We seek a highly motivated postdoctoral fellow to spear head research investigating the genomic basis of transcriptional specificity of cell signaling during the directed differentiation of human pluripotent stem cells into the digestive and respiratory organoids. The goal is to understand how different transcriptional programs are activated by the same cell signaling pathways, at different times in development and disease. You will join a multidisciplinary team in the Zorn Lab, Division of Developmental Biology at Cincinnati Children’s Hospital Research Foundation.
Qualified applicants will have a PhD with peer review research publications, a demonstrated expertise in genomic analysis, and a keen interest to develop an independent research program in the area of genomics, development and stem cell biology.
Please submit your application to aaron.zorn@cchmc.org with the following information: a cover letter, statement of interest, and CV with contact details for 3 referees.
Closing Date: May 31st, 2017
The Cincinnati Childrens Hospital Medical Center, and the University of Cincinnati are Affirmative Action/Equal Opportunity Employers. Qualified women and minority candidates are especially encouraged to apply.
Department/Location: Wellcome Trust – Medical Research Council Cambridge Stem Cell Institute
Reference: PS11650
Closing date: 31 March 2017
Collaborative PhD Studentship with Astra Zeneca
Elucidating cellular behaviours and potential oncogenicity utilising in vitro organoid culture system Project Description
Department name: Cambridge Stem Cell Institute, University of Cambridge Supervisors: Primary: Dr Joo-Hyeon Lee (University of Cambridge) and Dr Mick Fellows (AstraZeneca)
Assessment of cell transformation (i.e. the acquisition of malignant characteristics in morphology, growth control or function) has proven problematic to model in vitro. The soft agar cell transformation assay has been validated, but only in a limited number of immortalised or mixed population embryonic cells and the specificity of the assay has been questioned. Furthermore, the genetic basis of cell transformation in this model has not been fully delineated (Harvey et al 2015, Creton et al 2012). The use of genetically and phenotypically stable organoids derived from single human stem cells (Schwank et al 2013) should provide a better model for cell proliferation and transformation.
Organoids have the advantage of 3D morphology, being able to be derived from several major organs and can be engineered to demonstrate proliferative effects by introducing a cell type specific ki67 fluorescent protein to tag. Organoid stability is also dependent on several growth factors and inhibitors e.g. EGF, Wnt, BMP inhibitor, TGFb inhibitor. Growth following withdrawal of these factors, which are known pathways implicated in cancer genesis, will be indicative of cell phenotypic and/or transformative changes. It is also proposed that this model could be used to investigate mechanisms of pro-oncogenicity and also the potential off-target effect of genome editing by CRISPR/Cas9, for which there is currently an unmet need for in vitro assays to assess concerns around inappropriate editing in oncogenes.
The project will develop organoids from a variety of tissue sources, including organoids with the ki67 tag, and assessment of proliferative and morphological changes following withdrawal of growth factors after treatment with reference carcinogens and following transfection with specific and promiscuous guide RNA and associated Cas9 protein. Comparative data will be generated from alternative in vitro methodologies to analyse cell transformation e.g. the soft agar assay. Reference carcinogens will be analysed in this assay along with the potential of CRISPR/Cas9 editing to induce cell transformation (which has not been previously assessed). The student will also use next generation sequencing technologies to identify specific genes involved in morphological changes in both cell and organoid cultures. Success will provide new and more relevant model for understanding and assessment of morphological changes and carcinogenesis.
Eligibility and Funding
This studentship covers 4 years’ UK/EU tuition fees (see below for EU eligibility requirements) and a maintenance stipend.
Fully-funded PhD studentships (3 years) for projects in Mathematical/Computational Biology are available at the University of Southampton (Mathematical Sciences). The PhD projects will be supervised by Drs Philip Greulich and Ben MacArthur and will involve mathematical and computational modelling of stem cell dynamics in biological tissues, in particular related to cancer and development. The candidate should have, or should expect, an upper second-class degree or higher, and a genuine interest in biological problems.
