Arabidopsis Research Roundup: April 1st.

This weeks Arabidopsis Research Roundup contains an eclectic mix of investigations. Firstly is a study from Peter Unwin that investigates the molecular factors that control interactions between plants and nematode parasites. Secondly is a study led by John Christie that investigates the factors that control hypocotyl curvature. Thirdly is a fascinating proof-of-concept synbio-style study from Rothamstead Research where an algal gene is transferred into Arabidopsis in the hope of developing a phytomediation-based solution to heavy metal contamination. Fourthly is a study from David Bass that catalogues protist species that feed on leaf-microorganisms whilst finally John Carr heads a study that compares RNA-dependent RNA polymerases from Arabidopsis and Potato.

Eves-van den Akker S, Lilley CJ, Yusup HB, Jones JT, Urwin PE (2016) Functional C-terminally encoded plant peptide (CEP) hormone domains evolved de novo in the plant parasite Rotylenchulus reniformis. Mol Plant Pathol. http://dx.doi.org/10.1111/mpp.12402).CEP1

This study is a collaboration between researchers at the Universities of Dundee and Leeds, led by Peter Unwin. The focus of the paper is the interaction of Plant-Parasitic Nematodes (PPNs) with their plant hosts. PPNs stimulate redifferentiation of vascular tissues to form ‘feeding structures’ that benefit the parasite. This process is mediated by a diverse family of effector proteins termed C-terminally Encoded Peptide plant hormone mimics (CEPs). This study investigates the CEPs from the nematode Rotylenchulus reniformis and suggests that these peptides evolved de novo in this organism. They show that the activity of a synthetic peptide corresponding to RrCEPs causes a reduction in primary root elongation whilst upregulating a set of genes including the nitrate transporter AtNRT2.1. The authors propose that CEPs evolved in R. reniformis to allow sustained biotrophy by upregulating a specific set of feeding-responsive genes and by limiting the size of the feeding site produced. This study represents an exciting introduction to a currently under-researched area within plant-pathogen interactions.

Sullivan S, Hart JE, Rasch P, Walker CH, Christie JM (2016) Phytochrome A Mediates Blue-Light Enhancement of Second-Positive Phototropism in Arabidopsis. Front Plant Sci. 7:290 http://dx.doi.org/10.3389/fpls.2016.00290 Open AccessFrontiersPHOT1

John Christie (Glasgow) is the corresponding author on this investigation into the role of the blue-light receptor phototropin 1 (phot1) during hypocotyl phototropism. Curvature of this organ is enhanced by treatment by red-light mediated by the phytochromeA receptor. However this study shows that pre-treatment with blue-light can also enhance this hypocotyl curvature although this did not occur at higher light intensities. In addition phototropic enhancement was also lacking when PHOT1 is expressed only in the hypocotyl epidermis. Therefore the study shows that the phyA impact on phot1 signaling is restricted to low light intensities and in tissues other than the epidermis.

Zhong Tang, Yanling Lv, Fei Chen, Wenwen Zhang, Barry P. Rosen, and Fang-Jie Zhao (2016) Arsenic Methylation in Arabidopsis thaliana Expressing an Algal Arsenite Methyltransferase Gene Increases Arsenic Phytotoxicity J. Agric. Food Chem. http://dx.doi.org/10.1021/acs.jafc.6b00462 Open Access ArsM

This synthetic biology-focused study is led by Fang-Jie Zhao at Rothamstead Research. The authors take an algal gene (arsM) that allows the transformation of inorganic arsenic to a more volatile methylated version. The biological activity of this enzyme was successfully transferred to two different Arabidopsis ecotypes. However interestingly these transgenic plants became more sensitive to arsenic in growth media suggesting that the new methylated arsenic intermediate is more phytotoxic than inorganic arsenic. Therefore this study demonstrates a negative consequence of this project that attempted to engineer arsenic tolerance in plants. Once again this demonstrates that nature rarely acts predictably and any great ideas usually need to be tested in vivo.

