcr4b-/-embryos, we had to in-cross cxcr4a+/-; cxcr4b-/- fish because cxcr4a-/-; cxcr4b-/- adults are not viable. This meant only a quarter of our host embryos would have the desired genotype. Half would be cxcr4a+/-; cxcr4b-/- and a quarter would be cxcr4b-/-. We had no way of knowing the genotype of the chimeric host embryos until going through a genotyping protocol after imaging these embryos. Nevertheless, we got what we wanted and more.
We found that a few chemokine sensitive (wild-type) cells do not restore the migration phenotype of completely blind (cxcr4a-/-; cxcr4b-/-) primordia, an observation that is inconsistent with the classical leader-follower model. However, we had one embryo in which about half of the primordium consisted of wild-type cells, and the other half consisted of cxcr4a-/-; cxcr4b-/-cells, and this chimeric primordium migrated about half of its path. This made us consider the possibility that cells pull on each other during migration. When there are many cells which can see the chemokine, they might generate sufficient pulling forces in the right direction to drag their chemokine blind neighbors along—in the case of this chimeric embryo, half of the way.
We decided to follow up on this observation using a quantitative approach. We predicted that as we increase the number of chemokine-sensing cells in an otherwise chemokine-blind primordium, the distance migrated by the chimeric primordium should increase. To quantify this behavior, we needed a lot of samples. Using chemokine-blind primordia (cxcr4a-/-; cxcr4b-/-) as hosts posed two challenges. First, it is difficult to get them (remember that only one-quarter of the embryos are of this genotype). Second, it might require a lot of chemokine-sensing cells to pull their completely chemokine-blind neighbors along. To overcome these challenges, we considered using cxcr4a+/-; cxcr4b-/- primordia as hosts. These almost chemokine-insensitive primordia have only one copy of cxcr4a left. Gayatri’s quantification of the migration behavior showed that the migration of these cxcr4a+/-; cxcr4b-/- primordia are worse than cxcr4b-/-primordia and a little bit better than cxcr4a-/-; cxcr4b-/- primordia. Having just a little bit of Cxcr4a activity would help us resolve the relationship between migrated distance and chemokine-sensing cell number, we hoped.
A) Schematic of experimental design and predictions. B-D’) Examples of chimeric primordia with low, medium, and high donor cell contribution. E) Graph of the total number of wild-type cells in the chimeric primordia (total primordial wild-type cells include interneuromast, neuromast (nm) and primordium (prim) cells) versus the migration distance of cxcr4-deficient primordia. 48 hpf.
Luckily, we already imaged a lot of chimeric primordia that consisted of chemokine-sensitive (wild-type) and chemokine-insensitive (cxcr4a+/-; cxcr4b-/-) cells. So we went back to this data set and counted the number of wild-type cells and measured how far these chimeric primordia migrated. As we predicted, the chimeric primordia migrated further as the number of chemokine-sensitive cells increased. This observation was consistent with the idea of cells “pulling” on each other. Next, we asked what could facilitate this “pulling”. We turned to the obvious candidates: cell-cell adhesion molecules. And there are a few of them expressed by the primordium cells including E-cadherin (Cadherin 1, Cdh1) and N-cadherin (Cadherin 2, Cdh2) [8-10].
We first decided to focus on Cdh1 and Cdh2. Our first question was how cdh1-/- and cdh2-/- cells behave during migration. The simplest method of answering this question would be observing the migration behavior of cdh1 or cdh2 mutant primordia under a microscope. However, this was not an option for either of these two genotypes. The problem with cdh1-/- embryos is that they die during gastrulation before the primordium is formed. As for cdh2-/- embryos, they have problems with somite development that affect the proper expression of Cxcl12a (the directional cue). Thus, we went back to our favorite technique, chimeric analysis by blastomere transplantation.
