With an overwhelming majority, this crisp fly embryo staining won the third round of cover voting, in which readers of the Node chose from images taken in the 2011 Woods Hole Embryology course to select a cover for an upcoming issue of Development.

It shows a ventral view of stage 16 Drosophila melanogaster embryo immunostained for Tropomyosin (green; muscle), Pax 3/7 (blue; segmentally repeated nuclei in CNS and ectoderm), and anti-HRP (red; cell bodies and axons of the nervous system). All nuclei shown in gray (DAPI).

The winning image was captured by Julieta María Acevedo of the Fundacion Instituto Leloir in Argentina, and Lucas Leclere of the Sars International Centre for Marine Molecular Biology in Norway. Congratulations!

The runner-up was an immunostaining of a butterfly wing disk, taken by Alessandro Mongera, Maria Almuedo Castillo, and Jakub Sedzinski. The image of a C. elegans head, taken by Eric Brooks and John Young, got third place, and the final image, of a 3rd instar Drosophila wing disk, was the work of Lynn Kee.

Meanwhile, you’ll see the winner of the previous round on the cover of Development tomorrow, so keep an eye on the journal website.

(1 votes)

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British researchers working with human embryos for IVF have been wondering about the effects of components of the culture medium they use.

In the UK, culture media used for IVF and fertility research are regulated by the Medicines and Healthcare Regulatory Agency (MHRA) and at European level. Regulation ensures that all researchers and clinicians in the field are using industry standard medium, but manufacturers can change the composition of the media at any time without input from its end users.

When IVF researchers became curious about the role of the various components in the culture medium, and whether the manufacturers could justify changes to the medium, they contacted the HFEA (Human Fertilization & Embryology Authority) – an independent regulatory body in the UK that oversees IVF and related research.

Among many of its other tasks, the HFEA has been trying to make sure that culture medium used for in vitro maturation and preimplantation cultures is safe and optimized for use with human embryos. For example, in 2001, they surveyed IVF clinics about the culture medium they use, mainly to investigate any potential infection risks.

In December 2011, the authority’s Scientific and Clinical Advances Advisory Committee (SCAAC) published a report outlining the current knowledge about the effect of culture media components on embryo viability and development. It’s available on their website.

This most recent report notes that “Although generally considered to be safe based on past and current experience, there are still uncertainties about the long term effects of the culture media used for in vitro fertilisation, and questions remain about the effect of varying components in media. Varying concentrations of components such as growth factors, amino acids, energy substrates, and antibiotics may potentially impact on early embryo development and may have longer term health implications.”

The main conclusion of the report is that, at the moment, the research is inconclusive. They recommend more research into effects of growth medium on early and late development, as well as further investigation into long-term health effects.

The report summarizes some of the studies that have been done to date, which suggest that it’s crucial to keep an eye on what manufacturers add to the media. In one example, an article by Harper et al in Human Reproduction points out that culture medium used in human embryo development was designed based on studies in animal development. It’s not optimized for human development, because for many conditions we simply don’t know what the optimal human condition is. What we do know is that the conditions used in embryo cultures may have long-term effects. They cite a study by Dumoulin and colleagues that showed that different preimplantation media result in differences in birth weight after successful IVF pregnancies. Since birth weight may be associated with disease risk later in life, this research in particular highlights the importance of studying the effect of the various components of culture media.

Studies such as these suggest that any changes in culture media for human embryos need to be assessed very closely. However, the manufacturers of culture media are not obliged to disclose the exact composition of the media they sell. That makes it very difficult to understand the effect of various factors on development.

In an ongoing effort, the HFEA is collecting reports and studies such as these, and is gathering feedback from IVF clinics and researchers. They hope to find out what can be done to ensure that the effects of the culture media used in this field are well-regulated and understood, and plan to communicate their findings to the MHRA.

These studies may take years to complete, especially those assessing long-term effects, but a stronger scientific understanding of the role of in vitro culture medium in human development will eventually translate to a higher success rate and lower risk of IVF procedures.