Ploch S, Rose L, Bass D, Bonkowski M (2016) High Diversity Revealed in Leaf Associated Protists (Rhizaria: Cercozoa) of Brassicaceae J Eukaryot Microbiol. http://dx.doi.org/10.1111/jeu.12314

After a fantastic opening line in the abstract, ‘The largest biological surface on earth is formed by plant leaves’, this study includes the work of David Bass from the Natural History Museum in London. They investigate the abundance of protists that associate with leaf-inhabiting microorganisms, the “phyllosphere microbiome“. Their findings demonstrate that protists should be considered an important part of the diversity of plant-interacting microbial organisms.

Hunter LJ, Brockington SF, Murphy AM, Pate AE, Gruden K, MacFarlane SA, Palukaitis P, Carr JP (2016) RNA-dependent RNA polymerase 1 in potato (Solanum tuberosum) and its relationship to other plant RNA-dependent RNA polymerases Sci Rep. 6:23082 http://dx.doi.org/10.1038/srep23082 Open Access

John Carr (Cambridge) is the UK-lead on this collaboration with Slovenian and Korean researchers. They primarily investigate the role of the RDR1 RNA-dependent RNA polymerase (RDRs) in potato. In Arabidopsis the RDR1 gene contributes to basal viral resistance but potato plants deficient in StRDR1 do not show altered susceptibility to three different plant viruses. In addition they perform a phylogenetic analysis on the RDR genes and identify a novel RDR7 gene that is only found in Rosids (but not Arabidopsis.

Arabidopsis Research Roundup: August 27th

The Arabdopsis Research Roundup broadens its remit this week. As well as including three original research papers, which look at casparian strip formation, light and hormone signaling, we also highlight an important viewpoint article that aims to set standards for synthetic biology parts. In addition we include a meeting report from a plant synthetic biology summer school and interviews with plant scientists at the JIC, Caroline Dean and Anne Osbourn.

Kamiya T, Borghi M, Wang P, Danku JM, Kalmbach L, Hosmani PS, Naseer S, Fujiwara T, Geldner N, Salt DE (2015) The MYB36 transcription factor orchestrates Casparian strip formation Proc Natl Acad Sci USA http://dx.doi.org/10.1073/pnas.1507691112 Open Access

GARNet Advisory Board Chairman David Salt (Aberdeen) leads this international collaboration that looks at the (relatively) poorly understood Casparian strip (CS), a lignin-based filter that lies in root endodermal cells. Formation of the CS is initiated by Casparian strip domain proteins (CASPs) that recruit other proteins, which begin the process of lignin deposition. In this study the authors look upstream this process and identify the transcription factor MYB36 that directly regulates expression of CASPs and is essential for CS formation. Ectopic expression of MYB36 in root cortical tissues is sufficient to stimulate expression of CASP1-GFP and subsequent deposit a CS-like structure in the cell wall of cortex cells. These results have implications for the design of future experiments that aim to control how nutrients are taken up by the plant as even though myb36 mutants have a ‘root-defect’, they also have changes to their leaf ionome.

Sadanandom A, Ádám É, Orosa B, Viczián A, Klose C, Zhang C, Josse EM, Kozma-Bognár L, Nagy F (2015) SUMOylation of phytochrome-B negatively regulates light-induced signaling in Arabidopsis thaliana Proc Natl Acad Sci USA http://dx.doi.org/10.1073/pnas.1415260112 Open Access

Ari Sadanandom (Durham) and Ferenc Nagy (Edinburgh) are the leaders of this study that investigates the precise function of the PhyB photoreceptor protein. PhyB interacts with a wide range of downstream signaling partners including the PHYTOCHROME INTERACTING FACTOR (PIF) transcription factors. The small ubiquitin-like modifier (SUMO) peptide is conjugated to larger proteins to bring about a variety of signaling outcomes. In this case the authors find that SUMO is preferentially attached to the C-term of PhyB under red light conditions, a relationship that occurs in a diurnal pattern. SUMOylation of PhyB prevents interaction with PIF5 whilst the OVERLY TOLERANT TO SALT 1 (OTS1) protein likely de-SUMOlyates PhyB in vivo. Altered levels of PhyB SUMOylation cause distinct light-responsive phenotypes and as such this paper adds another level of regulation to the already complex known network that controls light signaling.