Live imaging of cdh1-/- or cdh2-/- cells in an otherwise wild-type primordium showed that lacking either of the cadherins does not affect migration. These mutant cells co-migrated normally with their wild-type neighbors. This raised the next question: Are these cadherins acting redundantly? To answer this question, we needed to generate cdh1-/-; cdh2-/- embryos to be used as donors. However, this is a real challenge, as the cadherin mutants are not adult viable. To obtain the desired genotype, we had to in-cross cdh1+/-; cdh2+/- fish. The chance of finding a double cadherin mutant embryo is one in sixteen! Despite the odds, we did this experiment—repeatedly. We never found a chimeric embryo whose donor embryo was cdh1-/-; cdh2-/-. But we obtained a good sample size that made us confident to think that either such double cadherin mutant cells do not become primordium cells or they dissociate from the primordium very early on. Despite not being able to find what we set out to find, we encountered some interesting genetic scenarios. For example, cdh1-/-; cdh2+/- cells dissociated from the migrating primordium when they were placed at the tip of the primordium, whereas cdh1+/-; cdh2-/- cells fell off the primordium when they were located at the rear of the primordium. These findings were puzzling until the next observation.
To observe Cdh1 and Cdh2 expression in the primordium, we made BAC transgenic lines that expressed Cdh1-GFP and Cdh2-mCherry. A closer look at the primordia in these transgenic fish revealed that Cdh1 was expressed more in the front region of the primordium and Cdh2 was expressed more in the rear. This could be a possible explanation for the observation mentioned above, that cdh1-/-; cdh2+/- cells fell off from the tip positions and cdh1+/-; cdh2-/- cells fell off from the rear positions. Perhaps Cdh1 was needed more in the front and Cdh2 more in the rear?
The observation of cadherin deficient cells falling off from the primordium also suggested that cell-cell adhesion through cadherins couples the migrating cells. However, is such cell coupling necessary for cells to pull on each other? To answer this question, we transplanted cdh1-/- or cdh2-/- cells into cxcr4b-/-primordia. While cxcr4b-/- primordia have a migration defect due to not being able to see the chemokine signal properly, cadherin mutant cells still have the chemokine receptors and should migrate directionally. To our surprise, cdh1-/- cells located in the front region split away from the cxcr4b-/- cells. But cdh2-/- cells located in the front region pulled the cxcr4b-/- cells along. This finding was consistent with the differential expression pattern of Cdh1 (more in front) and Cdh2 (more in rear) that we had observed earlier. Additionally, a little bit of literature digging revealed that in vitro studies suggested that Cdh1 could withhold more force than Cdh2 [for example 11]. It is plausible that there is increased tension between the donor population, which moves persistently in a specific direction, and the host population, which moves in random directions. A strong physical attachment might be necessary to keep these two groups together.
These findings suggested that the physical coupling of primordium cells is important for the group’s directional migration. But how does cell-cell adhesion affect individual cell directionality within a group? Up on discussions of our data with Martin Meier-Schellersheim, who is a physicist at the NIH and a co-author, we decided to take a quantitative approach using nuclear movement as a proxy for cell movement.
Next, we utilized our favorite technique, blastomere transplantation, once again and placed donor cells (wild type, cdh1-/-, cdh2-/-, or cxcr4b-/-) labeled with H2A-GFP into wild-type host primordia whose cell nuclei were labeled with H2A-mCherry.
Using commercial image software, we tracked the individual donor and host nuclei in different primordium locations at high spatial and temporal resolution. To analyze the tracking data, we used custom scripts to assess directional sensing using three measures: neighbor-neighbor distance, directionality index, and directionality angle. The nuclear tracking analysis showed that directional sensing of cadherin and cxcr4b deficient cells was impaired based on all three categories, lack of Cdh1 having the most severe effect. Together these data showed that in addition to directional cue sensing, efficient migration requires cadherin-mediated cell coupling.