Harper J, Magli MC, Lundin K, Barratt CL, & Brison D (2012). When and how should new technology be introduced into the IVF laboratory? Human reproduction (Oxford, England), 27 (2), 303-13 PMID: 22166806

Dumoulin JC, Land JA, Van Montfoort AP, Nelissen EC, Coonen E, Derhaag JG, Schreurs IL, Dunselman GA, Kester AD, Geraedts JP, & Evers JL (2010). Effect of in vitro culture of human embryos on birthweight of newborns. Human reproduction (Oxford, England), 25 (3), 605-12 PMID: 20085915

(3 votes)

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We are seeking to appoint post-doctoral researchers to study microenvironmental regulation of stem cells in mammalian skin.

The aim of our group is to gain a better understanding of the mechanisms underlying the ways in which tissue microenvironments are regionally specialized, and how the specialized microenvironments instruct cellar behaviour and communication, and organ formation. We are particularly interested in the role of the extracellular matrix (ECM) in the formation of the stem cell microenvironment or niche.
http://www.cdb.riken.jp/en/02_research/0202_creative30.html

A recent study by our team has shown that the molecular composition of the basement membrane in mouse hair follicle stem cell niche, the bulge, is highly specialized. One stem cell-derived component, nephronectin, is important for the development and positioning of the bulge-residing arrector pili muscles, which, among other functions, are responsible for goosebumps (Fujiwara et al. 2011. Cell 144, 577-589). This was the first report to show that stem cells regulate the fate and positioning of surrounding niche cells through the specialization of the basement membrane.

To gain further insight into fundamental aspects of the microenvironmental regulation of stem cells, we use mouse skin as a model and seek to better understand 1) the molecular landscape of basement membrane specialization in the stem cell niche, 2) mechanisms by which the basement membrane in the stem cell niche is regionally specialized, and 3) how the specialized basement membrane controls stem cell niche formation, stem cell behaviour and the conversation between stem cells and their neighbouring cells.

Successful candidates will receive an excellent salary commensurate with qualifications and experience. Our Centre, the Center for Developmental Biology (CDB), is a world-leading research institute in the fields of developmental and regenerative biology and has state-of-the-art research facilities. The Centre provides a truly international, collegial, and supportive environment for its nearly thirty laboratories, and the freedom and resources to pursue their research toward deeper understanding of developmental biology. All necessary communications can be conducted in English, and support services are available for non-Japanese-speakers.

Please contact Hironobu Fujiwara, PhD (hironobu.fujiwara@cancer.org.uk) for further information.

To apply, please send 1) a cover letter, 2) a CV with publication list, 3) names and contact details of two referees, 4) a brief summary of research achievements and future research interests (two page maximum) to Hironobu Fujiwara (hironobu.fujiwara@cancer.org.uk) via email. Please send the application as a single PDF file. This call will be closed when the positions are filled.

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What was new on the Node this past month?

“Developments in development” essay competition
Congratulations to Joanna Asprer and Máté Varga, whose essays were selected by the judges in our recent essay competition. Read both essays on the Node, and then vote for your favourite. The final winning essay will be published in Development.
–An Excitingly Predictable ‘Omic Future – Joanna Asprer
–There’ll Be dragons? – The coming era of artificially altered development – Máté Varga

Research
Karuna Sampath’s lab has a paper out in the most recent issue of Development, and shares the backstory on the Node. They discovered that knockdown of squint affected dorsal axis formation in zebrafish, but their initial findings didn’t seem to match the prevailing knowledge about the role of squint, or even the stages at which it was expressed. “It seemed as though we were ascribing a function to an RNA no one else could see – it was The Emperor’s New Clothes of the zebrafish embryo!” writes Karuna, while graduate student Shimin Lim gives the student perspective in a second post: “… very soon, my project was shrouded with controversy. Colleagues in the field had challenged Aniket’s findings, and I was following up on his work.”
In their Node posts and the paper, Karuna and Shimin describe how they solved the puzzle to match both their new observations and the existing literature in the field. It’s a story of science in action!