Schuster C, Gaillochet C, Lohmann JU (2015) Arabidopsis HECATE genes function in phytohormone control during gynoecium development Development. http://dx.doi.org/10.1242/dev.120444 Open Access

Christopher Schuster who is now a postdoc based at the Sainsbury lab in Cambridge is the lead author on this investigation into the role of the HECATE (HEC) family of bHLH transcription factors on fruit development in Arabidopsis. During this process HEC proteins are involved in the response to both the phytohormones auxin and cytokinin, the authors proposing that HEC1 plays an essential role in Arabidopsis gynoecium formation.

Patron N et al (2015) Standards for plant synthetic biology: a common syntax for exchange of DNA parts New Phytologist http://dx.doi.org/10.1111/nph.13532 Open Access

Carmichael RE, Boyce A, Matthewman C Patron N (2015) An introduction to synthetic biology in plant systems New Phytologist http://dx.doi.org/10.1111/nph.13433 Open Access

Although not strictly based on Arabdopsis work, there are a couple of articles in New Phytologist that have broad relevance to plant scientists who are interested in plant synthetic biology. In the first of these Nicola Patron (The Sainsbury Laboratory) leads a wide consortium that aims to set parameters for the standardisation of parts in plant synthetic biology. It is hoped that as the principles of synbio are used more widley in the plant sciences that the proposals in this paper will serve as a useful guide to standidise part production. GARNet has recently written a blog post on this topic.
SynBioWorkshopPic
The associated meeting report looks at the use of plant synthetic biology in a teaching context with a synopsis of the ERASynBio summer school hosted by John Innes Centre. In this event, young researchers from a range of backgrounds were introduced to the power and potential of plant synthetic biology through a diverse course of lectures, practical session and group projects.

 

Vicente C (2015) An interview with Caroline Dean Development http://dx.doi.org/10.1242/dev.127548 Open Access

An interview with Anne Osbourn (2015) New Phytologist <a href="http://dx.doi acheter cialis.org/10.1111/nph.13616″ onclick=”_gaq.push([‘_trackEvent’, ‘outbound-article’, ‘http://dx.doi.org/10.1111/nph.13616’, ‘http://dx.doi.org/10.1111/nph.13616 ‘]);” target=”_blank”>http://dx.doi.org/10.1111/nph.13616 Open Access

These are interviews with eminent female plant molecular biologists who both work at the John Innes Centre. Caroline Dean’s lab focuses on the epigenetic mechanisms that regulate vernalisation whilst Anne Osbourn is interested in using synthetic biology approaches to engineer metabolic pathways for the production of novel compounds.

Jackie Hunter, BBSRC: “Breakthroughs will happen where disciplines coalesce”

Categories: funding, synthetic biology
Comments: No Comments
Published on: November 12, 2014

Jackie Hunter, Chief Executive of BBSRC, delivered a lunchtime presentation at the University of Warwick’s School of Life Sciences on Monday this week. She gave an overview of BBSRC investments and strategy, and spent the final twenty minutes in discussion with the gathered researchers, who posed questions from the floor.

Supporting bioscience in the UK

BBSRC is the biggest source of plant science funding in the UK. Its charter is to fund research and training in world-class bioscience, deliver social and economic impact, and to promote public dialogue.