Our observations suggested that cadherin-mediated cell-cell adhesion is important for coordinating cell movements in the migrating primordium. To test this further, we decided to use a gene trap line that expresses functional alpha E-catenin tagged with Citrine (Ctnna1-Citrine) from the endogenous promoter that we recently obtained from Scott Fraser’s lab [12]. Ctnna connects Cadherins on the plasma membrane to the actin cytoskeleton; therefore, lack of Ctnna should impair cadherin-mediated cell-cell adhesion. Chimeric analysis using blastomere transplantation would be the way to approach this experiment since the ctnna-/- embryos die during somitogenesis, before the primordium develops. But luckily, our lab recently developed a protein degradation system named zGrad that could be an efficient alternative to chimeric analysis [13]. As expected, the time-lapse analysis showed that depletion of Ctnna1 resulted in primordia that migrate less directionally. Interestingly, cells separated from each other during migration forming large irregular gaps consistent with the idea that cadherin-catenin complexes mediate the adhesion among the cells in the primordium.
In summary, these results suggest that all the cells in the primordium interpret the directional information and are physically coupled to each other to achieve robust migration. This behavior is not unlike some Turkish folk dances, which are characterized by groups of dancers who hold hands tightly as they dance to a tune in a synchronized fashion. Just as each dancer needs to listen to the music, each cell needs to sense the directional signal in order to coordinate their movements. Through their tight connections, dancers and cells alike synchronize their individual motions, thereby moving in unison.
References
Colak-Champollion, T., et al., Cadherin-Mediated Cell Coupling Coordinates Chemokine Sensing across Collectively Migrating Cells.Curr Biol, 2019. 29(15): p. 2570-2579 e7.
Dambly-Chaudiere, C., N. Cubedo, and A. Ghysen, Control of cell migration in the development of the posterior lateral line: antagonistic interactions between the chemokine receptors CXCR4 and CXCR7/RDC1.BMC Dev Biol, 2007. 7: p. 23.
Valentin, G., P. Haas, and D. Gilmour, The chemokine SDF1a coordinates tissue migration through the spatially restricted activation of Cxcr7 and Cxcr4b.Curr Biol, 2007. 17(12): p. 1026-31.
Dona, E., et al., Directional tissue migration through a self-generated chemokine gradient.Nature, 2013. 503(7475): p. 285-9.
Venkiteswaran, G., et al., Generation and dynamics of an endogenous, self-generated signaling gradient across a migrating tissue.Cell, 2013. 155(3): p. 674-87.
Haas, P. and D. Gilmour, Chemokine signaling mediates self-organizing tissue migration in the zebrafish lateral line.Dev Cell, 2006. 10(5): p. 673-80.
Siekmann, A.F., et al., Chemokine signaling guides regional patterning of the first embryonic artery.Genes Dev, 2009. 23(19): p. 2272-7.
Matsuda, M. and A.B. Chitnis, Atoh1a expression must be restricted by Notch signaling for effective morphogenesis of the posterior lateral line primordium in zebrafish.Development, 2010. 137(20): p. 3477-87.
Revenu, C., et al., Quantitative cell polarity imaging defines leader-to-follower transitions during collective migration and the key role of microtubule-dependent adherens junction formation.Development, 2014. 141(6): p. 1282-91.
Kozlovskaja-Gumbriene, A., et al., Proliferation-independent regulation of organ size by Fgf/Notch signaling.Elife, 2017. 6.
Panorchan, P., et al., Single-molecule analysis of cadherin-mediated cell-cell adhesion.J Cell Sci, 2006. 119(Pt 1): p. 66-74.
Trinh le, A., et al., A versatile gene trap to visualize and interrogate the function of the vertebrate proteome.Genes Dev, 2011. 25(21): p. 2306-20.
Yamaguchi, N., T. Colak-Champollion, and H. Knaut, zGrad is a nanobody-based degron system that inactivates proteins in zebrafish.Elife, 2019. 8.
This project will generate transgenic G. mellonella lines with fluorescent haemocytes, to allow visualisation of the immune response in this non-mammalian model organism. The larvae will be injected with the human fungal pathogen Candida albicans and the haemocytes tracked via time-lapse photography. The aim is to build an evidence base to support the wider adoption of G. mellonella larvae for studying host/pathogen interactions, providing an alternative to the use of mice. We’re looking for an experienced and dynamic Postdoc (Grade E or F) and a 0.5FTE Technician to join the team, alongside an existing PhD student.