An outline of two potential PhD project topics can be found here:
Hanieh Falahati, Lewis–Sigler Institute for Integrative Genomics, Princeton University.
Eric Wieschaus, Howard Hughes Medical Institute, Department of Molecular Biology, Princeton University.
The whole universe is moving toward disorder; this is the second law of thermodynamics in simple terms. Yet, living organisms have found a way to keep themselves organized by spending energy. Cells need this organization in order to provide specialized microenvironments for different cellular functions. Two types of intracellular organizations can be distinguished in biological systems: The first type is membrane-bound organelles such as ER, or lysosome. The composition of these organelles is maintain by spending energy and active transport of molecules across a membrane. The second type of intracellular organization comes from membrane-less organelles such as nucleoli, histone locus bodies, and stress granules that are high-concentration assemblies of different proteins and RNAs. A puzzling question is that without a membrane, how are cells able to form and maintain these organelles.
A historical perspective
The question of how membrane-less organelles form immediately captured my attention the first time I heard about it in an introductory meeting with my doctoral advisor, Prof. Eric Wieschaus. Of course this question has fascinated many scientists since the initial observation of the nucleolus, the quintessential membrane-less organelle, in the 18th century. In 1898, Montgomery performed a very comprehensive study of the nucleoli of different cell types, hand-drawn in 346 figures (a sample is shown in Fig. 1), and concluded that the nucleolus forms when a substance from the cytoplasm enters the nucleus and is deposited there “in the form of masses of varying dimensions, which may be either globular or irregular in shape, according as they are fluid or viscid in consistency”. He further describes that nucleoli form via “coalescence of numerous small portions of nucleolar substance”, consistent with its fluidity (Montgomery 1898). However, this model faded away with the advancement in cell biology and genetics which showed that the nucleolus forms around ribosomal DNA repeats and is the site of active transcription and processing of ribosomal RNA, and ribosomal biogenesis. By the end of the 20th century, nucleolus was commonly thought to form actively, in the process of making ribosomes, until a number of influential works from Hyman and Rosen groups redirected the focus to the liquid nature of the membrane-less organelles (Brangwynne et al. 2009; Li et al. 2012). Based on this liquid property, it was proposed that formation of membrane-less organelles is a spontaneous liquid-liquid phase separation (LLPS), similar to the separation of oil from water.
Figure 1. Hand drawing of nuclei with nucleoli (n) of different cell types drawn by Montgomery in 1898. N. Gr.: nuclear granule. Chr.: Chromatin.
By the time I joined the Wieschaus lab at Princeton, two main hypotheses existed for the formation of membrane-less organelles: 1. The LLPS model, in which the components of these organelles assemble spontaneously in a thermodynamically-driven process due to their favorable intermolecular interactions, and form a liquid phase; 2. The active assembly model which suggests that these organelles form as a result of an active process, which happens inside the cells or inside the organelles. Such active processes are reactions carried out by enzymes that couple these reactions to an energy source such as ATP. The evidence for the LLPS model comes from experiments showing that many of the proteins which localize to the membrane-less organelles have the physical properties to phase separate in vitro. However, in most cases the phase separation is only observed at concentrations much higher than cellular levels. Therefore, a pressing question is whether the physical properties of these components would also drive their spontaneous assembly in vivo, or active processes are responsible for their congregation. Accordingly, we decided to develop an in vivo test that would allow us to distinguish between the LLPS and the active mechanism, and apply it to study nucleolus assembly as a model for the formation of membrane-less organelles.
Working out a path – How to test the phase separation model in vivo?