Hunter explained that BBSRC responsive mode funding (around £150m per year) aims “to ensure excellence in science, wherever it comes from.” It must be functioning well as the UK is top of citation impact index, and the UKPSF found that UK plant science, mainly funded by BBSRC, is second only to the US in terms of publication impact. Strategic funding, capital and campus capital funding to institutes (£6m, £73m and £30m respectively) is used to maintain skills and output in economically important areas of research at the institutes; though Hunter made it clear that ‘blue sky’ research, funded via responsive mode, is important for impact as it generates both top REF scores and top impact metrics. BBSRC also invests £29M per year in specific initiatives.

When asked for advice about increasing BBSRC funding to the department, Hunter emphasised that funding allocation is based on excellence, so departments should provide an environment where excellence can flourish. She also said, “Interdisciplinarity is important: breakthroughs will happen where disciplines coalesce.”

Training and skills

There are around 2000 PhD students at any one time in the Doctoral Training Partnerships that make up part of the £71M BBSRC investment in Knowledge Exchange, Training and Skills. During the discussion session, someone asked about support later in a researcher’s career and Hunter pointed out that investment in early career fellowships must come at the expense of something else. She suggested that BBSRC may consider the value of studentships versus early career fellowships carefully, and in consultation with the community, over the next few years.

Plant science and Agriculture

Jackie Hunter is on the Agri-tech Leadership Council, which aims to increase UK agricultural exports and the value of the UK agri-tech industry by aligning public and industry funding and building skills and research output in agriculture and agri-technology. She also spoke about future directions in BBSRC’s Agriculture and Food theme: improving the nutritional qualities of plants and biopesticides regulation are both likely to become priority areas of research.

Hunter trailed two documents intended to help make two arguments, both of value to the UK plant research community. The first is an upcoming review on animal and plant health, lead by Defra and with input from BBSRC. To be launched later this month, it will be a starting point for BBSRC and Defra to develop joint strategies in tackling current animal and plant health issues, and to work together to call for more funding in this area. The second is a discussion document about synthetic biology and other new ways of working; Hunter hopes this will help make the case for trait-based, rather than methods-based, regulation of new crops.

On-going activities

Hunter also highlighted a few current initiatives our readers might be interested in.

BBSRC has invested £18m in 13 Networks in Industrial Biotechnology and Bioenergy (NIBBs). Here at GARNet, we’re in touch with the High Value Chemicals from Plants Network about a synthetic biology event next year and I recommend you join (it’s free) if you’re interested in high-value plant products or synthetic biology. The other plant science network is the Lignocellulosic Biorefinery Network.

One of Hunter’s objectives as CEO is to promote dialogue between scientists and a broad audience, and the first step towards engaging with the general public is the Great British Bioscience Festival. It is taking place this Friday, Saturday and Sunday in Bethnal Green, London, and there will be some amazing plant science among the exhibits. Lisa will be visiting the Festival to cover it for the next issue of the GARNish newsletter so stay tuned for her report!

Plant synthetic biology takes centre stage

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Published on: October 27, 2014

On Monday and Tuesday last week I was at the Marriott Heathrow for the Global Engage Synthetic Biology Congress. Plant synthetic biology had a dedicated track, and while this meant I regretted missing some talks in the other sessions, it did enable me to be suitably impressed at the quality of plant synthetic biology research, mostly coming from the UK and Europe, and its exciting range of applications.

Plant synthetic biology at Global Engage

A highlight for me was Matias Zurbriggen’s excellent presentation on using plant signalling pathways to remotely control mammalian cells. His objective is to understand plant pathways by reconstructing them in other systems, and via research on phytochromes he has developed a tool to remotely control gene expression in mammalian cells (1) and a light-controlled switch for plant cells (2).

Birger Lindberg Møller gave an interesting and accessible talk about plant synthetic biology for high value product (HVP) synthesis. Whatever your level of expertise, if you’re interested in this area I recommend you watch this earlier version of his talk.