For further details of the PDRA/F post see: https://jobs.exeter.ac.uk/hrpr_webrecruitment/wrd/run/ETREC107GF.open?VACANCY_ID%3d066891Q4Cg%1BUSESSION=68D96D4363BB4B7192452B3DEB6F3F37&WVID=3817591jNg&LANG=USA
For further details of the 0.5FTE Tech D post see:
The Tunicate embryology Research Team and the MRI core imaging facility in Montpellier, France, have an opening for a computer scientist in charge of the analysis of 3D + time fluorescence imaging data of living embryos of a class of marine invertebrates, the ascidians. The contract is for up to 4 years.
The selected candidate will use and further develop a tool for the automatic segmentation of fluorescence imaging data obtained using a light sheet microscope, ASTEC (Automated Segmentation and Tracking of Embryonic Cells) developed by the team and its collaborators (for an example output of ASTEC, watch the Video showing a segmented Phallusia embryo).
Missions:
– Use ASTEC to segment and track the imaging data collected by the team’s biologists. Perform statistical analyses of embryonic morphology (reproducibility between individuals and between species, phenotyping,…).
– Develop ASTEC’s ergonomics and man-machine interface to make the system as user-friendly as possible for biologist users. Participate in its deployment on the MRI imaging facility.
– Train biologist users in the use of the tool and set up and maintain a website presenting the system and its documentation to the scientific community.
– Improve ASTEC’s performance after finding its limits and the most important imaging parameters for the successful segmentation and tracking of cells.
Context:
The work will be carried out mainly within the tunicate development research team in Montpellier, Southern France. Frequent interactions will take place with the core facility’s engineers and with the computer scientists developing the core of ASTEC in the Morpheme, ICAR and MOSAIC teams.
Prerequisites:
– Undergraduate studies level in computer science, bioinformatics, applied mathematics, image analysis or physics
– Good knowledge of Python
– Knowledge in statistics
– Interest in biological image analysis
– Some training in biology would be a plus, but is not required
– A sufficient level in written / spoken English to interact daily with non-French speaking scientists. French knowledge not required
Contact:
Apply through the CNRS employment website. An informal contact can be established beforehand with the scientist in charge of the project, Patrick Lemaire, CRBM, patrick.lemaire@crbm.cnrs.fr
The laboratory of Dr. Angelo Iulianella is seeking a graduate student and postdoctoral research scientist to study the establishment and maintenance of cell types identities during neural development. The ideal candidates will have experience in molecular biology, cell culture, microscopy, transcriptional profiling, and neural development. The positions are available from early 2020, although start time is flexible. For postdoctoral candidates, funding is available for 3 years and recent graduates are encouraged to apply.
About us: Dalhousie University is one of Canada’s leading research-intensive universities. Our lab is part of the multidisciplinary Brain Repair Centre (http://www.brainrepair.ca) and Atlantic Mobility Action Project (www.amap.ca), which seeks to understand the development and repair of the neural circuitry of movement. We are situated in a highly collaborative environment with access to confocal and super-resolution microscopy, cytometry, and proteomic facilities.
About Halifax, Nova Scotia: With a population of 400,000 people, Halifax is the capital city of Nova Scotia and the educational, cultural and economic hub of Atlantic Canada. Discover rugged shorelines, sandy beaches, and hiking trails all within reach of the urban centre. Achieve your ideal work-life balance in a beautiful part of the world, while doing amazing science!
Please forward your CV, a statement of your research interest, and reference information by e-mail to:
Cell-cell communication plays a central role in the coordination of morphogenesis and fate specification. Most components of the major signalling pathways have been identified. We however lack a quantitative understanding, in time and space, of the dynamics of signal transduction from the membrane to the nucleus. The CRBM Tunicate embryology Research Team uses molecular and 3D + time live imaging approaches to study this process during ascidian embryogenesis.
The PhD project:
One of our major projects combines experimental and mathematical modelling approaches to produce a quantitative model of the information flow between membrane and nucleus for two major signal transduction pathways.
