SEB Plant Symposium on New Breeding Technologies

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Published on: July 30, 2017

Thanks to Iulia German from the University of York and the Warwick Integrative Synthetic Biology Centre for providing this outstanding review of the SEB plant section symposium.


At the beginning of July, Gothenburg was host to a fascinating discussion on New Breeding Technologies (NBTs), attended by scientists from 17 countries. This covered the applications of the technologies themselves, their legal status in the EU, and the rest of Europe, and the US, and Australia and New Zealand, the importance of getting the general public involved early in discussions about NBTs, and whether CRISPR-modified cabbage really does taste better than cabbage modified only through conventional breeding.

Image from Molecular Plant Pathology October 2016 issue. Pyott et al. engineered Turnip Mosaic Virus-resistant Arabidopsis and created letters in trays by infecting wildtype and CRISPR-Cas9-engineered mutants with the virus.

This Society for Experimental Biology plant section symposium was very timely given the increasing trendiness of CRISPR-Cas9 for editing plant genomes, the legal limbo of the technology in EU courts, and the lack of public awareness of CRISPR and what it can do.

From the Web of Science. While not as extreme as the overall trend, there is still a growth in published papers using CRISPR-Cas9.

While the efficiency of CRISPR-Cas9 in plants seems to be limited in comparison to animal cells, this appears to come with the unexpected benefit of fewer off-target effects. In fact the frequency of off-target mutations is even lower than that observed during chemical/physical mutagenesis. This does make for a hard time screening thousands of plants but that is worth it for people like Mariette Andersson, who is looking to bring a new variety of potato to market years faster than would be possible using conventional technologies, and Jean-Denis Faure who is generating Camelina sativa lines with different amounts of oleic acid content by targeting different combinations of FAD2 alleles in its hexaploid genome. Not to mention that it’s possible to screen for plants that have desired mutation but are CRISPR-cas9 transgene free (like in Attila Molnar’s work to create potyvirus-resistant Arabidopsis).

The variety and flexibility of CRISPR gene editing is increasing, with the possibility for nicks, double-stranded breaks, Cas9-nuclease fusions, dead Cas9-repressor/activator fusions, polycistronic systems, and a myriad of promoters and terminators to choose from.


The second point of discussion, the legal debate surrounding CRISPR is nicely summarised in this SEB article. It started with a history lesson on GMO regulation in the 70s after the ability to transfer genes from any organism to any other organism sparked safety discussions and a moratorium as laid out by the 1974 Paul Berg letter. The US was then first to develop regulations against these novel organisms. As a side note, the reason “novel” organisms are regulated rather than “risky” organisms is mainly because in a legal context it was easier to define something novel than something risky – however this has encouraged the train of thought that “novel” organisms are somehow more dangerous than “natural” organisms (another thing that emerged from this NBT meeting is how much scientists hate misuse of the word “natural”). All countries seem to have some legal variation on this “novel organism” theme, with some more lenient than others – Canada for example does not regulate plants with novel traits that do not possess an environment risk.

However, this process-based regulation is very outdated and not reflective of the current situation and advances in biotechnology. It would be better to include product-based rather than process-based rules ie plants should not be regulated based solely on the methods used to produce them (process-based) but based on the risks presented by the GM plant products.

There is no strict regulation if the technique involves mutagenesis, transfer of genes from sexually compatible organisms or non-inheritable changes. As CRISPR-edited plants fall into the “mutagenesis” category, we agreed that the same regulation should apply.
The crux of the debate surrounding NBTs: are they more like conventional breeding techniques or gene modification techniques? Image from: [1]


The world (other than Sweden and Denmark, which have declared that CRISPR-edited organisms are not GM) is looking to the EU to lead the way in making regulations, but the EU has been stalling.