Developing such an in vivo test was not trivial, since compared to in vitro systems, living cells are far more limited in the ways they can be manipulated. But considering the expertise of my advisor, Eric Wieschaus, in manipulating Drosophila embryos, his lab was the best place to do this type of research. It was not the only reason for us to study this question in fly embryos: Development in Drosophila starts without a nucleolus, and the nucleolus forms at a particular developmental stage (Video 1). This allows us to manipulate the system and study its effect on the nucleolus assembly. In addition, nucleolus forms during the 13 synchronous divisions at the beginning of development, therefore at each time-point there are multiple nuclei at the exact same developmental stage, which helps with statistics. Finally, during these 13 division the nuclei share the cytoplasm, which means that all the nuclei have access to the same concentration of most molecules. These features make fly embryos an ideal system for studying nucleolus assembly.
Video 1. Nuclei of D. melanogaster embryos start development without a nucleolus, and form the nucleolus (bright foci) for the first time at nuclear cycle 13.
How can you test whether the formation of an organelle in vivo is a thermodynamically driven LLPS, or an active assembly process? In general, LLPS processes are affected by two factors: concentration and temperature (Fig. 2). Increasing the concentration results in an increase in the size of assemblies: The more oil you add to the water, the bigger the oil droplets. However, active processes are also affected similarly with concentration: The higher the concentration of the reactants, the more products (assemblies) are formed. Therefore, changes in the concentration cannot distinguish between an LLPS and an active assembly. On the contrary, temperature often has a differential effect on these two processes: In general, lowering the temperature causes more condensation and enhances phase separation, but slows down active enzymatic reactions. This means that by quantifying the effect of various temperatures on the assembly formation, one can distinguish between an LLPS and an active assembly model.
Figure 2. (Left) Phase separation processes are affected by concentration and temperature. Increasing the concentration and decreasing the temperature results in the formation of larger assemblies. (Right) Active processes are carried out by enzymes which couple them to an energy source. Therefore, the rate of these processes is determined by the rate of enzyme activity, which is reduced at lower temperatures.
At the time, our strategy to examine the effect of temperature on the formation of assemblies was received with skepticism among the community. The main concern was whether in the temperature range tolerable by biological systems, an effect would be detectable. A reassurance, however, was an exquisite work by Sarah Veatch, who showed that plasma membrane vesicles isolated from living cells exhibit phase transitions between 15 and 25°C (Veatch et al. 2008). Subsequently, we employed a microfluidic device that allowed us to control the temperature of a fly embryo between 6 and 31°C (Fig. 3). We used this microfluidic device to examine the effect of temperature on the first time formation of assemblies, and also to test the reversibility of assembly formation, for six different nucleolar proteins.
Figure 3. A microfluidic device was used to control the temperature of an embryo in a range of 6 to 31°C by flowing in warm or cold water.
The unexpected result of two independent mechanisms
Much to our surprise, the results of our test showed that two independent mechanisms govern the assembly of different subsets of nucleolar proteins: While four of the studied proteins depict properties of LLPS, the two others are recruited actively. At lower temperatures, the phase separating proteins assemble at an earlier developmental time, and dissolve upon heating (Video 2). The latter is consistent with the reversibility of thermodynamic processes. On the contrary, the formation of active proteins is inhibited at low temperatures, and is irreversible, an exclusive property for active processes.
Video 2. The nuclei of intact fly embryos are subjected to temperature changes in the surrounding fluid. As the temperature is shifted from low to high, the phase separated proteins dissolve, as can be seen in the disappearance of the bright spots.
rDNA: Come together, right now, over me
Our unexpected result that two independent mechanisms govern nucleolus assembly in vivo raised a new question: How are these two different mechanisms coordinated to form a single organelle? In a previous paper, we had shown that transcription of rDNA dictates the spatiotemporal precision in the formation of assemblies by the phase separating proteins (Falahati et al. 2016). Interestingly, we were also able to show that rDNA is necessary for the recruitment of the two actively assembling proteins. This suggests that rDNA can function as a coordinator between the two independent mechanism. In addition, it highlights the fact that in vivo, even the assembly of phase separating proteins is regulated by active processes such as transcription.