We are looking for a PhD student to develop an optogenetic control strategy for the FGF/SOS/Ras/ERK and Eph/RasGAP/Ras signaling pathways. This approach will open the way to a variety of questions including: how long does signal transduction take from the membrane to the nucleus? during which phase(s) of its cell cycle is the cell competent to respond to receptor activation? what is the minimum activation time of the receptor needed to produce a stable nuclear response? what is the function linking the activation level of the receptor and that of ERK? The experimental results will be integrated into a mathematical model, in collaboration with theoreticians, which will provide suggestions for further experiments.
Training:
This project is mostly experimental. It will give the selected student a solid expertise in embryology (microinjections, in vitro fertilization…), signal transduction and advanced light-sheet imaging. In addition, the PhD student will frequently interact with our computer science collaborators, the MOSAIC and ICAR teams. Participation in public outreach actions (Science festivals, My Thesis in 180 seconds, …) will be encouraged.
Necessary skills:
Master training in cell biology or development, with a strong interest for embryonic development
An interest in mathematical modelling (no specific mathematical knowledge needed).
A first experience in molecular cloning and confocal/light-sheet microscopy of live samples would be appreciated.
An experience in RNA or proteins microinjection into oocytes would be a plus but is not required.
No Knowledge of French required. Working knowledge in written / spoken English needed.
Application:
This project can be joined directly as a PhD student in fall/winter 2019, or as a Master intern in winter 2019, the PhD only starting in fall 2020. Funding is for 3 years.
The host research team is located at a major Cell Biology institute in Southern France, the CRBM (CNRS /U. Montpellier). All seminars and meetings are in English. The institute has a very well-equipped Imaging core facility, hosting a Luxendo MuViSPIM microscope on which lightsheet microscopy experiments will be carried out.
References linked to the project:
Leggio, B; Laussu J; Carlier, A; Godin, C; Lemaire, P and Faure, E (2019) MorphoNet: An interactive online morphological browser to explore complex multi-scale data. Nat Commun. 10(1):2812
Guignard*, U.-M.Fiuza*, B. Leggio, E. Faure, J.Laussu, L. Hufnagel, G. Malandain, C. Godin#, P.Lemaire# (2017) Contact-dependent cell communications drive morphological invariance during ascidian embryogenesis. bioRxiv 238741 https://www.biorxiv.org/content/early/2017/12/24/238741
U-M Fiuza, T. Negishi, A. Rouan, H. Yasuo, P. LemaireNodal and Eph signalling relay drives the transition between apical constriction and apico-basal shortening during ascidian endoderm invagination (2018) bioRxiv 418988 https://www.biorxiv.org/content/early/2018/09/15/418988
Lemaire P. (2011) Evolutionary crossroads in developmental biology: the tunicates, Development, 138(11):2143-52.
Tassy, O., Daian, F., Hudson, C., Bertrand, V., Lemaire, P. (2006) A quantitative approach to the study of cell shapes and interactions during early chordate embryogenesis. Current Biology 16:345-58
Postdoctoral Position in applying light sheet microscopy to understanding algal-spotted salamander endosymbiosis
A postdoctoral position is available in the laboratory of Dr. David Matus at Stony Brook University to investigate symbiotic and developmental processes with Selective Plane Illumination Microscopy. We have funding for a minimum of one year of postdoctoral support to develop protocols for extensive in vitro and in vivo imaging of cell invasion processes with a focus on the tissue and cellular entry of endosymbiotic algae as they enter their spotted salamander embryo hosts (Ambystoma maculatum). This work is funded by the Gordon and Betty Moore Foundation in collaboration with researchers from University of Arizona, the Bigelow Marine Laboratory, and Gettysburg College (see Shelf Life Episode 11). The project will combine molecular biology, embryology, cell biology, and extensive light sheet imaging. Preferred candidates will have backgrounds in light sheet and/or confocal microscopy, working with modern tissue clearing and staining methodologies and interests in advanced imaging methods at the intersections of cell and developmental biology with ecology and evolution. My laboratory is a part of a modern and well-equipped Department of Biochemistry and Cell Biology at Stony Brook University on Long Island, NY. For further information on our work, please see the following publications on techniques,cell invasion, and the symbiosis.