As to how to present CRISPR to the public, the situation boils down to: people make decisions based on their principles and beliefs when presented with very complex information, so if scientists unite behind one success story for CRISPR that will appeal to people’s imagination and values, then the battle is half won. It’s all about planting the flag early. As Craig Cormick learned from surveying Australians, the bulk of the population is neither highly opposed to or greatly in favour of new technologies (they are penguins – image below). Given convincing arguments, they are likely to move to either side. In the case of NBTs, the public are not likely to be in favour of them without being given a context: it’s the difference between asking “What do you think of CRISPR – should it be used?” and “What do you think of using CRISPR to make X product?”

Image from Craig Cormick

The public’s attitudes towards new technologies: at polar ends of the spectrum are small groups of “polar bears” which are either highly opposed to or highly in favour of the technology. Most of the population falls into a cautious group in the middle who decides based on the risks presented and the application of the technology.


One of the organisers, Swedish Stefan Jansson, has been doing a fair bit to normalise the use of NBTs in food by growing his own CRISPR-modified cabbage and cooking up a feast with it. The recipe for tagliatelle with CRISPRy fried vegetables, along with Stefan’s blog detailing how he grew the cabbage, fought moths and ate the first CRISPR meal ever with reporter Gustaf Klarin, can be found here. He’s gone on tour with his cabbage to countries like Norway in order to start a discussion and hopefully persuade the legal authorities to pass the same laws as Sweden regarding the classification of NBTs. He is also adamant about refusing to say what has been modified in the cabbage – good luck finding a targeted point mutation amongst all the other spontaneous mutations that arise in the genome! What we do know for sure is that CRISPRy cabbage tastes delicious – especially when prepared by the culinary geniuses at Sjömagasinet.

All in all, the SEB New Breeding Technologies was a fantastic meeting, with great food and company. I’m always looking forward to the GARNet plant gene editing workshop at the University of Bristol on March 26-27th 2018!

GCRF: the story so far…

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Published on: July 27, 2017

Unless you’ve been living under a rock then you’ll have heard of the Global Challenges Research Fund (GCRF). This is a RCUK-led program that is tasked with distributing £1.5 billion in Overseas Development Funding on behalf of the UK Government. This was released following the 2015 spending review and has to be spent over a five year period. When the scheme was announced there was plenty of interest as to how the finances would be allocated between individual research councils and also as to the type of grant that would be available. Over the past year there has been a steady stream of RCUK funding calls across topic areas. The one constant is that the proposed research must demonstrably benefit life in countries on the Overseas Development Aid (ODA) list (PDF).

In the early stages of the GCRF it was unclear what level of connection was needed between UK researchers and colleagues based in ODA countries. As the results of the first funding calls have been announced it appears that already established collaborations that have a track record have been favoured. For the Arabidopsis community it is clear that proof-of-concept research that might have a future benefit in an ODA country is unlikely to be directly funded. Therefore UK researchers undertaking basic research will have to rethink their focus in order to qualify for this type of funding.

To date there have been a number of funding calls in which plant scientists might feel they have opportunities for success. Last years initial BBSRC call entitled ‘GCRF Foundation Awards for Global Agriculture and Food Systems Research’  was designed for responsive mode type grants (£600K max) and was hugely oversubscribed. Finally about 20% of ‘Expression of interest’ applications were invited to provide full submissions (around 100 applications). Eventually 35 projects were funded that comprise an exciting mix of research areas that included work on a variety of species (rice, wheat, cassava, potato, chickpea, native orphan crops) and technologies (genomics, phenotyping, smart breeding of new varieties, improving pipelines to yield additional products). The full list is below or can be downloaded from the BBSRC website (PDF).


Other funding opportunities that have likely had multiple applications from plant scientists include a joint-call with USDA/NIH as part of the Ecology and Evolution of Infectious Diseases programme as well as a joint BBSRC-MRC call entitled Networks in Vector Borne Disease (VBD) Research. In addition applications to the annual Bioinformatics and Biological Resources Fund (BBR) can be directed into a separate sub-call if they have a GCRF-component. The results of each of these calls have not yet been announced but there is currently an active BBR call with a deadline at the end of September 2017.

