PhD - Engineering nitrogenase in plants

La Caixa INPhINIT Incoming Fellowships are dedicated and offered to early-stage researchers of any nationality to pursue their PhD studies in research centers accredited with the Spanish Seal of Excellence Severo Ochoa, María de Maeztu or Health Institute Carlos III and Portuguese units participating in the program.

• 3 years contract

• Candidates cannot have resided or carried out their main activity (work, studies, etc.) in Spain for more than twelve months in the three years immediately prior to the deadline for application.

• Program includes a training programme on transversal skills aiming in widening the chances for personal development, not only in academia, but also in industry.

• Candidates must be in the first four years of their research career and must not have previously obtained a PhD degree or be in a position to apply for one

• Candidates must accredit an advanced level of English (B2 or higher)

• Competitive salary with additional amount for research costs and a €7500 prize for fellows who deposit the thesis within 6 months after the third year of the fellowship has ended

Research Project / Research Group Description

The energy to synthesize nitrogen-based crop fertilizers accounts for 1.5% of global fossil fuel use, and fertilizer usage releases the powerful greenhouse gas nitrous oxide. The vision of the Rubio laboratory is to engineer cereal plants capable of fixing atmospheric N2 by transferring prokaryotic nitrogenase genes (encoding the protein complex that performs biological N fixation) to the plant genome. This project encompasses synthetic biology and gene circuit design for expression, isolation, and biochemical analysis of O2-sensitive metalloproteins from yeast and plants.

Important milestones have already been reached within this project, e.g. the assembly of extremely O2-sensitive nitrogenase (Nif) proteins and [Fe-S] clusters in yeast mitochondria (doi: 10.1038/ncomms11426, 10.1073/pnas.1904903116, 10.1111/1751-7915.13758), and the accumulation of Fe protein (the most O2-sensitive component of the nitrogenase) in plant chloroplasts and mitochondria (doi: 10.1111/pbi.13347, 10.1038/s42003-020-01536-6). While the Fe protein isolated from yeast was fully functional, that produced in tobacco and rice showed lower activity, emphasizing the need for further exploration of optimal conditions for nitrogenase in plants.

The focus of future work will be to increase the resilience of the Nif proteins to O2 levels typically found in plant cells, and to identify and engineer metabolic conditions that support N fixation and the delivery and stability of [Fe-S] clusters required for nitrogenase function. Monitored milestones and measurable indicators of success will include: (i) engineering nif gene expression cassettes induced under low O2 conditions; (ii) engineering chloroplast and mitochondria [Fe-S] cluster biosynthesis for nitrogenase; (iii) purification of Nif proteins from plastids and mitochondria of plants with increased [Fe-S] cluster occupation and specific activity.

Job position description

The “Biochemistry of nitrogen fixation” group at the Center for Plant Biotechnology and Genomics (UPM-INIA) opens a position for a PhD candidate within the INPhINIT program. The candidate would join a multidisciplinary and international research group that leads an international consortium to generate N-fixing plants, a grand challenge of plant biotechnology with potential to impact agricultural systems worldwide.

The host group is a world leader in nitrogenase bioengineering with experience in synthetic biology, microbial and plant biotechnology, and metalloprotein biochemistry. We use three eukaryotic model systems: yeast for the early screening and validation of nitrogenase components in aerobically cultured cells, tobacco for nitrogenase component expression and function assessments, and rice as target engineered crop. The candidate should have background in at least one of the areas listed above.

The candidate will obtain training in synthetic biology and gene circuit design, plant transformation, protein expression and purification, and metalloprotein analysis. The candidate will design and generate plants for regulated Nif protein expression in plastids and/or mitochondria. She/he will perform anaerobic Nif protein purifications from engineered plants and determine the protein biochemical properties and functionality using specialized sets of enzymatic and spectroscopic assays.

Despite being part of a highly collaborative and multidisciplinary group, the candidate should be able to work independently, design experiments and analyze results following rigorous scientific standards. Results and conclusions should be transmitted to specialized and non-specialized audiences in English, including weekly laboratory meetings. Documentation of laboratory work will be in digital format. Physical data such as DNA vectors, purified proteins, microbial strains, and seeds will be kept in the laboratory collections.

GROUP LEADER

Prof. Luis Manuel Rubio [email protected]

RESEARCH PRODUCT / RESEARCH GROUP

BIOCHEMISTRY OF NITROGEN FIXATION

http://www.cbgp.upm.es/index.php/en/scientific-information/synbio2/nitrogen-fixation

OTHER RELEVANT WEBSITES

“The Big Biology Breakthroughs of 2017”, the biggest biology stories and most important research of 2017 listed by the Royal Society of Biology (UK)

https://www.rsb.org.uk/biologist-features/158-biologist/features/1881-big-biology-breakthroughs-2017

The Fix is in” by Tracy Staedter. Discover Magazine, October 8, 2019

https://www.discovermagazine.com/environment/the-world-has-a-fertilizer-problem-bioengineered-corn-could-save-us

“The nitrogen fix” by Erik Stokstad. (2016) Science 353: 1225-1227

http://science.sciencemag.org/content/353/6305/1225