To apply, please send a letter of interest detailing your expertise, CV and names and contact information of three references to david.matus@stonybrook.edu and apply to the advertised position here.
The Developmental Biology Group (Prof. I. Lohmann) at the Centre for Organismal Studies (COS) in Heidelberg, Germany is looking for a
POST-DOC
to study the role of HOX transcription factors in controlling cellular plasticity and cell fate maintenance.
You will join the Lohmann lab (http://ilohmann-lab.org), which is located at the Centre for Organismal Studies (COS) at the University of Heidelberg in Germany, and studies the role of HOX transcription factors in defining cell type identities using Drosophilaas a model.
Cell fates are controlled by networks of transcription factors (TFs) that activate transcriptional programs realizing the distinct properties of cells of a given cell types.However, how TFs control different cell fates is still un unsolved question. HOX TFs represent an excellent model to address this fundamental problem, since they are broadly expressed yet perform highly specific functions within different cell types. We have previously shown that Hox TFs stabilize cell fate choices by suppressing the multipotency encoded in the genome via the interaction with the Polycomb complex. We now seek for enthusiastic new colleagues to analyse our hypothesis that elimination of the Hox code, which is maintained throughout the lifetime of an organism, results in “memory-less” naïve cells that are easy to reprogram.
The Heidelberg Molecular Life Science Community offers a vibrant molecular research community, as well as state-of-the-art core facilities.
Successful candidates should have experience in genomic approaches like RNA-seq, ChIP-seq, ATAC-seq, analysis of genomic data, confocal microscopy, advanced immunohistochemistry and possibly in Drosophila genetics and molecular biology. The ability to quickly integrate into an interdisciplinary team and work independently within an academic research environment is essential. The position is immediately available, the salary is according to TV-L regulations. Disabled persons with comparable skills will be preferentially considered.
The University of Oklahoma College of Arts and Sciences is excited to announce three open faculty positions at any rank in the Department of Biology. As part of our Biology of Behavior strategic initiative, the department invites individuals with creative, innovative, and dynamic research programs who are interested in joining a strong group of researchers to apply for these faculty positions:
A Geneticist who uses integrative molecular approaches to understand the evolution, specification, and/or regulation of how genes affect organismal behavior.
A Physiologist who studies the endocrine regulation and modulation of behavior.
An Evolutionary Developmental Biologist who studies how developmental processes give rise to organismal morphology, nervous system structures, and/or physiology that lay the foundation for the generation of behavior.
The anticipated start date is August 2020. For additional details on these positions, applicant qualifications, and how to apply, please visit http://ou.edu/bb and http://www.ou.edu/cas/biology.
Screening of candidates will begin October 15, 2019 and will continue until the positions are filled.
The University of Oklahoma is an EO/Affirmative Action institution http://www.ou.edu/eoo/. Individuals with disabilities and protected veterans are encouraged to apply.
How would you create a hole between two sticky surfaces? Simply crack it!
At a first glance, trying to pull apart the two surfaces seems to be a good idea, but in practice, you might need a lot of energy. However, it seems that the mouse embryo has found a smart and efficient way to do so during its pre-implantation development. After three rounds of cellular divisions, the 8-cell stage embryo starts to compact: cell-cell contacts are expanding, making the embryo more spherical instead of a collection of bubble-like cells [1]. After another round of cleavage, it also internalizes the cells that are more contractile [2]. They will become the Inner Mass Cell (ICM), the future fetus proper, while less contractile cells, the Trophectoderm cells (TE) form a squamous epithelium, that surrounds the ICM and will become part of the placenta. From this step, the embryo is almost spherical, with two layers of cells.
Then, at the 32-cell stage, the embryo shows a new feature: a lumen, a fluid-filled cavity, that breaks the previous radial symmetry by forming at the interface of TE and ICM cells. To grow a lumen, three conditions are needed: 1- to have a sealed compartment, here ensured by the tight junctions between TE cells at the embryo surface; 2- to draw water towards the sealed compartment: in our case, the mouse embryo builds an osmotic gradient by pumping ions in the intercellular medium and lets the water flow through pores; 3- and finally, you have to make room for the accumulated fluid. But here is the problem: the blastocoel forms systematically on the basolateral side of the TE cells, where the cells strongly adhere together! In most other examples of lumen formation, the opening happens at the apical side of the epithelium, where adhesion is repressed! Thus, arises the question: how can you create a lumen at the adhesive side of cells?