Continuing the HVP theme were Brian King, Vincent Martin and plenary speaker Jules Beekwilder. They all aim to make HVPs using simple chassis instead of relatively energy-intensive, and often inefficient, plants. (more…)

Goldenbraid 2.0: A Standardised DNA Assembly Framework

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Published on: May 27, 2014

As promised in my posts about the Plant Engine meeting I attended a couple of weeks ago (1, 2), here is Diego Orzaez to explain his GoldenBraid cloning method and online DNA assembly framework. Diego co-leads the Plant Genomics and Biotechnology Lab at the Instituto de Biología Molecular y Celular de Plantas in Valencia, Spain. 

goldenbraid

GOLDENBRAID 2.0: A STANDARDIZED DNA ASSEMBLY FRAMEWORK FOR MULTIGENE ENGINEERING IN PLANTS

Engineering large multigenic constructs for Plant Synthetic Biology, such as complex metabolic pathways or intricate gene networks, requires efficient, flexible DNA synthesis and assembly technologies. Although custom gene synthesis is becoming increasingly affordable, the direct synthesis of large multigenic constructs remains prohibitive for most labs. Moreover, custom gene synthesis gives little room for combinatorial engineering, something that is highly valued by biotechnologists.

An alternative “building” strategy for multigene engineering is Modular Construction, that is, the fabrication of new devices by combination of prefabricated standard modules. Modular DNA Construction brings a number of advantages as speed, versatility, lab autonomy, combinatorial potential and often lower cost. As in any standardized methodology, the more users adopt the standard, the bigger the advantages.

GoldenBraid is a Modular DNA Construction method developed at the Plant Genomics and Biotechnology lab (IBMCP-Spain), especially designed for building exchangeable multigenic constructs for Plant Synthetic Biology. Routinely, 15-20 Kb constructs comprising 4-6 transcriptional units made of dozens of individual pre-fabricated modules (GBparts) can be created in few days. Longer constructs can be assembled with little additional effort.

To facilitate the process of genetic design using GoldenBraid, and to stimulate the exchange of genetic modules among laboratories, we have recently launched GoldenBraid2.0 (GB2.0), a web-based DNA assembly framework available at www.gbcloning.org site. The GB2.0 webpage hosts the public GB2.0 database, an increasingly populated collection of pre-made “GBparts” that conform to the GB standard. An embedded software tool named GBdomesticator provides users with personalized lab protocols for creating their own collection of standard genetic parts. Users can always combine their own parts with those deposited in the public GB2.0 database. Moreover, building new GB2.0 multigenic constructs is highly facilitated by the GB assembler tool, a software package that assists in the design of new multigenic constructs.

We believe that adopting common standards and creating of public repositories of exchangeable genetic parts will speed up progress in Plant Biotechnology. If you are interested in this field, we encourage you to explore the gbcloning.org webpages. The details of GB assembly system are described in the publications listed below, and there are tutorials online.

Comments on how to enhance community efforts towards the development of public repositories of standard DNA parts are most welcome, and can be addressed to goldenbraid@ibmcp.upv.es

 

References

Plant synthetic biology in Europe

Categories: GARNet, synthetic biology
Comments: 1 Comment
Published on: May 20, 2014

Helsinki

On 8-9 May I attended a meeting hosted by PlantEngine to discuss the idea of a synthetic biology repository in Europe. The presentations were varied and interesting, and hopefully the other delegates enjoyed mine (PDF) too,

The meeting was at VTT in Espoo, a city very close to Helsinki. The local host Heiko Rischer gave a brief introduction to VTT, which is a Finnish institute but has bases all over the world. VTT is very separate from the university system, and although fundamental research gets done there it has a big commercial focus and strong links to Finnish industry. For example, VTT developed Arctic cloudberry stem cell technology for cosmetics with the R&D team from Finnish skincare company Lumene Oy.

PlantEngine itself was introduced by lead PI Heribert Warczecha. It is a European network focused on enhancing capacity in plant metabolic engineering by activities like defining target pathways, disseminate new technologies, and setting standards. There are currently over 70 labs in 23 EU countries in the network. They run training schools and workshops, and fund short-term scientific missions – check it out if you’re looking for funding for a short research trip to another lab.