We are very enthusiastic about the publication of this paper as it is a significant advancement in the field both from a methodological and mechanistic standpoint. We took an interdisciplinary approach and developed an invaluable tool for closing the gap between the current knowledge of the in vitro self-assembly and the formation of membrane-less organelles in vivo at its full complexity. From a mechanistic perspective, the large number of proteins studied here allowed us to unravel the unappreciated complexity in the formation of intracellular assemblies, as our results show the presence of at least two independent mechanisms for the recruitment of nucleolar proteins. We are confident that the method and the results introduced in this paper will set a framework to better understand how normal or pathological intracellular assemblies form.
Our latest monthly trawl for developmental biology (and other cool) preprints. See June’s introductory post for background, and let us know if we missed anything
Another month, another bumper crop of preprints covering everything from calcium waves in the fly to the first mutant ant line, plant superhero genes to rat embryonic stem cells. Plus, as usual, a whole bunch of tools and resources (including a 3D printed micropipete!), and right at the end some enigmatic bacterial structures.
The preprints were hosted predominantly on bioRxiv, though we also found a handful on arXiv and PeerJ.
Multiple roles of PIWIL1 in mouse neocorticogenesis. Barbara Viljetic, Liyang Diao, Jixia Liu, Zeljka Krsnik, Sagara H. R. Wijeratne, Ryan Kristopovich, Marina Dutre-Clarke, Matthew L. Kraushar, Jimin Song, Jinchuan Xing, Kevin Chen, Mladen-Roko Rasin
Laser ablation tomography image of a nodal root from a maize plant, from Bucksch, et al.
Morphological plant modeling: Unleashing geometric and topological potential within the plant sciences. Alexander Bucksch, Acheampong Atta-Boateng, Akomian Fortune Azihou, Mathilde Balduzzi, Dorjsuren Battogtokh, Aly Baumgartner, Brad Binder, Siobhan Braybrook, Cynthia Chang, Viktoriya Coneva, Thomas DeWitt, Alexander Fletcher, Malia Gehan, Diego Hernan Diaz Martinez, Lilan Hong, Anjali Iyer-Pascuzzi, Laura Klein, Samuel Leiboff, Mao Li, Jonathan Lynch, Alexis Maizel, Julin Maloof, RJ Cody Markelz, Ciera Martinez, Laura Miller, Washington Mio, Wojtek Palubicki, Hendrik Poorter, Christophe Pradal, Charles Price, Eetu Puttonen, John Reese, Ruben Rellan-Alvarez, Edgar Spalding, Erin Sparks, Chris Topp, Joseph Williams, Daniel Chitwood
Tex19.1 Restricts LINE-1 Mobilisation in Mouse Embryonic Stem Cells. Marie MacLennan, Marta Garcia-Canadas, Judith Reichmann, Elena Khazina, Carmen Salvador-Palomeque, Abigail Mann, Paula Peressini, Laura Sanchez, Christopher J Playfoot, David Read, Chao-Chun Hung, Ragnhild Eskeland, Richard R Meehan, Oliver Weichenrieder, Jose Luis Garcia- Perez, Ian R Adams
Sagittal sections of mandibular molars from Feng, et al.
