Overexpression of miR160 affects root growth and nitrogen-fixing nodule number in Medicago truncatula

2013 ◽  
Vol 40 (12) ◽  
pp. 1208 ◽  
Author(s):  
Pilar Bustos-Sanmamed ◽  
Guohong Mao ◽  
Ying Deng ◽  
Morgane Elouet ◽  
Ghazanfar Abbas Khan ◽  
...  
Keyword(s):  
Root Growth ◽  
Medicago Truncatula ◽  
Expression Profiles ◽  
Nodule Development ◽  
Auxin Response ◽  
Symbiotic Interaction ◽  
Nodule Number ◽  
Race Pcr ◽  
Regulatory Loop

Auxin action is mediated by a complex signalling pathway involving transcription factors of the auxin response factor (ARF) family. In Arabidopsis, microRNA160 (miR160) negatively regulates three ARF genes (ARF10/ARF16/ARF17) and therefore controls several developmental processes, including primary and lateral root growth. Here, we analysed the role of miR160 in root development and nodulation in Medicago truncatula Gaertn. Bioinformatic analyses identified two main mtr-miR160 variants (mtr-miR160abde and mtr-miR160c) and 17 predicted ARF targets. The miR160-dependent cleavage of four predicted targets in roots was confirmed by analysis of parallel analysis of RNA ends (PARE) data and RACE-PCR experiments. Promoter-GUS analyses for mtr-miR160d and mtr-miR160c genes revealed overlapping but distinct expression profiles during root and nodule development. In addition, the early miR160 activation in roots during symbiotic interaction was not observed in mutants of the nodulation signalling or autoregulation pathways. Composite plants that overexpressed mtr-miR160a under two different promoters exhibited distinct defects in root growth and nodulation: the p35S:miR160a construct led to reduced root length associated to a severe disorganisation of the RAM, whereas pCsVMV:miR160a roots showed gravitropism defects and lower nodule numbers. Our results suggest that a regulatory loop involving miR160/ARFs governs root and nodule organogenesis in M. truncatula.

scholarly journals Expression of small GTPases in the roots and nodules of Medicago truncatula cv. Jemalong

2019 ◽  
Vol 78 (1) ◽  
pp. 1-8 ◽  
Author(s):  
Abdul Razaque Memon ◽  
Christiane Katja Schwager ◽  
Karsten Niehaus
Keyword(s):  
Medicago Truncatula ◽  
Binding Proteins ◽  
Polymerase Chain Reaction Analysis ◽  
Small Gtpases ◽  
Infection Process ◽  
Nodule Development ◽  
Gtp Binding Proteins ◽  
Gtp Binding ◽  
Differential Expression Pattern

Abstract In this study we used Medicago truncatula, to identify and analyze the expression of small GTP-binding proteins (Arf1, Arl1, Sar1, Rabs, Rop/Rac) and their interacting partners in the infection process in the roots and nodules. A real-time polymerase chain reaction analysis was carried out and our results showed that Arf1 (AtArfB1c-like), MtSar1, AtRabA1e-like, AtRabC1-like, MsRab11-like and AtRop7-like genes were highly expressed in the nodules of rhizobium inoculated plants compared to the non-inoculated ones. On the contrary, AtRabA3 like, AtRab5c and MsRac1-like genes were highly expressed in non-infected nitrogen supplied roots of M. truncatula. Other Rab genes (AtRabA4a, AtRabA4c and AtRabG3a-like genes) were nearly equally expressed in both treatments. Interestingly, RbohB (a respiratory burst NADPH oxidase homologue) was more highly expressed in rhizobium infected than in non-infected roots and nodules. Our data show a differential expression pattern of small GTP-binding proteins in roots and nodules of the plants. This study demonstrates an important role of small GTP-binding proteins in symbiosome biogenesis and root nodule development in legumes.

scholarly journals MtBZR1 Plays an Important Role in Nodule Development in Medicago truncatula

2019 ◽  
Vol 20 (12) ◽  
pp. 2941
Author(s):  
Can Cui ◽  
Hongfeng Wang ◽  
Limei Hong ◽  
Yiteng Xu ◽  
Yang Zhao ◽  
...  
Keyword(s):  
Medicago Truncatula ◽  
Sinorhizobium Meliloti ◽  
Normal Growth ◽  
Dry Mass ◽  
Growth Conditions ◽  
Nodule Development ◽  
Functional Study ◽  
Loss Of Function ◽  
Wild Type

