Characterizing the functional impact of Sinorhizobium meliloti bacA allelic variation on symbiosis with legume hosts
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Authors
Huang, Rui
Date
2024-01-04
Type
thesis
Language
eng
Keyword
rhizobia , rhizobia and legume host specificity , NCR peptides , BacA transporter
Alternative Title
Abstract
The symbioses between legume plants and rhizobial bacteria are highly specific and require both partners to be compatible with each other during the process of symbiotic development. A range of host and symbiont properties regulate the specificity of these relationships. In certain clades of legume plants, the rhizobial BacA transporter and legume-expressed NCR peptides are thought to play a role in defining host/symbiont compatibility. The inverted repeat-lacking clade (IRLC) of legumes expresses NCR peptides to initiate and maintain terminal differentiation of rhizobia, a developmental process that leads to mature, nitrogen-fixation bacteroids. The BacA transporter is essential in transporting NCR peptides and protecting the rhizobia from the antimicrobial activity of these peptides. A previous study found that replacing the bacA allele of Sinorhizobium meliloti with the ortholog from Rhizobium leguminosarum allowed symbiosis with the host Melilotus officinalis but not with Medicago sativa. In this thesis, I investigated the mechanisms underlying the symbiotic specificity of bacA alleles. On the host legume side, the nodule transcriptomes of M. sativa and M. officinalis were assembled, and their NCR peptide transcripts were identified. The transcriptomic data revealed an approximately 2-fold greater abundance of transcripts encoding highly cationic NCR peptides (isoelectric point > 9.5), which are known to have antimicrobial properties, in M. sativa vs M. officinalis. I hypothesize that the difference in the abundance of highly cationic NCR peptide contributes to why the R. leguminosarum BacA can support symbiosis with M. officinalis but not M. sativa. In addition, I hypothesize that unlike the S. meliloti BacA, the R. leguminosarum BacA is unable to efficiently transport all of the highly cationic NCR peptides of M. truncatula, resulting in a build-up in the bacterial periplasm, leading to cell death. On the rhizobia side, I combined genetic analysis and computational modelling to better understand the BacA transporter. Site-directed mutagenesis analyses showed that S. meliloti
Q193G and N312G BacA mutants recreated the symbiotic phenotypes of S. meliloti carrying the R. leguminosarum bacA allele when paired with hosts M. sativa and M. officinalis. In addition, I generated mutant libraries of S. meliloti and R. leguminosarum bacA through error-prone PCR that identified amino acid residues important for BacA function. Together with BacA structural models that I generated, these data provide novel insight and hypotheses into the functional differences between S. meliloti and R. leguminosarum BacA and their ability to transport NCR peptides. Collectively, the data presented in this thesis provides a deeper understanding of how BacA and NCR peptides may contribute to host/symbiont compatibility in rhizobium – legume symbioses.
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Attribution-NonCommercial 4.0 International
ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.
Attribution-NonCommercial 4.0 International