Functional Annotation of Chemical Libraries across Diverse Biological Processes. Jeff S Piotrowski, Sheena C Li, Raamesh Deshpande, Scott W Simpkins, Justin Nelson, Yoko Yashiroda, Jacqueline Barber, Hamid Safizadeh, Erin Wilson, Hiroki Okada, Abraham Gebre, Karen Kubo, Nikko Torres, Marissa LeBlanc, Kerry Andrusiak, Reika Okamoto, Mami Yoshimura, Eva DeRango-Adem, Jolanda van Leeuwen, Katsuhiko Shirahige, Anastasia Baryshnikova, Grant W Brown, Hiroyuki Hirano, Michael Costanzo, Brenda Andrews, Yoshikazu Ohya, Hiroyuki Osada, Minoru Yoshida, Chad L Myers, Charles Boone
TOWARDS COORDINATED INTERNATIONAL SUPPORT OF CORE DATA RESOURCES FOR THE LIFE SCIENCES. Warwick Anderson, Rolf Apweiler, Alex Bateman, Guntram A Bauer, Helen Berman, Judith A Blake, Niklas Blomberg, Stephen K Burley, Guy Cochrane, Valentina Di Francesco, Tim Donohue, Christine Durinx, Alfred Game, Eric Green, Takashi Gojobori, Peter Goodhand, Ada Hamosh, Henning Hermjakob, Minoru Kanehisa, Robert Kiley, Johanna McEntyre, Rowan McKibbin, Satoru Miyano, Barbara Pauly, Norbert Perrimon, Mark A Ragan, Geoffrey Richards, Yik-Ying Teo, Monte Westerfield, Eric Westhof, Paul F Lasko
Uncharacterized bacterial structures revealed by electron cryotomography. Megan J Dobro, Catherine M Oikonomou, Aidan Piper, John Cohen, Kylie Guo, Taylor Jensen, Jahan Tadayon, Joseph Donermeyer, Yeram Park, Benjamin A Solis, Andreas Kjaer, Andrew I Jewett, Alasdair W McDowall, Songye Chen, Yi-Wei Chang, Jian Shi, Poorna Subramanian, Cristina V Iancu, Zhuo Li, Ariane Briegel, Elitza I Tocheva, Martin Pilhofer, Grant J Jensen
A postdoctoral research fellow position is available in the laboratory of Gregor Andelfinger at the CHU Sainte Justine Research Center, Montréal, Québec, Canada. Our work focuses on the genetic origins of cardiovascular diseases in the young and includes both genomics and functional approaches. In this project, the successful candidate will apply genomic and molecular biology approaches to develop disease models. Postdoctoral fellows will have a unique opportunity to do interdisciplinary research at all interfaces of translational medicine and publish their results in high impact journals.
The candidate will work in collaboration with all lab members and is expected to independently perform human and mouse genetic studies, as well as in vitro molecular analyses. Experience with a wide variety of techniques (developmental studies, histology, confocal microscopy, cell culture assays, RNAseq) is desired. Experience with genome editing techniques (CRISPR) and/or and bioinformatics analysis is an asset.
The successful candidate must have a recent Ph.D. degree in life sciences, molecular cell biology, or a related field. We are seeking highly motivated and well published researchers with excellent oral, written, and interpersonal communication skills.
Montreal is a vibrant bilingual city with a high quality of life. With a greater metropolitan area of 3.5 M, Montreal offers all scientific and cultural benefits of a large North American agglomeration with a European flair and beautiful surroundings.
The Andrew Yoo lab at WUSTL is looking for a lab manager/technician. The Yoo lab uses microRNA-mediated direct reprogramming of fibroblasts to neurons to study aging and neurodegenerative diseases.
PhD position opened in Switzerland for a developmental or cell biologist
Development & Evolution of structural colours in vertebrates
In the context of a multidisciplinary study combining cell biology, evolutionary developmental biology and physics, we offer one PhD student position for an outstanding, highly motivated, and creative experimental wet-lab biologist with strong skills in developmental biology or cell biology.
A major research project in Michel Milinkovitch’s group (Laboratory of Artificial & Natural Evolution; www.lanevol.org) at the University of Geneva (UNIGE) is to better understand the molecular developmental mechanisms generating a diversity of skin colours and patterns in reptiles.
One aim is to understand the development & evolution of surface gratings and intracellular nano-structures generating colours through the physical phenomenon of light interference. The successful candidates will use state-of-the-art microscopy and biochemical/genetic manipulations in both the zebrafish and chameleons to understand the development and evolution of surface gratings and intracellular photonic crystals.
Candidates must have a Master in biology or biochemistry. Skills and experience with developmental biology and/or cell biology are mandatory. The successful candidate will have a genuine interest for organismal biology and will appreciate interactions with developmental biologists, evolutionary biologists, physicists and computer scientists. The PhD student will be localised in the Milinkovitch lab (http://www.lanevol.org) and will have very regular interactions/collaborations with members of Marcos Gonzalez-Gaitan’ lab (https://cms.unige.ch/sciences/biochimie/-Marcos-Gonzalez-Gaitan-Lab-.html).