Currently RCUK are inviting applications for larger scale projects in order to establish Interdisciplinary Research Hubs that will tackle an ‘intractable development challenge‘ in ODA countries. With maximum grants of £20million available this call represents a significant investment of up to £300M. In these proposals the Research Hub will be situated in the UK but with possible partners in both developed countries (for which an equal investment is expected) and in countries on the ODA-list (for which 100% of direct costs can be met). The deadline for the Intention to Submit is the 29th September with the full proposal due in early November.


The GCRF has received some excellent publicity over the past couple of weeks with the announcement of the 37 successful proposals that received funding through the GCRF Collective Fund, which provide up to £8million of funding for each grant. RCUK produced a smart brochure outlining each of these projects, which included the TIGR2ESS program that is a UK-India collaboration aiming to help facilitate a ‘Second Green Revolution’ by using smart technology to support female smallholder farmers to maintain crop yields in the face of increased urbanisation. This is led by Professor Howard Griffiths at the University of Cambridge. Elsewhere Professor Federica de Palma at the Earlham Institute  is leading a project that aims to use NGS to scrutinise the untapped potential of the biodiversity in Colombia as well as aiming to characterise the genetics of native agricultural germplasm in order to make farming more efficient.

From https://twitter.com/TIGR2ESS/status/888354428855742464/photo/1

The TIGR2ESS team!


Given the large amount of money available from the GCRF there will be many other funding calls over the next few years, keep your eye out for something relevant for your research…..and collaborate!

 

 

 

Anne Osterrieder talks to GARNet

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Published on: July 21, 2017

Anne Osterrieder speaks to GARNet about a recent paper published in JXB entitled ‘Stacks off tracks: a role for the golgin AtCASP in plant endoplasmic reticulum-Golgi apparatus tethering‘. Please listen here, on the GARNet YouTube channel or subscribe to GARNet on iTunes.

Arabidopsis Research Roundup: July 18th

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Published on: July 18, 2017

This weeks Arabidopsis Research Roundup includes four studies from around the UK. Firstly is a systems-level study of the drought response that includes Alessandra Devoto from RHUL as a co-author. Secondly Anne Osbourn’s group at the JIC investigates sesterterpenoid biosynthesis across plant species. Thirdly Paul Jarvis from Oxford University adds to this groups portfolio of research on the mechanisms that control thylakoid import. Finally Patrick Gallois (University of Manchester) provides further insight into the regulation of programmed cell death.


Kim JM, To TK et al (2017) Acetate-mediated novel survival strategy against drought in plants Nature Plants http:/​/​dx.​doi.​org/10.1038/nplants.2017.97

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Alessandra Devoto (Royal Holloway) is a co-author of this study led by Jong-Myong Kim, Mototaki Seki (RIKEN, Yokohama) and Taiko Kim Ko (University of Toyko) that investigates the system-wide alterations that plants make in response to drought stress. They demonstrate that the histone deacetylase HDA6 is the primary regulator of an epigenetic switch that leads to a metabolic flux conversion from glycolysis into acetate synthesis. This in turn stimulates the jasmonate signaling pathway that causes increased drought tolerance. Importantly the authors show that this critical survival response is evolutionarily conserved through monocots and dicots.


