Nitrogen-fixation inside legume nodules requires a steady supply of oxygen to support high rates of bacterial respiration. However, the bacterial nitrogenase enzyme that fixes nitrogen is highly sensitive to oxygen. This contradiction, the ‘oxygen paradox of biological nitrogen-fixation’, is addressed by small heme-containing proteins produced by the plant that can buffer cellular O2 levels
On October 29, 2021, the top international journal SCIENCE published a paper entitled “NIN-like Protein transcription factors regulate leghemoglobin genes in legume nodules” by Jeremy Murray's team and colleagues at CEMPS and overseas. This study is the first to identify the molecular mechanism by which the NLP family of transcription factors regulates the expression of leghemoglobin genes in root nodules.
NLPs (NIN-like Proteins) are plant specific transcription factors that can bind the target gene promoters through special nitrate-responsive elements (NREs) to activate the downstream gene expression to participate in the process of plant nitrogen metabolism. Dr. Murray's team found that two members of the NLP family, NLP2 and NIN, have high expression levels in root nodules. When analyzing nodules of the NLP2 mutant, they unexpectedly found that they contained less leghemoglobin and fixed less N.
Further work by the team revealed that NIN and NLP2 activate leghemoglobin gene expression in root nodules by directly binding to two overlapping NREs, which were called ‘double nitrate responsive elements (dNREs) which are conserved in legumes, balancing the oxygen microenvironment necessary for nitrogen fixation. Phylogenetic analysis suggests that dNREs and NLP2 evolved in legumes to help increase the expression levels of leghemoglobins in nodules. Leghemoglobins evolved from non-symbiotic hemoglobins which play an important role in O2 -scavenging in plants which helps them survive in low oxygen (hypoxic) conditions that occur during flooding. The Murray team found that other NLPs activate expression of non-symbiotic hemoglobins through NREs. This suggests NLP-hemoglobin modules with roles in hypoxic survival were recruited to nodulation to address the O2 paradox of biological N-fixation.
Legumes have been around for tens of millions of years, and their importance for agriculture was noted by scholars of the Chou dynasty. As energy costs increase year by year, the production cost of nitrogen fertilizer is also increasing. In addition, overuse of nitrogen fertilizers is harmful to the environment. Therefore, as a source of nitrogen fertilizer, biological nitrogen fixation is of great significance to the development of sustainable of agriculture. This study, which reveals a central regulatory mechanism in legume N-fixation, offers a chance to improve the N-fixation in legumes and provides a theoretical basis to transfer N-fixation in non-legumes plants such as rice and maize. Such efforts can help reduce the use of nitrogen fertilizer in agriculture, help improve global food security, and protect the ecological environment.
This work was mainly done at the CEMPS. Suyu Jiang is the first author and Jeremy Murray is the corresponding author. The research team is also part of the International Joint Unit Centre for CAS-JIC (CEPAMS). Ertao Wang, (CEMPS), Ping Xu, Shanghai Normal University, Pascal Gamas, University of Toulouse, France, and Phillip Poole, University of Oxford, UK also participated in the work. The research was supported by the CAS Project for Young Scientists in Basic Research (YSBR-011), the National Science Foundation of China (3217020272), the Ministry of Science and Technology (2019FA0904703), Strategic Priority Research Program of the Chinese Academy of Sciences (XDB27040209), the National Key R&D Program of China (2016YFA0500500), the Chinese Academy of Science (153D31KYSB20160074), and the Biotechnology and Biological Sciences Research Council (BBSRC; grant nos. BB/L010305/1[David Phillips Fellowship]; BB/J004553/1) to JM, BB/N003608/1 to CSC, and by a grant to SJ from the China Postdoctoral Science Foundation (2018M642099). PG, MFJ and YP were supported by ANR grants EPISYM (ANR-15-CE20-0002) and Laboratoire d’Excellence (LABEX) TULIP (ANR-10-LABX-41).
Dr. Jeremy Murray, Group leader
CAS-JIC Center of Excellence for Plant and Microbial Sciences (CEPAMS), CAS Center for Excellence in Molecular Plant Sciences (CEMPS), Chinese Academy of Sciences