Brassinosteroid (BR) is an essential hormone in plant growth and development. The BR signaling pathway was extensively studied, in which BRASSINAZOLE RESISTANT 1 (BZR1) functions as a key regulator. Here, we carried out a functional study of the homolog of BZR1 in Medicago truncatula R108, whose expression was induced in nodules upon Sinorhizobium meliloti 1021 inoculation. We identified a loss-of-function mutant mtbzr1-1 and generated 35S:MtBZR1 transgenic lines for further analysis at the genetic level. Both the mutant and the overexpression lines of MtBZR1 showed no obvious phenotypic changes under normal growth conditions. After S. meliloti 1021 inoculation, however, the shoot and root dry mass was reduced in mtbzr1-1 compared with the wild type, caused by partially impaired nodule development. The transcriptomic analysis identified 1319 differentially expressed genes in mtbzr1-1 compared with wild type, many of which are involved in nodule development and secondary metabolite biosynthesis. Our results demonstrate the role of MtBZR1 in nodule development in M. truncatula, shedding light on the potential role of BR in legume–rhizobium symbiosis.

scholarly journals NO Network for Plant–Microbe Communication Underground: A Review

2021 ◽  
Vol 12 ◽  
Author(s):  
Anjali Pande ◽  
Bong-Gyu Mun ◽  
Da-Sol Lee ◽  
Murtaza Khan ◽  
Geun-Mo Lee ◽  
...  
Keyword(s):  
Mycorrhizal Fungi ◽  
Nodule Development ◽  
Symbiotic Interaction ◽  
Plant Microbe Interaction ◽  
Nodule Senescence ◽  
Infection Threads ◽  
Molecular Patterns ◽  
Defense Studies ◽  
Hair Curling

Mechanisms governing plant–microbe interaction in the rhizosphere attracted a lot of investigative attention in the last decade. The rhizosphere is not simply a source of nutrients and support for the plants; it is rather an ecosystem teeming with diverse flora and fauna including different groups of microbes that are useful as well as harmful for the plants. Plant–microbe interaction occurs via a highly complex communication network that involves sophisticated machinery for the recognition of friend and foe at both sides. On the other hand, nitric oxide (NO) is a key, signaling molecule involved in plant development and defense. Studies on legume–rhizobia symbiosis suggest the involvement of NO during recognition, root hair curling, development of infection threads, nodule development, and nodule senescence. A similar role of NO is also suggested in the case of plant interaction with the mycorrhizal fungi. Another, insight into the plant–microbe interaction in the rhizosphere comes from the recognition of pathogen-associated molecular patterns (PAMPs)/microbe-associated molecular patterns (MAMPs) by the host plant and thereby NO-mediated activation of the defense signaling cascade. Thus, NO plays a major role in mediating the communication between plants and microbes in the rhizosphere. Interestingly, reports suggesting the role of silicon in increasing the number of nodules, enhancing nitrogen fixation, and also the combined effect of silicon and NO may indicate a possibility of their interaction in mediating microbial communication underground. However, the exact role of NO in mediating plant–microbe interaction remains elusive. Therefore, understanding the role of NO in underground plant physiology is very important, especially in relation to the plant’s interaction with the rhizospheric microbiome. This will help devise new strategies for protection against phytopathogens and enhancing plant productivity by promoting symbiotic interaction. This review focuses on the role of NO in plant–microbe communication underground.

scholarly journals Genome-Wide Identification and Expression Analysis of the Aux/IAA and Auxin Response Factor Gene Family in Medicago truncatula

2021 ◽  
Vol 22 (19) ◽  
pp. 10494
Author(s):  
Rui Liu ◽  
Zhenfei Guo ◽  
Shaoyun Lu
Keyword(s):  
Medicago Truncatula ◽  
Reference Genome ◽  
Expression Profiles ◽  
Interaction Network ◽  
Auxin Response ◽  
Cis Acting ◽  
Protein Protein Interaction ◽  
Genome Wide ◽  
Auxin Response Factor Gene ◽  
Protein Protein Interaction Network

Aux/IAA and auxin response transcription factor (ARF) genes are key regulators of auxin responses in plants. A total of 25 MtIAA and 40 MtARF genes were identified based on the latest updated Medicago truncatula reference genome sequence. They were clustered into 10 and 8 major groups, respectively. The homologs among M. truncatula, soybean, and Arabidopsis thaliana shared close relationships based on phylogenetic analysis. Gene structure analysis revealed that MtIAA and MtARF genes contained one to four concern motifs and they are localized to eight chromosomes, except chromosome 6 without MtARFs. In addition, some MtIAA and MtARF genes were expressed in all tissues, while others were specifically expressed in specific tissues. Analysis of cis-acting elements in promoter region and expression profiles revealed the potential response of MtIAA and MtARF genes to hormones and abiotic stresses. The prediction protein–protein interaction network showed that some ARF proteins could interact with multiple Aux/IAA proteins, and the reverse is also true. The investigation provides valuable, basic information for further studies on the biological functions of MtIAA and MtARF genes in the regulation of auxin-related pathways in M. truncatula.