In our research [4], we combine developmental biology and physics to decipher the mechanisms of the embryogenesis. We have found that the apparition and the positioning of the lumen, the so-called blastocoel, can be explained using simple physical and biological concepts.
The formation of the blastocoel was a long-time debated topic. Studies have mainly focused on the expansion phase, when the blastocoel is already positioned, while its initiation and positioning are still poorly understood.
What Julien did first, with the help of Francesca and Ludmilla, was to look at the steps preceding the apparition of the blastocoel. In the last decades, efforts have been done on culture conditions and advances in microscopy have permitted to reduce light exposition while improving spatiotemporal resolution. Using resolutive imaging in space and time, that involved the use of both transgenics and microscopy techniques, heobserved the embryo literally boiling!Hundreds of bubbles appeared at the intercellular contacts before the final lumen, forming a network of small microlumens throughout the embryo. Some of those microlumens grew in size, while others disappeared (Fig. 1). As biologist, this observation might seem not significant, but for physicists, this coarsening process immediately rang a bell: looking at the movies, it was really analogous to a well-known process in soft-matter physics: Ostwald ripening. Basically, it describes how in a vinaigrette, the droplets of vinegar will coarsen into fewer drops, the bigger droplets growing to the detriment of the smaller.
From this observation, the collaboration between the two teams emerged, with one team of biologists (Julien, Francesca, Ludmilla and Jean-Léon), the other of physicists (Mathieu, Annette and Hervé), with two questions: i- how these water pockets form in spite of cell adhesion? ii- as they form ubiquitously through the embryo, what mechanisms ensure the formation of a single blastocoel and its final positioning?
Figure 1: Microlumens appear at the onset of cavitation and coarsen to form a final fluid-filled lumen, the blastocoel, breaking the symmetry of the embryo.
The microlumens form and expand in the extracellular space, at the interface between cells. Julien looked at those cellular contacts, showing that during the formation of microlumens, the spatial distribution of adhesion molecule (E-cadherins) evolves from a homogeneous to a localized heterogeneous distribution. From this observation came the idea of hydraulic fracturing, where water pressure cracks cell-cell contacts exactly like it would crack the rock in oil fracking [3].
After discussions, we came with two main scenarii. a- as cells are active material, they could autonomously regulate their adhesion and create weak points where the fluid could accumulate; b- adhesion is a force that opposes to fluid accumulation, and the expansion of microlumens is capable of pushing adhesion molecules away. To answer this, we had no direct way to measure inside the embryo how cells react against an increase of pressure in the intercellular space. So instead, we chose to inhibit (in three different ways) the formation of the microlumens. in the absence of microlumen, we couldn’t see any reorganization of the E-Cadherin. Our favorite interpretation from this result: this is the hydraulic pressure that breaks locally the adhesion between the cells, and from these breaking points, microlumen can expand. In a nutshell, the embryo seems to generate an increase of hydraulic pressure to break apart all cells contacts instead of specifically regulating its adhesive properties.
A coarsening process is generally made possible by the exchange of matter between different compartments. In the mouse embryo, the microlumens can exchange fluid via the intercellular contacts, which connect them throughout the embryo. A coarsening process akin to Ostwald ripening furthermore involves two other key features: the only stable state is a single droplet, and it requires a surface tension at droplet interfaces, which generate the pressure driving fluid exchange. In the embryo, we invariably observe the formation of a single lumen, and it is furthermore always located at the interface in between the TE and ICM. Thus, we quickly came to the idea that playing on the cell “surface tension” would give us great insights into the mechanical aspects of the blastocoel formation. Indeed, according to previous studies [1], we knew that ICM and TE cells have different levels of contractility, that can physically be translated into surface tensions.