Conferences and courses
This month we heard from Gi Fay Mok, who attended the 12th International Conference on Limb Development and Regeneration in Mont-Tremblant, Canada. It sounds like it was a great conference!

A few weeks later, developmental biologists gathered in Canada again, for the annual SDB meeting, which was held in Montreal this year. Patricia Gongal summarized the meeting’s highlights and we collected tweets from the conference on Storify.

We also received more updates from the Woods Hole Embryology Course. Priti wrote about making connections and meeting people while Manuela reflected on the course after it finished.

Meanwhile, there is currently an open voting round to choose a Development cover from these images from last year’s embryology course. Which image is your favourite?

Also on the Node
– Clare Cox reviewed the film “Stem Cell Revolutions”. This is an educational documentary about advances in stem cell research, produced by Amy Hardie and Clare Blackburn. Read Claire’s review, and then watch the documentary!

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Here I share the background story on my graduate work that was recently published in Development, “Dorsal activity of maternal squint is mediated by a non-coding function of the RNA”: I first joined Karuna Sampath’s group at Temasek Life Sciences Laboratory in 2005 during my undergraduate days. It was an exciting year in the lab because Aniket Gore, a former student, had published an interesting story on the identification of asymmetrically localized maternal sqt RNA as the earliest dorsal marker in zebrafish. I found the role of sqt in dorsal specification immensely intriguing, therefore I started working on it as a graduate student.

However, very soon, my project was shrouded with controversy.  Colleagues in the field had challenged Aniket’s findings, and I was following up on his work.  The first hurdle I encountered was to convince Karuna (yes, you are reading this correctly) that sqt RNA has a function independent of Sqt protein.  The sqt insertion mutants were thought to be nulls. I was stunned when I saw that sqt^cz35 RNA elicited transient dorsal expansion in wild-type embryos. I remember showing Karuna the sqt^cz35 RNA-injected embryos stained for goosecoid and chordin, excited that I had identified a non-coding function. But the first thing she asked was, “Are you sure your sqt^cz35 prep is not contaminated with traces of wild-type sqt?”  I re-transformed pCS2+sqt^cz35, made countless mini-preps, sequenced them, synthesized capped RNA, injected and assayed each for dorsal expansion in the embryos to show there was no contamination.  In any case, wild-type sqt RNA did not show transient dorsal expansion, so I knew this was something unusual.

The series of experiments that followed was rather straight-forward. I tested different forms of non-coding sqt RNA (sqt^mut RNAs) and found that even heterologous sequences fused with sqt UTR could induce dorsal, and this does not require Oep-dependent Nodal signaling. Since the dorsal activity was in the sqt UTR, Steve Cohen and Mark Featherstone, who serve on my thesis committee, asked if microRNAs had any part to play. MZdicer embryos were still able to respond to the sqt^mut RNAs, ruling out miRNAs in this process.  This was a new function I had found in the sqt 3’UTR.

No words can describe my excitement at being able to share my findings with the readers of Development. Although the journey was quite arduous, I am pleased that my work has addressed some of the issues surrounding the debate regarding maternal sqt and DV axis formation, and that I have found an interesting area of biology that can be further mined upon.  My focus is now to understand the precise mechanism by which maternal non-coding sqt functions in dorsal axis formation.

