Visualizing the Subcellular Distribution of Atmospheric Nitrogen Fixed by Gluconacetobacter diazotrophicus in Maize Plants

Główne pojęcia

Nanoscale secondary ion mass spectrometry (NanoSIMS) imaging reveals the heterogeneous incorporation of atmospheric nitrogen fixed by Gluconacetobacter diazotrophicus bacteria into chloroplasts, the key energy source for plant growth, in maize leaves.

Streszczenie

The study used nanoscale secondary ion mass spectrometry (NanoSIMS) imaging to directly visualize the distribution of atmospheric nitrogen fixed by the nitrogen-fixing bacteria Gluconacetobacter diazotrophicus (Gd) in maize plants.

Key findings:

• Significant uptake of the fixed 15N nitrogen was observed in the chloroplasts of the maize leaves, with the majority being incorporated heterogeneously into these organelles.
• The cell nuclei and xylem cell walls also showed 15N enrichment, but to a lesser extent compared to the chloroplasts.
• Within individual chloroplasts, the stromal regions containing the Calvin-Benson cycle enzymes like Rubisco had significantly higher 15N incorporation compared to the thylakoid membrane regions.
• Sporadic regions with exceptional 15N enrichment (up to 27,000% above natural levels) were observed, likely corresponding to individual Gd bacteria, suggesting a symbiotic relationship where the plant provides energy from photosynthesis to drive the bacterial nitrogen fixation.

The findings help explain the observed enhanced growth rates in plants inoculated with this nitrogen-fixing bacteria, as the fixed nitrogen is directly incorporated into the chloroplasts, the key energy source driving plant growth. The methodology provides a template for future correlative studies combining genomic and spatial transcriptomic analyses to further understand the mechanisms of nitrogen fixation and transfer in plant-bacteria symbioses.

Statystyki

The nitrogen isotope ratio (15N/14N) in the control sample was 0.0039, which was weakly but statistically significantly different from the theoretical natural ratio of 0.0037.
The chloroplasts showed significantly higher nitrogen isotope ratios compared to the cell nuclei and xylem cell walls.
Within individual chloroplasts, the stromal regions had significantly higher 15N incorporation compared to the thylakoid membrane regions.
Regions with exceptional 15N enrichment (up to 27,000% above natural levels) were observed, likely corresponding to individual Gd bacteria.

Cytaty

"Significant uptake of 15N into chloroplasts (shown by the white arrows in Fig. 1a) are clearly observed in the nitrogen isotope ratio images in Fig. 1e and 1f."
"The chloroplasts had significantly higher (p < .0001) nitrogen isotope ratio values compared to those measured in the nuclei and xylem cell walls, the latter of which were only slightly higher than the natural value."
"These regions show immense enrichment in 15N (∼10,000% to over ∼27,000% increase above natural ratio)."

Kluczowe wnioski z

Subcellular visualization of the distribution of atmospheric dinitrogen fixed by Gluconacetobacter diazotrophicus bacteria in maize

by McMahon,G., ... o www.biorxiv.org 04-30-2024

https://www.biorxiv.org/content/10.1101/2024.04.29.591714v1

Głębsze pytania

How can the insights from this subcellular nitrogen fixation visualization be leveraged to optimize the use of Gluconacetobacter diazotrophicus as a sustainable nitrogen source for crop production?

The subcellular visualization of nitrogen fixation by Gluconacetobacter diazotrophicus (Gd) in maize plants provides valuable insights that can be leveraged to enhance the efficiency and sustainability of using this bacterium as a nitrogen source for crop production. By understanding the distribution of fixed nitrogen within plant cells, particularly the heterogenous incorporation into chloroplasts, researchers and agronomists can optimize the application of Gd to maximize nitrogen uptake by plants. This knowledge can inform targeted inoculation strategies to ensure that Gd colonizes plant tissues effectively, especially chloroplasts, where nitrogen assimilation is crucial for plant growth and development.
Furthermore, the high-resolution spatial mapping of nutrient transfer and assimilation within plant cells can guide the development of precision agriculture techniques tailored to the specific needs of crops. By visualizing how Gd interacts with plant cells at a subcellular level, researchers can design innovative delivery methods, such as nano-scale carriers for Gd inoculants, to enhance the efficiency of nitrogen fixation and uptake by plants. This targeted approach can minimize the environmental impact of synthetic fertilizers while promoting sustainable agricultural practices.
Overall, leveraging the insights from subcellular nitrogen fixation visualization can lead to the development of customized strategies for utilizing Gluconacetobacter diazotrophicus as a sustainable nitrogen source for crop production, ultimately contributing to enhanced crop yields, reduced environmental pollution, and improved agricultural sustainability.