Huang AC, Kautsar SA, Hong YJ, Medema MH, Bond AD, Tantillo DJ, Osbourn A (2017) Unearthing a sesterterpene biosynthetic repertoire in the Brassicaceae through genome mining reveals convergent evolution. PNAS http:/​/​dx.​doi.​org/10.1073/pnas.1705567114

Open Access

Anne Osbourn (JIC) leads this study in which her group works with collaborators from Cambridge, Wageningen and UC Davis to perform a cross-species genome-wide analysis of sesterterpenoid biosynthesis. They use a novel search algorithm to identify paired enzymatic components that comprise sesterterpene synthases (STS). These enzymes were transiently overexpressed in tobacco leaves, resulting in the formation of fungal-like sesterterpenes, suggestive of convergent evolution of plant and fungal STS. This study illuminates possible future strategies for the beneficial use of sesterterpenes through metabolic and protein engineering


Bédard J, Trösch R, Wu F, Ling Q, Flores-Pérez Ú, Töpel M, Nawaz F, Jarvis P (2017) New Suppressors of the Chloroplast Protein Import Mutant tic40 Reveal a Genetic Link between Protein Import and Thylakoid Biogenesis. Plant Cell. http:/​/​dx.​doi.​org/10.1105/tpc.16.00962 Open Access

Paul Jarvis (Oxford University) leads this global collaboration that focuses on the chloroplast protein import protein Tic40. A suppressor screen identified two novel regulators of Tic40, ALB4 and STIC2 that they postulate are involved in the thylakoid targeting of a subset of proteins and that their influence becomes more important in the absence of Tic40.


Cai YM, Yu J, Ge Y, Mironov A, Gallois P (2017) Two proteases with caspase-3-like activity, cathepsin B and proteasome, antagonistically control ER-stress-induced programmed cell death in Arabidopsis. New Phytol.

http:/​/​dx.​doi.​org/10.1111/nph.14676 Open Access

Patrick Gallois is the corresponding author on this study that originates at the University of Manchester. They attempt to establish a role for cathepsin B and proteasome subunit PBA1 in the control of programmed cell death (PCD) and whether their functions interest with those of caspase-3. They reveal a complex system of regulation where aspects of PCD are differentially impacted by each of these proteins. They propose the role of cathepsin B might occur late in PCD following tonoplast rupture.

Arabidopsis Research Roundup: July 3rd.

The Arabidopsis Research Roundup returns this week with selection of publications from institutions across the UK. Firstly George Bassel (Birmingham) leads a study that investigates the integration of inductive signals in the embryonic root. Secondly a group from the Oxford Brookes plant science group look into the literal linkages between the golgi apparatus and ER. Thirdly John Christie (Glasgow) and co-workers define a new variant of the phototropin receptor. Next Caroline Dean and Martin Howard (John Innes Centre) collaborate on work that defines the relationship between FLC, COOLAIR and cell size. The fifth paper is led by members of SLCU and investigates the regulatory influence of the Evening Complex of the circadian clock. The penultimate paper features work from Alison Smith’s group at the JIC that looks at dynamics of starch accumulation and degradation. Lastly is research from NIAB that defines the pathogeniticity of novel UK isolates of the fungal pathogen Verticillium longisporum.


Topham AT, Taylor RE, Yan D, Nambara E, Johnston IG, Bassel GW (2017) Temperature variability is integrated by a spatially embedded decision-making center to break dormancy in Arabidopsis seeds. PNAS

http:/​/​dx.​doi.​org/10.1073/pnas.1704745114

Open Access

George Bassel (University of Birmingham) leads this study that identifies a decision making centre in the embryonic root that is defined by the intimate interaction between the hormones abscisic acid (ABA) and gibberellin (GA). The activity of this ‘decision centre’ is linked to both hormone transport and changes in temperature, the overall output of which is the decision to promotes seed germination or to delay until more favourable environmental conditions.

George discusses this paper on the GARNet YouTube channel.



Osterrieder A, Sparkes IA, Botchway SW, Ward A, Ketelaar T, de Ruijter N, Hawes C (2017) Stacks off tracks: a role for the golgin AtCASP in plant endoplasmic reticulum-Golgi apparatus tethering. J Exp Bot. http:/​/​dx.​doi.​org/10.1093/jxb/erx167

Open Access

Anne Osterrieder and Chris Hawes (Oxford Brookes University) continue their work that looks at  the cellular dynamics of the golgi apparatus with this study that identifies the AtCASP protein as a important component that tethers the golgi to the ER. They use live-cell imaging to visualise golgi formation in cells that have different levels of AtCASP, allowing the authors to confirm that ER-golgi tethering is disrupted without the activity of this protein.