We therefore built a theoretical model of the network of microlumens as a two-dimensional graph of connected hemispherical drops, to test in silico the physical predictions with an algorithm developed by Annette and Mathieu, and we designed experiments to test in situ the biological predictions. Our combined results suggest that, due to osmotic gradient and active pumping, the cells inject fluid that pressurizes the intercellular space, hence creating the hundreds of microlumens by disrupting the cell-cell adhesive contacts. The newly formed microlumens then coarsen into a single final lumen, with a characteristic biphasic dynamic of collective growth then shrinkage, observed both for the model and for the myriad microlumens measured by Julien.
From there, Mathieu predicted with the model the formation of the blastocoel on the side of the embryo, between TE and ICM cells, hence breaking the symmetry of the embryo (Fig. 2., left panel). This prediction was tested using chimeric embryos that Julien made (Fig. 2, center and right panels): an equal mixture of wild-type cells and low-contractility cells, deficient in myosin activity, shows a clear bias for the final position of the lumen toward the low-contractility domain of the chimera, as the theory predicted. The experiments of Julien on low-adhesion mutants, lacking half of E-cadherin activity, also predicted effects of the partial loss of adhesion, that were then tested by Mathieu on the theoretical model, confirming the whole process as being a trade-off between adhesion and contractility.
Figure 2: (Left) Probability of blastocoel formation for ICM-ICM (blue) or TE-ICM (red) multi-cellular lumens vs the tension asymmetry ratio from simulations. (Right) In chimeric embryos, the final position of the blastocoel is biased towards less contractile part of the embryo (Myh9 deficient cells).
One fascinating aspect of the process is how much it is robust: even though breaking adhesive cell-cell contacts may not be thought to be the best process to form a lumen, the mouse embryo succeeds to form an internal cavity. Moreover, despite the immense molecular complexity of embryo development, the formation of the blastocoel follows rather simple physical mechanisms.
The project was going back and forth between theory and experiment all along. The geographic proximity and the excellent relationship between the two teams were key factors, speeding up the process and the constant exchanges, helping us to remove the barrier between theoretical biophysics and developmental biology.
It was a quick and extremely stimulating project for both our young teams. Part of the pleasure of the project was to gather many people with various expertise, and to see all the pieces matching together to give a comprehensive model at many levels, and of course to work with such enjoyable people.
I, Julien, come from the zebrafish community, and I was used to image embryos that develop fast, as the embryo looks like a fish in 24 hours after fertilization. Then I started to work in Jean-Léon’s team and image relatively slow embryonic development (the mouse embryo takes 3 days to build the blastocyst). Nonetheless, I was convinced that we were missing (and most probably are still missing) key steps of mammalian development and decided to push the system further. Having time resolution of minutes or seconds led to these incredible observations and were key is the direction in which we pushed our research. What I will retain from this work is the exciting collaboration with Annette, Mathieu and Hervé, that opened a new field for me: I must confess, I never heard of coarsening before! I am really happy to see that physicists can be as amazed as developmental biologists by embryogenesis and that these enthusiastic interactions can lead to exciting discoveries.
As far as I (Mathieu) am concerned, this was my first real scientific contribution, ending with a beautiful paper. The fact that such a key step in the mouse embryo development can be simply seen as a fracking and coarsening process still amazes me. Starting the study of the morphogenesis of the mouse embryo was a real challenge with my background of theoretical physicist. Hopefully, Julien and others from his team were always more than happy to speak, and to show me what they were doing, which was an invaluable help. I could not think of better conditions as a start for my PhD, and I am really thrilled to see where it will go.
Figure 3 : Representative set of collaborators (n = 6). From left to right: Ludmilla, Julien, Francesca, Jean-Léon, Hervé and Mathieu.
It opened so many foods for thoughts, promising new and exciting results about the development of the mouse embryo, that we are now trying to push forward. Are there factors that favor the final position within the embryo or is it a stochastic phenomenon? What are the consequences of this increase of pressure on cells at molecular and genetic levels? What triggers the initiation and nucleation of the microlumens?
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