What other plant-microbe symbioses could benefit from similar high-resolution spatial mapping of nutrient transfer and assimilation within plant cells and tissues?

Several other plant-microbe symbioses could benefit from high-resolution spatial mapping of nutrient transfer and assimilation within plant cells and tissues to enhance our understanding of their interactions and optimize agricultural practices. Some examples include:

Rhizobia-Legume Symbiosis: Similar to the Gd-maize interaction, studying the subcellular distribution of nitrogen fixation in legume nodules can provide insights into how rhizobia transfer fixed nitrogen to plant cells. Understanding the spatial dynamics of nutrient transfer within nodules can improve the efficiency of nitrogen fixation and enhance crop productivity.

Mycorrhizal Associations: Mapping the uptake and transport of nutrients, especially phosphorus, in mycorrhizal fungi-plant symbioses can elucidate the mechanisms underlying nutrient exchange between the symbiotic partners. This knowledge can help optimize the use of mycorrhizae in agriculture to improve nutrient uptake and plant growth.

Endophytic Bacteria in Crop Plants: Investigating the subcellular localization of nutrient uptake by endophytic bacteria in various crop plants can shed light on how these microbes contribute to plant nutrition and growth. Understanding the spatial distribution of nutrient assimilation can aid in developing targeted strategies for enhancing nutrient availability to plants.

By applying high-resolution spatial mapping techniques to these plant-microbe symbioses, researchers can uncover novel insights into nutrient transfer mechanisms, symbiotic interactions, and plant-microbe communication, leading to more sustainable and efficient agricultural practices.

Given the exceptional 15N enrichment observed in putative individual Gd bacteria, what are the specific mechanisms and signaling pathways that enable this tight coupling between the plant host and the nitrogen-fixing endophyte?

The exceptional 15N enrichment observed in putative individual Gluconacetobacter diazotrophicus (Gd) bacteria suggests a tight coupling between the plant host and the nitrogen-fixing endophyte, facilitated by specific mechanisms and signaling pathways. Several key mechanisms and signaling pathways contribute to this symbiotic relationship:

Chemotaxis and Colonization: Gd bacteria likely possess chemotactic abilities that enable them to sense and move towards plant roots, facilitating colonization and establishment within plant tissues. Chemotaxis allows Gd to navigate through the rhizosphere and enter plant cells, where they can access nutrients and establish a symbiotic relationship with the host plant.

Quorum Sensing: Quorum sensing mechanisms in Gd bacteria may play a role in coordinating gene expression and metabolic activities during nitrogen fixation. By detecting cell density and responding to signaling molecules, Gd can regulate nitrogenase activity and optimize nitrogen fixation in response to environmental cues and plant signals.

Plant Signaling Pathways: Plants likely release signaling molecules, such as flavonoids and phytohormones, that attract and activate Gd bacteria within root tissues. These plant signaling pathways can modulate gene expression in Gd, promoting nitrogen fixation and establishing a beneficial symbiosis between the plant host and the endophyte.

Nutrient Exchange: The symbiotic relationship between the plant host and Gd involves a complex exchange of nutrients, including fixed nitrogen from Gd to the plant and carbon sources from the plant to Gd. Specific transporters and channels facilitate the transfer of nutrients between the two partners, ensuring a mutually beneficial interaction.

Oxygen Regulation: Gd bacteria possess mechanisms to regulate oxygen levels within plant tissues to protect the nitrogenase enzyme from inhibition. By controlling oxygen concentrations and maintaining a microaerobic environment, Gd can sustain nitrogen fixation activity and support plant growth without oxygen-induced damage to the nitrogenase complex.

Overall, the tight coupling between the plant host and the nitrogen-fixing endophyte, such as Gluconacetobacter diazotrophicus, is orchestrated by a combination of chemotaxis, quorum sensing, plant signaling pathways, nutrient exchange mechanisms, and oxygen regulation strategies. These intricate mechanisms ensure efficient nitrogen fixation, nutrient transfer, and symbiotic interactions, contributing to the growth and development of both the plant and the endophytic bacteria.