The University of Geneva is world-renowned for its research in Biology and Physics and is among the top 1% best universities in the world.
PhD students are remunerated according to the standards of UNIGE, which are very generous when compared to other international programs.
Geneva is an international city occupying a privileged geographical situation with its beautiful lake and the close-by Alps.
Refs: Saenko et al.Precise colocalization of interacting structural and pigmentary elements generates extensive color pattern variation in Phelsuma lizards.BMC Biology 2013, 11: 105; Teyssier et al.Photonic Crystals Cause Active Colour Change in Chameleons.Nature Communications 6: 6368 (2015); Tzika et al. Reptilian Transcriptomes v2.0: An Extensive Resource for Sauropsida Genomics and Transcriptomics. Genome Biol. Evol. 7: 1827-1841 (2015); Ullate-Agote et al.The genome sequence of the corn snake (Pantherophis guttatus), a valuable resource for EvoDevo studies in squamates.Int. J. Dev. Biol. 58: 881-888 (2014); Saenko et al.Amelanism in the corn snake is associated with the insertion of an LTR-retrotransposon in the OCA2 gene.Scientific Reports 5, 17118 (2015); Di-Poï & Milinkovitch. The Anatomical Placode in Reptile Scale Morphogenesis Indicates Shared Ancestry Among Skin Appendages in Amniotes. Science Advances 2, e1600708 (2016).
Candidates must send their application — in the form of a single PDF file including a brief letter of interest, a CV, as well as contact information (not support letters) of two persons of reference — to: Prof. Michel Milinkovitch (michel.Milinkovitch@unige.ch).
We are looking for a postdoc interested in studying neural development in tunicates, the sister group to the vertebrates. The larval nervous system of the tunicate Ciona has only 177 neurons and its entire “connectome” has been recently mapped (Ryan et al., eLife 2016). The Ciona genome is also highly compact and easily manipulated using CRISPR/Cas9 (Stolfi et al., Development 2014). This tractability offers an unprecedented opportunity to understand gene networks underlying the development of every single neuron in a chordate nervous system.
We have several potential projects centered on understanding the link between transcriptional regulation and cell behavior during the development of the Ciona nervous system. Specific cells of interest include homologs of vertebrate spinal cord neurons (Stolfi et al., Development 2011) and neural crest-derived sensory neurons (Stolfi et al., Nature 2015). Projects will involve isolating neural progenitors from transgenic Ciona embryos, profiling their transcriptome and chromatin states using next-generation sequencing, and targeting candidate regulators and effectors using CRISPR/Cas9.
Our lab is located at the Georgia Institute of Technology in Atlanta, USA. Georgia Tech offers first-rate research facilities and a highly collaborative research environment with traditional strengths in engineering, computing, and math. We are dedicated to promoting equity of under-represented minorities in academia.
Position highlights: The Grant Strategist will support the productivity of Zuckerman Institute scientists by editing and consulting on grants, particularly sections on research strategy, aims, and narratives; acquire and maintain sound knowledge of the Institute and the research priorities; use that knowledge and understanding to seek and recommend grants for individual as well as multi-investigator grants within and between disciplines.
The Grant Strategist will serve as a project manager to pursue, identify and promote funding opportunities for multi-PI grants. The Grant Strategist will also consult with and keep abreast of the current granting environment to provide suggestions to individual researchers on specific grant opportunities.
Position requirements: Ph.D. in the biological sciences or related field, including extensive writing experience; or advanced degree in journalism with deep expertise in science. Minimum of four (4) years of successful science grant writing experience or related experience in science communication field is required. Extensive knowledge of scientific granting processes, particularly in the biological sciences; successful grant writing or consulting record is vital.
(No Ratings Yet)
Loading...
(2 votes)