Petersen J, Inoue SI, Kelly SM, Sullivan S, Kinoshita T, Christie JM (2017) Functional Characterization of a Constitutively Active Kinase Variant of Arabidopsis Phototropin 1

J Biol Chem. http:/​/​dx.​doi.​org/10.1074/jbc.M117.799643

Open Access

John Christie (University of Glasgow) collaborates with Japanese colleagues to identify a novel variant of the phototropin receptor. Study of this variant allows a greater understanding regarding the mode of action of this protein under different light conditions, as controlled by phosphorylation.


Ietswaart R, Rosa S, Wu Z, Dean C, Howard M (2017) Cell-Size-Dependent Transcription of FLC and Its Antisense Long Non-coding RNA COOLAIR Explain Cell-to-Cell Expression Variation. Cell Syst. http:/​/​dx.​doi.​org/10.1016/j.cels.2017.05.010

Open Access

Martin Howard and Caroline Dean (John Innes Centre) again collaborate on research that analyses the mode of regulation of FLC. They dissect RNA dynamics of FLC expression by single molecule in situ RNA fluorescence, showing that this is dependent on the presence of the antisense COOLAIR regulatory DNA. In the absence of COOLAIR they show FLC expression has a linear relationship with cell size but in the presence of the antisense transcript, FLC expression decreases with cell size. Overall they demonstrate FLC expression is tightly dependent on the presence of the antisense COOLAIR transcript.


Ezer D, Jung JH, Lan H, Biswas S, Gregoire L, Box MS, Charoensawan V,, Cortijo S, Lai X,, Stöckle D, Zubieta C, Jaeger KE, Wigge PA (2017) The evening complex coordinates environmental and endogenous signals in Arabidopsis. Nat Plants.

http:/​/​dx.​doi.​org/10.1038/nplants.2017.87

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Phil Wigge and Katja Jaeger (SLCU) lead this study that investigates how the evening complex of the circadian clock coordinates the expression of numerous important growth regulators. This genome wide regulation is determined by temperature and concides with the binding of phytochrome B, which provides a cellularly mechanism of this level of environmental control.


Fernandez O, Ishihara H, George GM, Mengin V, Flis A, Sumner D, Arrivault S, Feil R, Lunn JE, Zeeman SC, Smith AM, Stitt M (2017) Foliar starch turnover occurs in long days and in falling light at the end of the day. Plant Physiol. http:/​/​dx.​doi.​org/10.1104/pp.17.00601

Open Access

On this paper Alison Smith (John Innes Centre) is a co-corresponding author together with Mark Stitt from the Max Planck Institute for Molecular Plant Physiology in Potsdam. They continue their work to investigate the dynamics of starch metabolism in Arabidopsis leaves. Broadly they show the rate of starch accumulation corresponds to the photosynthetic rate whilst degradation is linked to correct functioning of the circadian clock. They investigate this process in more detail by determining how the rate of starch degradation alters dependent on the time after dawn.


Depotter J, Rodriguez-Moreno L, Thomma BP, Wood T (2017) The emerging British Verticillium longisporum population consists of aggressive Brassica pathogens. Phytopathology http:/​/​dx.​doi.​org/10.1094/PHYTO-05-17-0184-R

Tom Wood (NIAB) is the corresponding author of this study that characterises four new UK isolates of the fungal pathogen Verticillium longisporum. The pathogenticity of V.longisporum was tested on Arabidopsis alongside three other Brassica crops. They demonstrate that the UK isolates were unusually aggressive yet this was not consistent across all Brassica cultivars with different fungal lineages showing different effects on oil seed rape, cabbage or cauliflower.

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