Shimin Lim

Shimin Lim, Pooja Kumari, Patrick Gilligan, Helen Ngoc Bao Quach, Sinnakaruppan Mathavan, & Karuna Sampath (2012). Dorsal activity of maternal squint is mediated by a non-coding function of the RNA Development, 139 (16) : 10.1242/dev.077081

(13 votes)

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You’ve probably already read Patricia Gongal’s summary of the recent SDB meeting (and if not, go do that now!) but many others have also reported from Montreal through Twitter. We’ve collected some of the tweets from the conference below:

(more…)

(1 votes)

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Here is the backdrop for our recent paper in Development, “Dorsal activity of maternal squint is mediated by a non-coding function of the RNA”:  This work follows up a previous publication from my laboratory where we showed that knock-down of maternal squint (sqt) or ablation of sqt-containing cells led to loss of dorsal structures in zebrafish (Gore et al., 2005).  It was a surprising finding, and did not fit with the prevailing view of sqt function or the sqt and one-eyed pinhead (oep) mutant phenotypes.  Furthermore, authors of two papers (Pei et al., 2007 and Bennett et al., 2007) reported that they could not detect sqt RNA in early embryos.  It seemed as though we were ascribing a function to an RNA no one else could see – it was The Emperor’s New Clothes of the zebafish embryo!  Our view was that the sqt insertion mutants used by the other groups were not RNA nulls.  We also couldn’t understand why they did not detect maternal sqt RNA in the mutants when it was readily visible to us (Gore et al., 2007).  There were many unresolved questions, and I was encouraged (by my institute’s scientific advisory board and other colleagues) to get to the bottom of the debated issues.  So we set out to understand the basis for the differences in the findings from the various groups, and the mechanism of maternal sqt in dorsal axis formation.

As it turns out, the differences in the datasets between the groups were very informative, and uncovered really interesting biology, which we report in Shimin Lim’s paper.  We think the initial lack of processing of maternal sqt RNA (which explains some of the differences in the RNA expression data), is likely important for other aspects of sqt function such as localization and translation.  We were very surprised to find that over-expression of mutant sqt RNA (that is incapable of making a functional signaling protein) or the sqt 3’UTR fused to any reporter could expand the expression domain of dorsal genes.  Shimin had uncovered a new non-coding activity of the RNA.

However, the dorsal expansion disappeared later in gastrulation, which was really perplexing, difficult to track, and it took us a while to figure out what was happening.  My colleague, Steve Cohen, then pointed out to us the example of bicoid transgenes that transiently expand the head anlagen in Drosophila embryos.  Shimin’s finding that the requirement for oep is not absolute for dorsal expansion by non-coding sqt RNA was another piece of the puzzle that fit well, given the differences in the phenotypes of the signaling mutants and our morphants.  That the sqt morpholinos block both RNA localization as well as protein synthesis, and lead to loss of nuclear beta-catenin helped us make sense of our previous results showing loss of dorsal structures in sqt morphants; the maternal sqt puzzle seemed to be coming together.

But there are still several pieces missing in the puzzle.  The findings we report in Shimin’s paper explain many, but not all the differences in the RNA expression data between the groups.  Alleles that are RNA nulls are required to unequivocally demonstrate the function(s) of maternal sqt.  And the biggest piece of the puzzle remains to be found – how exactly does sqt RNA function in dorsal axis formation?  So there is more interesting biology to be uncovered.

Despite the shadow of skepticism surrounding our sqt work, and the frustration of trying to make sense of data that simply doesn’t fit with prevailing models, my enthusiastic young colleagues who did this work remain excited about uncovering the unexpected, discovering novel findings, and I hope they will see the satisfaction that comes from solving a challenging puzzle.

Shimin Lim, Pooja Kumari, Patrick Gilligan, Helen Ngoc Bao Quach, Sinnakaruppan Mathavan, & Karuna Sampath (2012). Dorsal activity of maternal squint is mediated by a non-coding function of the RNA Development, 139 (16) : 10.1242/dev.077081

Gore, A., Maegawa, S., Cheong,A., Gilligan,P., Weinberg,E.  and Sampath, K. The zebrafish dorsal axis is apparent by the four-cell stage. Nature 438: 1030-1035 (2005).

Bennett, J. T., Stickney, H. L., Choi, W. Y., Ciruna, B., Talbot, W. S. and Schier, A. F. Maternal nodal and zebrafish embryogenesis.  Nature 450(7167): E1-2 (2007).