Role of microRNAs in the legume - rhizobium nitrogen fixing symbiosis

2019 ◽  
Author(s):  
◽  
Nhung Thi Huyen Hoang
Keyword(s):  
Transcription Factor ◽  
Nitrogen Fixation ◽  
Biological Nitrogen Fixation ◽  
Nodule Development ◽  
Atmospheric Nitrogen ◽  
Ammonia Production ◽  
Nitrogen Fixing ◽  
Nodule Number ◽  
Biological Nitrogen

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT REQUEST OF AUTHOR.] Nitrogen is a macronutrient that is critical for plant growth and development because it provides the building blocks of nucleic acids, proteins, chlorophyll, and energy- transfer compounds, such as ATP. Although 78% of the atmosphere is diatomic nitrogen, this form is inert and unavailable to plants due to the strong nitrogen-nitrogen triple bond. Plants can only absorb nitrogen in the forms of NH4+ or NO3-. Most of the inorganic nitrogen available to crop plants is provided through fertilizers synthesized based on the Haber-Bosch process. This process converts atmospheric nitrogen (N2) into ammonia (NH3) by a reaction with hydrogen (H2) using a metal catalyst (iron) under high temperatures (~500 [degrees]C) and high pressures (150-300 bar). Ammonia production by this method consumes a lot of energy, which is derived from burning fossil fuels. Synthetic ammonia production by the Haber-Bosch process causes losses of biodiversity through eutrophication, soil acidification and global increase in N2O atmospheric concentration, which is the third most significant greenhouse gas. An alternative approach to provide a sustainable nitrogen source to plants without causing such damage to the environment is through biological nitrogen fixation between legume species and Rhizobium bacteria. The symbiotic interaction between legume plants and rhizobia results in the formation of root nodules, specialized organs within which rhizobia convert atmospheric nitrogen into ammonia for plant consumption. In return, the legume host plants provide rhizobia with photosynthate as a carbon source for their growth. The legume - Rhizobium symbiosis is a sophisticated process that requires numerous regulators including the 20-24 nucleotide-long microRNAs which negatively regulate the expression of their target messenger RNAs. In my study, we provide two examples that demonstrate the significant role of microRNAs in the symbiotic interplay between soybean, an important legume crop, and rhizobia. In the first example, our results suggest that gma-miR319i functions as a positive regulator of nodule number during the soybean - Bradyrhizobium symbiosis by targeting the TCP33 transcription factor. Overexpression and CRISPR/cas9-mediated gene mutation of gma-miR319i increased and reduced nodule number after rhizobial inoculation, respectively. gma-miR319i and TCP33 showed an inverse expression pattern in different stages of nodule development. TCP33 modulated nodule development in a gma-miR319i dependent manner. The expression of gma-miR319i and TCP33 was differentially regulated in one soybean mutant line that exhibits a hypernodulation phenotype. In the second example, we further investigated the mechanism by which two identical microRNAs, gma-miR171o and gma-miR171q, function in modulating the spatial and temporal aspects of soybean nodulation. Although sharing the identical mature sequence, gma-miR171o and gma-miR171q genes are divergent and show unique, tissue-specific expression patterns. The expression levels of the two miRNAs are negatively correlated with that of their target genes. Ectopic expression of these miRNAs in transgenic hairy roots resulted in a significant reduction in nodule formation. Both gma-miR171o and gma-miR171q target members of the GRAS transcription factor superfamily, namely GmSCL-6 and GmNSP2. Besides those two above-mentioned examples, we were able to generate and characterize an enhancer trap insertional mutant of the NODULATION SIGNALING PATHWAY 2 (NSP2) gene which is the target gene of Gma-miR171 and also an important regulator of nodulation. Overall, our study shows the importance of microRNAs in the regulation of nitrogen-fixing symbiosis. Our results contribute to efforts to fully understand the molecular mechanisms controlling the legume - Rhizobium interaction. Our ultimate hope is that the information gained through my studies can lead to an increased utilization of biological nitrogen fixation for sustainable agriculture and environment protection.