One of PlantEngine’s aims is to explore synthetic biology for engineering plant products, which is the reason for the meeting I attended. (more…)

Golden Gate cloning: Tips and resources

Comments: 2 Comments
Published on: May 14, 2014

goldengate

Last week I was in Helsinki for a plant synthetic biology meeting, and I learned a lot about existing European synbio tools, resources and research. There’s a short Storify of Tweets from the meeting here, and I’ll do a round-up post very soon. But today I’m highlighting a tool presented at the workshop, which was also presented at our SynBio workshop last year and at PlantSci 2014 but still hasn’t really featured on this blog (rather remiss of me, I know).

The Golden Gate cloning and related MoClo systems were presented (PDF) by one of its inventors, Sylvestre Marillonnet, at our synbio workshop last year. Sylvestre has worked with Nicola Patron, Head of Synthetic Biology at The Sainsbury Laboratory, to make a MoClo toolkit and set of parts available on Addgene. The toolkit includes 39 parts encoding promoters and 5′ untranslated regions; antigenic tags; sub-cellular localisation signals; reporter genes; selectable marker genes; terminators; 3′ untranslated regions; a suppressor of silencing; and two linkers.

Unfortunately the paper describing the toolkit is behind a paywall, but I’ve been tipped off as to where to find all the practical information you need:

1. The supplementary data is accessible to anyone, and it is very informative. SD 2 and 4 list modules in the toolkit and parts kit respectively.

2. Nicola’s website, Synbio@TSL, has pages on how Golden Gate cloning works, making modules, and an assembly protocol.

3. Nicola presented the toolkit at PlantSci 2014 in May and her poster gives a good overview of the paper’s content: GG_Plant_Kit_Poster

Nicola has generated many other parts, which are listed on her website. Some of them can be obtained from Addgene, while others have to be requested from her lab. Synbio@TSL also has a nice introduction to synthetic biology, synthetic biology news, links to online resources and synbio centres, and guides to the major genome editing and DNA assembly techniques.

GoldenBraid is another modular cloning technique which has its own web resources and toolkit available. There’s a guest post coming up soon about that one though so no spoilers here!

The Golden Gate Toolkit is published in: Engler C, Youles M, Grüetzner R, Ehnert T-M, Werner S, Jones JDG, Patron N, Marillonnet S. (2014) A Golden Gate Modular Cloning Toolbox for Plants. ACS Synthetic Biology DOI: 10.1021/sb4001504

The MoClo system is published in: Weber E, Engler C, Gruetzner R, Werner S, Marillonnet S (2011) A Modular Cloning System for Standardized Assembly of Multigene Constructs. PLoS ONE 6(2): e16765. doi:10.1371/journal.pone.0016765

The GoldenBraid system was most recently published as: Sarrion-Perdigones A, Vazquez-Vilar M, Palací J, Castelijns B, Forment J, Ziarsolo P, Blanca J, Granell A, Orzaez D. (2013) GoldenBraid 2.0: A Comprehensive DNA Assembly Framework for Plant Synthetic Biology. Plant Physiol. 2013 162: 1618-1631. doi:10.1104/pp.113.217661

Image credit: Nicola Patron

Report launch: Developing Plant Synthetic Biology in the UK

Categories: GARNet, synthetic biology
Comments: No Comments
Published on: March 31, 2014

plant synthetic biology

 

We’ve been working on the meeting report from last year’s An Introduction to Plant Synthetic Biology workshop for months, so we’re delighted that GARNet Chair Professor Jim Murray is going to launch it tomorrow during his talk at PlantSci 2014!

You’ll be able to download the report, Developing Plant Synthetic Biology in the UK: Opportunities and Recommendations tomorrow on the GARNet website, or if you’re at the conference come and see Jim, Lisa or Charis to get a printed version.

To keep up with all the news from PlantSci 2014, follow #PlantSci2014

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