Pei, W., Williams, P. H., Clark, M. D., Stemple, D. L. and Feldman, B. Environmental and genetic modifiers of squint penetrance during zebrafish embryogenesis.  Dev Biol 308(2): 368-78 (2007).

Gore A.V., Cheong A., Gilligan P.C., Sampath K.   Gore et al. reply.  Nature 450(7167):E2-E4 (2007).

Karuna Sampath

(14 votes)

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Do you need to learn a new technique?  Are you planning a collaborative visit?  If so please have a look at our Travelling Fellowships – http://www.biologists.com/fellowships.html.  The next deadline is the 31st August.

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Open University – Stazione Zoologica Anton Dohrn International Ph.D. Program: 7 Positions on offer, 2012 – 2013

7 Ph.D. fellowships are available to carry out interdisciplinary training in Biological Sciences at the Stazione Zoologica ‘Anton Dohrn’ Naples Italy.

Applications are invited from suitably qualified postgraduate candidates (see details)

The closing date for applications is August 31st 2012

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Here are the research highlights from the current issue of Development:

Less air, more muscle repair

Oxygen levels in stem cell niches are often lower than those in surrounding tissues, and hypoxia is known to regulate the self-renewal and proliferation of multiple stem cell types. Now, Shihuan Kuang and co-workers report that hypoxia regulates the ‘stemness’ of satellite cells (muscle stem cells) (see p. 2857). Upon muscle injury, quiescent satellite cells, which lie below the basal lamina of muscle fibres, are activated to proliferate, differentiate and fuse into myofibres, which regenerate the damaged muscle. The researchers show that hypoxic conditions favour satellite cell quiescence by promoting self-renewal divisions without affecting the overall proliferation of primary myoblasts. Hypoxia, they report, activates Notch signalling, which suppresses the microRNAs miR-1 and miR-206, thereby upregulating Pax7, a key regulator of satellite cell self-renewal. Moreover, hypoxic conditioning enhances the efficiency of myoblast transplantation and the self-renewal of implanted cells in vivo. These results identify oxygen level as a physiological regulator of satellite cell activity and suggest that manipulation of oxygen exposure could improve muscle regeneration after injury.

Embryonic stem cells achieve XEN state

Early mammalian embryogenesis is characterised by a gradual restriction in the developmental potential of embryonic cells. By the blastocyst stage, embryonic and extra-embryonic cells have diverged in their fate and function. However, on p. 2866, Kathy Niakan and co-workers describe a robust method for converting mouse embryonic stem cells (mESCs) derived from the inner cell mass of the blastocyst into extra-embryonic endoderm (XEN) stem cells. Previous studies have derived XEN from mESCs by overexpression of Gata4 or Gata6, but here the researchers achieve the same outcome by adding growth factors to a standard culture medium. They confirm that downregulation of the pluripotency transcription factor Nanog and expression of primitive endoderm-associated genes are necessary for XEN cell derivation, and they show that FGF signalling is required to exit mESC self-renewal but not for XEN cell maintenance. Intriguingly, the researchers also show that not all pluripotent stem cells respond equivalently to differentiation-inducing signals. Together, these results suggest that mESCs have a broader differentiation capacity than previously appreciated.

Gut feelings about mesothelial development

The vertebrate body cavity (coelom) and its internal organs are lined with a simple squamous epithelium called the mesothelium. Mesothelia, which generate the vasculature of coelomic organs, are relatively quiescent in healthy adults but are implicated in pathogenic conditions such as peritoneal sclerosis. The mesothelium of the heart develops from the proepicardium, a localized exogenous population of cells that migrates to and envelops the myocardium; but what are the developmental origins of other mesothelia? Here (p. 2926), David Bader and colleagues identify a novel mechanism for the generation of mesothelia by investigating intestinal development in avian embryos. Using long-term lineage tracing, they show that mesothelial progenitors of the intestine are intrinsic to the gut tube anlage. Moreover, using a new chick-quail chimera model of gut morphogenesis, they show that mesothelial progenitors are distributed throughout the gut primordium and are not derived from a localized, exogenous proepicardium-like source of cells. Thus, cardiac and intestinal mesothelia have strikingly different developmental histories, despite their similarities in ultrastructure and developmental potential.

