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Plant Promoter Analysis: Identification and Characterization of Root Nodule Specific Promoter in the Common Bean
Published on: December 23, 2017
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Conserved cis-elements enable NODULES WITH ACTIVATED DEFENSE1 regulation by NODULE INCEPTION during nodulation
Haixiang Yu1,2, Aifang Xiao1,3, Zhongmin Zou1
1National Key Laboratory of Agricultural Microbiology, Hubei Hongshan Laboratory, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
The Plant Cell
|August 13, 2024
Summary
Symbiotic nitrogen fixation in nitrogen-fixing clade (NFC) plants involves the NODULES WITH ACTIVATED DEFENSE1 (NAD1) gene, regulated by nodule inception (NIN). This NIN-NAD1 module is crucial for accommodating rhizobia in plant nodules.
Area of Science:
- Plant molecular biology
- Evolutionary biology
- Microbiology
Background:
- Symbiotic nitrogen fixation in nitrogen-fixing clade (NFC) plants is a key agricultural trait.
- The evolutionary origin of nodulation is hypothesized to involve gene gain and loss, but molecular evidence is scarce.
- NODULES WITH ACTIVATED DEFENSE1 (NAD1) is a candidate gene for nodulation.
Purpose of the Study:
- To investigate the evolutionary conservation and function of NAD1 in NFC plants.
- To identify regulatory elements controlling NAD1 expression in nodules.
- To elucidate the role of the nodule inception (NIN) protein in regulating NAD1.
Main Methods:
- Bioinformatic analysis of plant genomes to confirm NAD1 presence and identify cis-regulatory elements.
- Quantitative reverse transcription PCR to analyze NAD1 gene expression in nodules.
- Promoter deletion analysis and site-directed mutagenesis to identify functional nodulation-associated cis-regulatory elements (NACEs).
- Electrophoretic mobility shift assay (EMSA) to confirm NIN binding to NACEs.
- Analysis of CRISPR-Cas9-induced LjNAD1 mutants in Lotus japonicus.
Main Results:
- NAD1 is confirmed as an NFC-specific gene, exclusively expressed in nodules.
- Three conserved nodulation-associated cis-regulatory elements (NACE1-3) were identified in the LjNAD1 promoter.
- NACE1 and NACE2 show specificity to Fabales and Papilionoideae, respectively, while NACE3 is conserved across NFC plants.
- Nodule inception (NIN) directly binds to NACE1-3, activating NAD1 expression.
- LjNAD1 mutants exhibit abnormal symbiosome formation and bacteroid breakdown, similar to Mtnad1 and Mtnin mutants.
Conclusions:
- The NIN-NAD1 regulatory module is essential for rhizobial accommodation in NFC plant nodules.
- NIN-mediated regulation of NAD1 represents a critical evolutionary adaptation for nodulation.
- This study provides molecular evidence supporting the evolutionary model of nodulation in NFC plants.
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