Investigating the regulation by O2 of the nif and fix genes in Rhizobia - AfS, ENWW

Lead Research Organisation: University of Oxford
Department Name: Interdisciplinary Bioscience DTP

Abstract

Nif and fix genes are responsible for diazotrophic biological nitrogen fixation (BNF), the ability of specialised bacteria to convert atmospheric N2 into N-containing compounds utilisable by plants. BNF can enable plants to be grown without the application of artificial N containing fertilisers, providing the potential to greatly enhance crop yields in areas without ready fertiliser access. A better understanding of the regulation of these genes will allow the manipulation and possible transfer of BNF to other organisms. Low O2 is a known regulator of some nif and fix genes but not all of them. This work aims to discover the other regulators involved and use a proteomics-based approach to understand the metabolic processes that these regulators control.

Research themes:
Sustainably enhancing agricultural production.
Global food security

Publications

10 25 50

Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/M011224/1 01/10/2015 30/09/2023
1810130 Studentship BB/M011224/1 01/10/2015 30/09/2019 Nick Crang
 
Description It has been determined that the isocitrate dehydrogenase gene icd is solely responsible for the isocitrate dehydrogenase activity seen in Rhizobium leguminosarum bv. viciae 3841and it is essential for growth under all conditions. As a result of this discovery a statistical error in the Hidden Markov Model previously utilised during InSeq screenings by our group has been identified and corrected.

PHB has been demonstrated to be per se non-essential for N2 fixation in Azorhizobium caulinodans ORS571. It does however, play a key role in the regulatory activities of phaR which can severely reduce growth and fixing ability when misregulated.

The theoretical "Malonate Shunt" in Azorhizobium Caulinodans which allowed for generation of acetyl CoA from malate without a pyruvate intermediate, has been shown to be non-existent in the form first proposed by Pauling et. al 2001. Furthermore, any form of malonate semialdehyde dehydrogenase activity has been shown to be functionally irrelevant for growth and fixation on C4 dicarboxylates. Instead the two primary fluxes that enable acetyl CoA production and growth on C4 dicarboxylates are shown to operate via PEP carboxykinase and the malic enzyme pathways
Exploitation Route The PHB findings have been written for publication after a second viewing by the reviewers. This work was important as A. caulinodans was exceptional amongst rhizobia in being unable to fix N2 in the absence of PHB production. Our findings were able to demonstrate that this is not due to a unique flux balancing effect observed only in this species but was entirely due to the misregulation of central carbon metabolism by phaR.

The identification of the malonate shunt as functionally irrelevant has informed flux balance maps previously generated for A. caulinodans, allowing their metabolic models to be refined as has the demonstration that the primary and secondary metabolic fluxes to acetyl coA are via malic enzyme and PEP carboxykinase respectively. It is intended to publish this refined flux model as a paper and we are currently awaiting the refining of the flux map by our collaboration partner.
Sectors Manufacturing, including Industrial Biotechology

 
Description Metabolic bottlenecks in rhizobia affecting competitive nodulation during infection stages
Amount SFr. 103,400 (CHF)
Funding ID P400PB_183901 / 1 
Organisation Swiss National Science Foundation 
Sector Public
Country Switzerland
Start 03/2019 
End 02/2021
 
Description TEM imagining to help characterise PHB mutants 
Organisation James Hutton Institute
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution We produced a mutant strains of azorhizobium caulinodans with novel nitrogen fixing phenotypes. These mutant strains were applied to the host legume Sesbania Rostra and nodules were sent to our partner.
Collaborator Contribution Our collaboration partner performed transmission electron microscopy nodules, allowing us to visualise clearly the phenotype of the bacteroids within
Impact We will be using the images obtained as part of a publication
Start Year 2017
 
Description Development of Poole Lab website (Rhizosphere.org) 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact The Rhizosphere website is used to describe work performed, profiles lab members, lists outreach activities and celebrates achievements. Also includes a list of Lab publications.
Year(s) Of Engagement Activity 2014,2015,2016,2017,2018,2019,2020
URL https://rhizosphere.org
 
Description Plant Sciences Masterclass 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact 30 Secondary school students between 14 and 17 years of age attended a lecture at the Oxford Museum of Natural History on the GM work on tomatoes by Professor Lee Sweetlove from the Oxford Department of Plant Sciences. Students were split into groups of around a dozen and discussed through the benefits of GM, how it differs to traditional agricultural breeding techniques and whether or not the results Professor Sweetlove had achieved could have been reached via conventional means.

I was a discussion facilitator for one of the groups, answering questions, asking leading questions and trying my best to draw them out into the discussion.
Year(s) Of Engagement Activity 2017
 
Description Posters at ICNF, Granada, Spain, Sept. 2017 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact International conference. 11 posters from Lab. 2 short presentations
Year(s) Of Engagement Activity 2017
URL https://rhizosphere.org/lab-news/
 
Description Rothamsted Research invited seminar - Phil Poole 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Professional Practitioners
Results and Impact Phil gave a lunch-time research seminar.
Year(s) Of Engagement Activity 2019
URL https://rhizosphere.org/lab-news/
 
Description Twitter account Rhizosphere @PooleLabOxford 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Twitter account to publicise the work of the Poole Lab
Year(s) Of Engagement Activity 2017,2018,2019,2020
URL https://twitter.com
 
Description World Science Day for Peace and Development 2018 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact As part of my internship at UNESCO Bangkok I organised, in collaboration with the Thai Ministry of Science and Technology as well as the Asian Institute of Technology (AIT), the World Science Day for Peace and Development event. This focused on the theme of plastic pollution and involved four international speakers, taking place across two days at UNESCO Bangkok and AIT. 100 guests from NGOs, the Thai government, universities, schools and industry attended the event, which included Q&A sessions plus poster presentations.
Year(s) Of Engagement Activity 2018
URL https://bangkok.unesco.org/index.php/content/strategic-solutions-deal-global-deluge-plastic-pollutio...