Neural lineage pools: REST assured

During nervous system development, coordinated activation and suppression of transcriptional activators and repressors regulates a network of lineage-specific genes that maintains the neural stem/progenitor (NS/P) pool and ensures the orderly acquisition of neural cell fates. Here (p. 2878), Nurit Ballas and colleagues characterise the role played by REST (RE1 silencing transcription factor, a known master repressor of neuronal genes) during neural development. By analyzing the function of REST at different stages of mouse neural development, the researchers show that, although it represses neuronal genes in embryonic stem (ES) cells, it is not required for the maintenance of ES pluripotency or for the conversion of ES cells to neurogenic or gliogenic NS/P cells. REST is required, however, for NS/P cell self-renewal. Moreover, it maintains the proper ratio of neurons and glia during differentiation and regulates genes involved in both neuronal and oligodendrocyte specification and maturation. Thus, REST plays a central role during neural development by regulating the pool size of different neural lineages.

The long and short of developmental cell migration

Long-distance cell migration is an important feature of embryonic development, but the mechanisms that underlie these migratory phenomena are unclear. Now, by combining computational modelling and experimental analysis of neural crest (NC) cell migration in chick embryos, Paul Kulesa and co-workers provide a new mechanistic explanation for long-distance cell migration (see p. 2935). The researchers begin by hypothesizing that chemotaxis drives cells to move in a directional manner towards a distant target. This simple model is insufficient, however, to explain their new experimental data. Instead, model simulations that include tissue growth predict that directed NC cell migration requires leading cells to respond to long-range guidance signals and trailing cells to respond to short-range cues. The researchers subsequently identify two NC cell subpopulations with distinct gene expression profiles and cellular orientations within migratory streams, and experimentally confirm model predictions of the effects of exchanging leading and trailing cell positions. Thus, they conclude, a two-component mechanism that includes chemotaxis and cell-cell contact drives long-distance cell migration.

miR-7 fine-tunes pancreatic development

MicroRNAs (short non-coding RNAs that post-transcriptionally silence target mRNAs) are essential for early pancreas development. But how specific microRNAs are intertwined into the transcriptional network that controls endocrine differentiation is poorly understood. On p. 3021, Eran Hornstein and colleagues investigate the involvement of miR-7, which is highly expressed in the endocrine pancreas of several vertebrates, during mouse endocrine cell differentiation. They show that miR-7 is expressed in mouse endocrine precursors and mature endocrine cells, and that the transcription factor Pax6, which is required for endocrine cell differentiation, is an important miR-7 target. The overexpression of miR-7 in developing pancreas explants or in transgenic mice downregulates Pax6 and inhibits α- and β-cell differentiation, they report, whereas miR-7 knockdown has opposite effects. Notably, Pax6 downregulation reverses the effects of miR-7 knockdown on insulin promoter activity. These findings, which suggest that miR-7 and Pax6 are wired into a transcriptional network that ensures the precise control of endocrine cell differentiation, may help in the development of cell-based therapies for diabetes.

Plus…

Vascular instruction of pancreas development

Recent studies suggest that blood vessels provide organs with non-nutritional signals that control their development and homeostasis.  In this issue, Ondine Cleaver and Yuval Dor review the contribution of the vasculature to developing tissues, with a focus on the pancreas.

See the Review article on p. 2833

Satellite cells are essential for skeletal muscle regeneration: the cell on the edge returns centre stage

In this issue, Frederic Relaix and Peter Zammit review recent recombination-based studies that have furthered our understanding of satellite cells – muscle stem cells. The clear consensus is that skeletal muscle does not regenerate without satellite cells, confirming their pivotal and non-redundant role.

See the Review article on p. 2845

(1 votes)

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