scholarly journals The Medicago truncatula PIN2 auxin transporter mediates basipetal auxin transport but is not necessary for nodulation

2019 ◽  
Vol 71 (4) ◽  
pp. 1562-1573 ◽  
Author(s):  
Jason L P Ng ◽  
Astrid Welvaert ◽  
Jiangqi Wen ◽  
Rujin Chen ◽  
Ulrike Mathesius
Keyword(s):  
Medicago Truncatula ◽  
Lateral Root ◽  
Auxin Transport ◽  
Nodule Development ◽  
Moderate Increase ◽  
Loss Of Function ◽  
Auxin Response ◽  
Root Numbers ◽  
Auxin Transporter ◽  
Basipetal Auxin Transport

Abstract The development of root nodules leads to an increased auxin response in early nodule primordia, which is mediated by changes in acropetal auxin transport in some legumes. Here, we investigated the role of root basipetal auxin transport during nodulation. Rhizobia inoculation significantly increased basipetal auxin transport in both Medicago truncatula and Lotus japonicus. In M. truncatula, this increase was dependent on functional Nod factor signalling through NFP, NIN, and NSP2, as well as ethylene signalling through SKL. To test whether increased basipetal auxin transport is required for nodulation, we examined a loss-of-function mutant of the M. truncatula PIN2 gene. The Mtpin2 mutant exhibited a reduction in basipetal auxin transport and an agravitropic phenotype. Inoculation of Mtpin2 roots with rhizobia still led to a moderate increase in basipetal auxin transport, but the mutant nodulated normally. No clear differences in auxin response were observed during nodule development. Interestingly, inoculation of wild-type roots increased lateral root numbers, whereas inoculation of Mtpin2 mutants resulted in reduced lateral root numbers compared with uninoculated roots. We conclude that the MtPIN2 auxin transporter is involved in basipetal auxin transport, that its function is not essential for nodulation, but that it plays an important role in the control of lateral root development.

scholarly journals Auxin Response Factor 2 (ARF2), ARF3, and ARF4 Mediate Both Lateral Root and Nitrogen Fixing Nodule Development in Medicago truncatula

2021 ◽  
Vol 12 ◽  
Author(s):  
Cristina Kirolinko ◽  
Karen Hobecker ◽  
Jiangqi Wen ◽  
Kirankumar S. Mysore ◽  
Andreas Niebel ◽  
...  
Keyword(s):  
Medicago Truncatula ◽  
Root Development ◽  
Lateral Root ◽  
Lateral Roots ◽  
Nodule Development ◽  
Nodule Formation ◽  
Mrna Levels ◽  
Nitrogen Fixing ◽  
Auxin Response ◽  
Small Interference Rna

Auxin Response Factors (ARFs) constitute a large family of transcription factors that mediate auxin-regulated developmental programs in plants. ARF2, ARF3, and ARF4 are post-transcriptionally regulated by the microRNA390 (miR390)/trans-acting small interference RNA 3 (TAS3) module through the action of TAS3-derived trans-acting small interfering RNAs (ta-siRNA). We have previously reported that constitutive activation of the miR390/TAS3 pathway promotes elongation of lateral roots but impairs nodule organogenesis and infection by rhizobia during the nitrogen-fixing symbiosis established between Medicago truncatula and its partner Sinorhizobium meliloti. However, the involvement of the targets of the miR390/TAS3 pathway, i.e., MtARF2, MtARF3, MtARF4a, and MtARF4b, in root development and establishment of the nitrogen-fixing symbiosis remained unexplored. Here, promoter:reporter fusions showed that expression of both MtARF3 and MtARF4a was associated with lateral root development; however, only the MtARF4a promoter was active in developing nodules. In addition, up-regulation of MtARF2, MtARF3, and MtARF4a/b in response to rhizobia depends on Nod Factor perception. We provide evidence that simultaneous knockdown of MtARF2, MtARF3, MtARF4a, and MtARF4b or mutation in MtARF4a impaired nodule formation, and reduced initiation and progression of infection events. Silencing of MtARF2, MtARF3, MtARF4a, and MtARF4b altered mRNA levels of the early nodulation gene nodulation signaling pathway 2 (MtNSP2). In addition, roots with reduced levels of MtARF2, MtARF3, MtARF4a, and MtARF4b, as well as arf4a mutant plants exhibited altered root architecture, causing a reduction in primary and lateral root length, but increasing lateral root density. Taken together, our results suggest that these ARF members are common key players of the morphogenetic programs that control root development and the formation of nitrogen-fixing nodules.

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